WO2021043961A1

WO2021043961A1 - Dosing regimen for the treatment of cancer with an anti icos agonistic antibody and chemotherapy

Info

Publication number: WO2021043961A1
Application number: PCT/EP2020/074728
Authority: WO
Inventors: Marc S. BALLAS; Catherine E. Ellis
Original assignee: Glaxosmithkline Intellectual Property Development Limited
Priority date: 2019-09-06
Filing date: 2020-09-04
Publication date: 2021-03-11

Abstract

The present disclosure relates to a method of treating cancer in a human in need thereof, the method comprising administering to the human an agonist ICOS binding protein or antigen binding portion thereof at a dose of about 0.08 mg to about 240 mg and administering to the human a chemotherapeutic agent.

Description

DOSING REGIMEN FOR THE TREATMENT OF CANCER WITH AN ANTI ICOS AGONISTIC ANTIBODY AND CHEMOTHERAPY

FIELD OF THE INVENTION

The present invention relates to a method of treating cancer in a human. In particular, the present invention relates to dosing of combinations of an agonist ICOS binding protein and a chemotherapeutic agent.

BACKGROUND TO THE INVENTION

Effective treatment of hyperproliferative disorders, including cancer, is a continuing goal in the oncology field. Generally, cancer results from the deregulation of the normal processes that control cell division, differentiation and apoptotic cell death and is characterized by the proliferation of malignant cells which have the potential for unlimited growth, local expansion and systemic metastasis. Deregulation of normal processes includes abnormalities in signal transduction pathways and response to factors that differ from those found in normal cells.

Immunotherapies are one approach to treat hyperproliferative disorders. A major hurdle that scientists and clinicians have encountered in the development of various types of cancer immunotherapies has been to break tolerance to self-antigen (cancer) in order to mount a robust antitumor response leading to tumor regression. Unlike traditional development of small and large molecule agents that target the tumor, cancer immunotherapies target cells of the immune system that have the potential to generate a memory pool of effector cells to induce more durable effects and minimize recurrences.

Though there have been many recent advances in the treatment of cancer, there remains a need for more effective and/or enhanced treatment of an individual suffering the effects of cancer. The methods herein that relate to combining therapeutic approaches for enhancing anti-tumor immunity address this need.

SUMMARY OF THE INVENTION

In one aspect, there is provided a method of treating cancer in a human in need thereof, the method comprising administering to the human an agonist ICOS binding protein or antigen binding portion thereof at a dose of about 0.08 mg to about 240 mg and administering to the human a chemotherapeutic agent.

In one aspect, there is provided an agonist ICOS binding protein or antigen binding portion thereof for use in treating cancer, wherein the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg and is administered with a chemotherapeutic agent.

In another aspect, there is provided a combination comprising an agonist ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent for use in treating cancer, wherein the ICOS binding protein or antigen binding portion thereof is to be administered at a dose of about 0.08 mg to about 240 mg.

In one aspect, there is provided use of an agonist ICOS binding protein or antigen binding portion thereof in the manufacture of a medicament for treating cancer, wherein the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg and is administered with a chemotherapeutic agent.

In another aspect, there is provided a pharmaceutical kit comprising about 0.08 mg to about 1000 mg of an ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent.

In one embodiment of the method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of the invention, the agonist ICOS binding protein or antigen binding portion thereof comprises one or more of: CDRH1 as set forth in SEQ ID NO:l; CDRH2 as set forth in SEQ ID NO:2; CDRH3 as set forth in SEQ ID NO:3; CDRL1 as set forth in SEQ ID NO:4; CDRL2 as set forth in SEQ ID NO:5 and/or CDRL3 as set forth in SEQ ID NO:6 or a direct equivalent of each CDR wherein a direct equivalent has no more than two amino acid substitutions in said CDR.

In another embodiment of the method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of the invention, the agonist ICOS binding protein or antigen binding portion thereof comprises a VH domain comprising an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:7 and/or a VL domain comprising an amino acid sequence at least 90% identical to the amino acid sequence as set forth in SEQ ID NO:8 wherein said agonist ICOS binding protein specifically binds to human ICOS.

In a further embodiment of the method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of the invention, the agonist ICOS binding protein is a monoclonal antibody.

In another embodiment of the method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of the invention the agonist ICOS binding protein is a humanized or fully human monoclonal antibody.

In one embodiment of the method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of the invention the agonist ICOS binding protein comprises an hIgG4PE scaffold.

In another embodiment of the method, agonist ICOS binding protein, combination or use of the invention, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg, about 0.24 mg, about 0.8 mg, about 2.4 mg, about 8 mg, about 24 mg, about 80 mg, or about 240 mg.

In a further embodiment, the method, agonist ICOS binding protein, combination or use of the invention, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg. In yet another embodiment of the method, agonist ICOS binding protein, combination or use of the invention, the agonist ICOS binding protein or antigen binding portion thereof is administered once every three weeks or every 6 weeks.

In one embodiment of the method, agonist ICOS binding protein, combination or use of the invention, the agonist ICOS binding protein or antigen binding portion thereof and/or the chemotherapeutic agent is administered via IV infusion.

In one embodiment of the method, agonist ICOS binding protein, combination or use of the invention, the cancer is a solid tumor.

In a further embodiment of the method, agonist ICOS binding protein, combination or use of the invention, the cancer is selected from NSCLC, HNSCC, urothelial cancer, cervical cancer and melanoma.

In one embodiment of the method, agonist ICOS binding protein, combination, use, or kit of the invention, the chemotherapeutic agent is docetaxel, carboplatin, cisplatin, paclitaxel, fluorouracil, pemetrexed, gemcitabine or a combination thereof.

In one embodiment of the method, agonist ICOS binding protein, combination or use of the invention, docetaxel is administered at a dose of about 30 to about 100 mg/m2 or about 75 mg/m2; wherein carboplatin is administered at a dose of about AUC 4-7 mg/ml per min or about AUC 5 mg/ml per min; wherein cisplatin is administered at a dose of about 20 mg/m2 to about 120 mg/m2 or 100 mg/m2; wherein paclitaxel is administered at a dose of about 135 mg/m2 to about 225 mg/m2 or 200 mg/m2; wherein fluorouracil is administered at a dose of about 200 mg/m2 to about 1200 mg/m2 or 1000 mg/m2; wherein pemetrexed is administered at a dose of about 500 mg/m2; or wherein gemcitabine is administered at a dose of about 1000 mg/m2 to about 1250 mg/m2 or 1250 mg/m2.

In another embodiment of the method, agonist ICOS binding protein, combination, use, or kit of the invention, the chemotherapeutic agent is a platinum-based chemotherapy doublet.

In a further embodiment of the method, agonist ICOS binding protein, combination, use, or kit of the invention, the chemotherapeutic agent is a doublet of pemetrexed and carboplatin, paclitaxel and carboplatin or gemcitabine and carboplatin.

In one embodiment of the method, agonist ICOS binding protein, combination, use, or kit of the invention, the chemotherapeutic agent is a doublet of fluorouracil and carboplatin or fluorouracil and cisplatin.

In another embodiment of the method, agonist ICOS binding protein, combination or use of the invention, the agonist ICOS binding protein or antigen binding portion thereof and the chemotherapeutic agent is administered concurrently and/or sequentially.

It is to be understood that the embodiments described herein relate to the method of treatment, the agonist ICOS binding protein or antigen binding portion thereof for use, combination for use, use of the agonist ICOS binding protein or antigen binding portion thereof in the manufacture of a medicament, the composition and the pharmaceutical kit of the invention.

DESCRIPTION OF DRAWINGS/FIGURES

FIGS. 1A-1D show plots showing the effect of anti-mICOS agonist antibody (at 5, 20, 100 and 200 mg/animal) alone and in combination with carboplatin in CT26 murine colon carcinoma model in female BALB/c mice. Carbo = carboplatin; ICOS = anti-mouse ICOS agonist antibody (7E.17G9); Veh = vehicle; Isotype = Rat IgG2b isotype.

FIGS. 2A-2D show plots showing the effect of anti-mICOS agonist antibody (at 5, 20, 100 and 200 mg/animal) alone and in combination with paclitaxel in CT26 murine colon carcinoma model in female BALB/c mice. Carbo = carboplatin; ICOS = anti-mouse ICOS agonist antibody (7E.17G9); Veh = vehicle; Isotype = Rat IgG2b isotype.

FIGS. 3A-3B are plots showing duration of H2L5 IgG4PE monotherapy treatment: individual patient data. FIG. 3A shows monotherapy dose escalation cohort. FIG. 3B shows PK/PD cohort.

FIG. 4A-4D are plots showing PK and receptor occupancy. FIG 4A shows dose-proportional PK from 0.01 mg/kg to 3 mg/kg; FIG. 4B shows peak receptor occupancy corresponding to maximum plasma concentration; similar relationship for CD8+ receptor occupancy (data not shown). FIG. 4C shows CD4+RO with H2L5 IgG4PE 0.3 mg/kg and 1.0 mg/kg monotherapy (Part 1A). FIG. 4D is a plot showing receptor occupancy (RO) H2L5 IgG4PE concentration.

FIGS. 5A-5C show PK/PD and immunofluorescence data characterising immune phenotype of TIL. FIG. 5A shows cytotoxic T cell to Treg ratio across H2L5 IgG4PE concentrations. FIG. 5B shows MultiOmyx™ dose-response curves. FIG 5C. shows ratio of cytotoxic T cell proliferation: Treg proliferation.

FIG. 6 show results from a patient case study, showing a set of scans of Patient 1 (H2L5 IgG4PE monotherapy treatment).

FIG. 7 shows the tumour response of combination treatment with H2L5 IgG4PE and 5-FU/cisplatin or H2L5 IgG4PE and 5-FU/carboplatin. irCR = immune-related Complete Response; irPR = immune- related Partial Response; irSD = immune-related Stable Disease; irPD = immune-related Progressive Disease; NE = Not Evaluable. H2L5 IgG4PE comprises of subjects dosed with either 24 mg or 80 mg. FIG. 8 is a plot showing percentage change from baseline in tumour measurement of combination treatment with H2L5 IgG4PE and 5-FU/cisplatin or H2L5 IgG4PE and 5-FU/carboplatin. * indicates PD- (L)l Experienced Subjects; ® denotes ongoing treatment. H2L5 IgG4PE comprises of subjects dosed with either 24 mg or 80 mg.

FIG. 9 is a plot of most common treatment related adverse events (AE) observed for combination treatment with H2L5 IgG4PE and 5-FU/cisplatin or H2L5 IgG4PE and 5-FU/carboplatin.

FIG. 10 is a schematic showing the study design of docetaxel alone (SoC) and ICOS in combination with docetaxel.

DETAILED DESCRIPTION OF THE INVENTION

DEFINITIONS

Antigen Binding Protein (ABP)" means a protein that binds an antigen, including antibodies or engineered molecules that function in similar ways to antibodies. Such alternative antibody formats include triabody, tetrabody, miniantibody, and a minibody. Also included are alternative scaffolds in which the one or more CDRs of any molecules in accordance with the disclosure can be arranged onto a suitable non-immunoglobulin protein scaffold or skeleton, such as an affibody, a SpA scaffold, an LDL receptor class A domain, an avimer (see, e.g., U.S. Patent Application Publication Nos. 2005/0053973, 2005/0089932, 2005/0164301) or an EGF domain. An ABP also includes antigen binding fragments of such antibodies or other molecules. Further, an ABP may comprise the VH regions of the invention formatted into a full length antibody, a (Fab02 fragment, a Fab fragment, a bi-specific or biparatopic molecule or equivalent thereof (such as scFv, bi- tri- or tetra-bodies, TANDABS etc.), when paired with an appropriate light chain. The ABP may comprise an antibody that is an IgGl, IgG2, IgG3, or IgG4; or IgM; IgA, IgE or IgD or a modified variant thereof. The constant domain of the antibody heavy chain may be selected accordingly. The light chain constant domain may be a kappa or lambda constant domain. The ABP may also be a chimeric antibody of the type described in WO86/01533, which comprises an antigen binding region and a non-immunoglobulin region. The terms "ABP", "antigen binding protein", "binding protein", "antigen binding agent" and "binding agent" are used interchangeably herein. For example, disclosed herein are agonist ICOS binding proteins.

"Antigen binding site" refers to a site on an antigen binding protein that is capable of specifically binding to an antigen, this may be a single variable domain, or it may be paired VH/VL domains as can be found on a standard antibody. Single-chain Fv (scFv) domains can also provide antigen-binding sites.

The term "antibody" is used herein in the broadest sense to refer to molecules comprising an immunoglobulin-like domain (for example IgG, IgM, IgA, IgD or IgE) and includes monoclonal, recombinant, polyclonal, chimeric, human, humanized, multispecific antibodies, including bispecific antibodies, and heteroconjugate antibodies; a single variable domain (e.g. VH, VHH, VL, domain antibody (DAB)), antigen binding antibody fragments, Fab, F(ab02, Fv, disulphide linked Fv, single chain Fv, disulphide-linked scFv, diabodies, TANDABS, etc. and modified versions of any of the foregoing (for a summary of alternative "antibody" formats see, e.g. Holliger and Hudson, Nature Biotechnology, 2005, Vol 23, No. 9, 1126-1136).

A "chimeric antibody" refers to a type of engineered antibody that contains a naturally- occurring variable region (light chain and heavy chains) derived from a donor antibody in association with light and heavy chain constant regions derived from an acceptor antibody.

A "humanized antibody" refers to a type of engineered antibody having its CDRs derived from a non-human donor immunoglobulin, the remaining immunoglobulin-derived parts of the molecule being derived from one or more human immunoglobulin(s). In addition, framework support residues may be altered to preserve binding affinity (see, e.g. Queen et al. Proc. Natl Acad Sci USA, 86:10029- 10032 (1989), Hodgson et al. Bio/Technology, 9:421 (1991)). A suitable human acceptor antibody may be one selected from a conventional database, e.g. the KABAT database, Los Alamos database, and Swiss Protein database, by homology to the nucleotide and amino acid sequences of the donor antibody. A human antibody characterized by a homology to the framework regions of the donor antibody (on an amino acid basis) may be suitable to provide a heavy chain constant region and/or a heavy chain variable framework region for insertion of the donor CDRs. A suitable acceptor antibody capable of donating light chain constant or variable framework regions may be selected in a similar manner. It should be noted that the acceptor antibody heavy and light chains are not required to originate from the same acceptor antibody. The prior art describes several ways of producing such humanized antibodies - see, for example, EP-A-0239400 and EP-A-054951.

The term "fully human antibody" includes antibodies having variable and constant regions (if present) derived from human germline immunoglobulin sequences. The human sequence antibodies of the invention may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g. mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo). Fully human antibodies comprise amino acid sequences encoded only by polynucleotides that are ultimately of human origin or amino acid sequences that are identical to such sequences. As meant herein, antibodies encoded by human immunoglobulin-encoding DNA inserted into a mouse genome produced in a transgenic mouse are fully human antibodies since they are encoded by DNA that is ultimately of human origin. In this situation, human immunoglobulin-encoding DNA can be rearranged (to encode an antibody) within the mouse, and somatic mutations may also occur. Antibodies encoded by originally human DNA that has undergone such changes in a mouse are fully human antibodies as meant herein. The use of such transgenic mice makes it possible to select fully human antibodies against a human antigen. As is understood in the art, fully human antibodies can be made using phage display technology wherein a human DNA library is inserted in phage for generation of antibodies comprising human germline DNA sequence. The term, full, whole or intact antibody, used interchangeably herein, refers to a heterotetra meric glycoprotein with an approximate molecular weight of 150,000 daltons. An intact antibody is composed of two identical heavy chains (HCs) and two identical light chains (LCs) linked by covalent disulphide bonds. This H2L2 structure folds to form three functional domains comprising two antigen-binding fragments, known as 'Fab' fragments, and a 'Fc' crystallisable fragment. The Fab fragment is composed of the variable domain at the amino-terminus, variable heavy (VH) or variable light (VL), and the constant domain at the carboxyl terminus, CHI (heavy) and CL (light). The Fc fragment is composed of two domains formed by dimerization of paired CH2 and CH3 regions. The Fc may elicit effector functions by binding to receptors on immune cells or by binding Clq, the first component of the classical complement pathway. The five classes of antibodies IgM, IgA, IgG, IgE and IgD are defined by distinct heavy chain amino acid sequences which are called m, a, g, e and d respectively, each heavy chain can pair with either a K or l light chain. The majority of antibodies in the serum belong to the IgG class, there are four isotypes of human IgG, IgGl, IgG2, IgG3 and IgG4, the sequences of which differ mainly in their hinge region.

Fully human antibodies can be obtained using a variety of methods, for example using yeast- based libraries or transgenic animals (e.g. mice) which are capable of producing repertoires of human antibodies. Yeast presenting human antibodies on their surface which bind to an antigen of interest can be selected using FACS (Fluorescence-Activated Cell Sorting) based methods or by capture on beads using labelled antigens. Transgenic animals that have been modified to express human immunoglobulin genes can be immunised with an antigen of interest and antigen-specific human antibodies isolated using B-cell sorting techniques. Human antibodies produced using these techniques can then be characterised for desired properties such as affinity, developability and selectivity.

The term "domain" refers to a folded polypeptide structure that retains its tertiary structure independent of the rest of the polypeptide. Generally, domains are responsible for discrete functional properties of polypeptides and in many cases may be added, removed or transferred to other polypeptides without loss of function of the remainder of the protein and/or of the domain.

The term "single variable domain" refers to a folded polypeptide domain comprising sequences characteristic of antibody variable domains. It therefore includes complete antibody variable domains such as VH, VHH and VL and modified antibody variable domains, for example, in which one or more loops have been replaced by sequences which are not characteristic of antibody variable domains, or antibody variable domains which have been truncated or comprise N- or C-terminal extensions, as well as folded fragments of variable domains which retain at least the binding activity and specificity of the full-length domain. A single variable domain is capable of binding an antigen or epitope independently of a different variable region or domain. A "domain antibody" or "DAB" may be considered the same as a "single variable domain". A single variable domain may be a human single variable domain, but also includes single variable domains from other species such as rodent, nurse shark and Camelid VHH DABS. Camelid VHH are immunoglobulin single variable domain polypeptides that are derived from species including camel, llama, alpaca, dromedary, and guanaco, which produce heavy chain antibodies naturally devoid of light chains. Such VHH domains may be humanized according to standard techniques available in the art, and such domains are considered to be "single variable domains". As used herein VH includes camelid VHH domains.

The terms "VH" and "VL" are used herein to refer to the heavy chain variable region and light chain variable region respectively of an antigen binding protein.

"CDRs" are defined as the complementarity determining region amino acid sequences of an antigen binding protein. These are the hypervariable regions of immunoglobulin heavy and light chains. There are three heavy chain and three light chain CDRs (or CDR regions) in the variable portion of an immunoglobulin. Thus, "CDRs" as used herein refers to all three heavy chain CDRs, all three light chain CDRs, all heavy and light chain CDRs, or at least two CDRs.

Throughout this specification, amino acid residues in variable domain sequences and variable domain regions within full length antigen binding sequences, e.g. within an antibody heavy chain sequence or antibody light chain sequence, are numbered according to the Kabat numbering convention. Similarly, the terms "CDR", "CDRL1", "CDRL2", "CDRL3", "CDRH1", "CDRH2", "CDRH3" used in the Examples follow the Kabat numbering convention. For further information, see Kabat et al. Sequences of Proteins of Immunological Interest, 5th Ed., U.S. Department of Health and Human Services, National Institutes of Health (1991).

It will be apparent to those skilled in the art that there are alternative numbering conventions for amino acid residues in variable domain sequences and full length antibody sequences. There are also alternative numbering conventions for CDR sequences, for example those set out in Chothia et al. (1989) Nature 342: 877-883. The structure and protein folding of the antigen binding protein may mean that other residues are considered part of the CDR sequence and would be understood to be so by a skilled person.

Other numbering conventions for CDR sequences available to a skilled person include "AbM" (University of Bath) and "contact" (University College London) methods. The minimum overlapping region using at least two of the Kabat, Chothia, AbM and contact methods can be determined to provide the "minimum binding unit". The minimum binding unit may be a sub-portion of a CDR.

CDRs or minimum binding units may be modified by at least one amino acid substitution, deletion or addition, wherein the variant antigen binding protein substantially retains the biological characteristics of the unmodified protein, such as an antibody comprising SEQ ID NO:7 and SEQ ID NO:8.

CDRs or minimum binding units may be modified by at least one amino acid substitution, deletion or addition, wherein the variant antigen binding protein substantially retains the biological characteristics of the unmodified protein, such as an antibody comprising SEQ ID NO:7 and SEQ ID

NO:8. It will be appreciated that each of CDR HI, H2, H3, LI, L2, L3 may be modified alone or in combination with any other CDR, in any permutation or combination. In one embodiment, a CDR is modified by the substitution, deletion or addition of up to 3 amino acids, for example 1 or 2 amino acids, for example 1 amino acid. Typically, the modification is a substitution, particularly a conservative substitution (referred herein also as a direct equivalent), for example as shown in Table 1 below.

Table 1

The VH or VL (or HC or LC) sequence may be a variant sequence with up to 10 amino acid substitutions, additions or deletions. For example, the variant sequence may have up to 9, 8, 7, 6, 5, 4, 3, 2 or 1 amino acid substitution(s), addition(s) or deletion(s). The sequence variation may exclude one or more or all of the CDRs, for example the CDRs are the same as the VH or VL (or HC or LC) sequence and the variation is in the remaining portion of the VH or VL (or HC or LC) sequence, so that the CDR sequences are fixed and intact. Typically, the variation is a substitution, particularly a conservative substitution, for example as shown in Table 1.

"Percent identity" between a query amino acid sequence and a subject amino acid sequence is the "Identities" value, expressed as a percentage, that is calculated using a suitable algorithm or software, such as BLASTP, FASTA, DNASTAR Lasergene, GeneDoc, Bioedit, EMBOSS needle or EMBOSS infoalign, over the entire length of the query sequence after a pair-wise global sequence alignment has been performed using a suitable algorithm/software such as BLASTP, FASTA, ClustalW, MUSCLE, MAFFT, EMBOSS Needle, T-Coffee, and DNASTAR Lasergene. Importantly, a query amino acid sequence may be described by an amino acid sequence identified in one or more claims herein.

The query sequence may be 100% identical to the subject sequence, or it may include up to a certain integer number of amino acid or nucleotide alterations as compared to the subject sequence such that the % identity is less than 100%. For example, the query sequence is at least 50, 60, 70, 75, 80, 85, 90, 95, 96, 97, 98, or 99% identical to the subject sequence. Such alterations include at least one amino acid deletion, substitution (including conservative and non-conservative substitution), or insertion, and wherein said alterations may occur at the amino- or carboxy-terminal positions of the query sequence or anywhere between those terminal positions, interspersed either individually among the amino acids or nucleotides in the query sequence or in one or more contiguous groups within the query sequence. The % identity may be determined across the entire length of the query sequence, including the CDRs. Alternatively, the % identity may exclude one or more or all of the CDRs, for example all of the CDRs are 100% identical to the subject sequence and the % identity variation is in the remaining portion of the query sequence, e.g. the framework sequence, so that the CDR sequences are fixed and intact.

The variant sequence substantially retains the biological characteristics of the unmodified protein, such as an agonist for ICOS.

An antigen binding fragment may be provided by means of arrangement of one or more CDRs on non-antibody protein scaffolds. "Protein Scaffold" as used herein includes, but is not limited to, an immunoglobulin (Ig) scaffold, for example an IgG scaffold, which may be a four chain or two chain antibody, or which may comprise only the Fc region of an antibody, or which may comprise one or more constant regions from an antibody, which constant regions may be of human or primate origin, or which may be an artificial chimera of human and primate constant regions.

The protein scaffold may be an Ig scaffold, for example an IgG, or IgA scaffold. The IgG scaffold may comprise some or all the domains of an antibody (/^'.e. CHI, CH2, CH3, VH, ML). The antigen binding protein may comprise an IgG scaffold selected from IgGl, IgG2, IgG3, IgG4 or IgG4PE. For example, the scaffold may be IgGl. The scaffold may consist of, or comprise, the Fc region of an antibody, or is a part thereof.

The subclass of an antibody in part determines secondary effector functions, such as complement activation or Fc receptor (FcR) binding and antibody dependent cell cytotoxicity (ADCC) (Huber et al. Nature 229(5284): 419-20 (1971); Brunhouse et al. Mol Immunol 16(11): 907-17 (1979)). In identifying the optimal type of antibody for a particular application, the effector functions of the antibodies can be taken into account. For example, hlgGl antibodies have a relatively long half- life, are very effective at fixing complement, and they bind to both FcyRI and FcyRII. In contrast, human IgG4 antibodies have a shorter half-life, do not fix complement and have a lower affinity for the FcRs. Replacement of serine 228 with a proline (S228P) in the Fc region of IgG4 reduces heterogeneity observed with hIgG4 and extends the serum half-life (Kabat et al. "Sequences of proteins of immunological interest" 5.sup.th Edition (1991); Angal et al. Mol Immunol 30(1): 105-8 (1993)). A second mutation that replaces leucine 235 with a glutamic acid (L235E) eliminates the residual FcR binding and complement binding activities (Alegre et al. J Immunol 148(11): 3461-8 (1992)). The numbering of the hIgG4 amino acids was derived from EU numbering reference: Edelman et al. Proc. Natl. Acad. USA, 63, 78-85 (1969). PMID: 5257969. In one embodiment of the present invention the ICOS antibody is an IgG4 isotype. In one embodiment, the ICOS antibody comprises an IgG4 Fc region comprising the replacement S228P and L235E or a functional variant thereof. Such an antibody may have the designation IgG4PE. In a preferred embodiment, the agonist ICOS binding protein is H2L5 IgG4PE. The term "donor antibody" refers to an antibody that contributes the amino acid sequences of its variable regions, CDRs, or other functional fragments or analogs thereof to a first immunoglobulin partner. The donor, therefore, provides the altered immunoglobulin coding region and resulting expressed altered antibody with the antigenic specificity and neutralising activity characteristic of the donor antibody.

The term "acceptor antibody" refers to an antibody that is heterologous to the donor antibody, which contributes all (or any portion) of the amino acid sequences encoding its heavy and/or light chain framework regions and/or its heavy and/or light chain constant regions to the first immunoglobulin partner. A human antibody may be the acceptor antibody.

Affinity, also referred to as "binding affinity", is the strength of binding at a single interaction site, i.e. of one molecule, e.g. an antigen binding protein of the invention, to another molecule, e.g. its target antigen, at a single binding site. The binding affinity of an antigen binding protein to its target may be determined by equilibrium methods (e.g. enzyme-linked immunoabsorbent assay (ELISA) or radioimmunoassay (RIA)), or kinetics (e.g. BIACORE analysis).

Avidity, also referred to as functional affinity, is the cumulative strength of binding at multiple interaction sites, e.g. the sum total of the strength of binding of two molecules (or more, e.g. in the case of a bispecific or multispecific molecule) to one another at multiple sites, e.g. taking into account the valency of the interaction.

As used herein an "immuno-modulator" or "immuno-modulatory agent" refers to any substance including monoclonal antibodies that affects the immune system. In some embodiments, the immuno-modulator or immuno-modulatory agent upregulates an aspect of the immune system. Immuno-modulators can be used as anti-neoplastic agents for the treatment of cancer. For example, immuno-modulators include, but are not limited to, anti-PD-1 antibodies (e.g. dostarlimab, OPDIVO/nivolumab, KEYTRUDA/pembrolizumab and LIBTAYO/cemiplimab), anti-CTLA-4 antibodies and anti-ICOS antibodies.

As used herein the term "agonist" refers to an antigen binding protein including, but not limited to, an antibody, that is capable of activating the antigen to which it binds to induce a full or partial antigen-mediated response that is above the response measured in the absence of the antigen binding protein. Examples of agonistic responses include but are not limited to transduction of a survival, growth, proliferation, differentiation and/or maturation signal. In one embodiment, the agonist upon contact with a co-signalling receptor causes one or more of the following (1) stimulates or activates the receptor, (2) enhances, increases or promotes, induces or prolongs an activity, function or presence of the receptor and/or (3) enhances, increases, promotes or induces the expression of the receptor. Agonist activity can be measured in vitro by various assays know in the art such as, but not limited to, measurement of cell signalling, cell proliferation, immune cell activation markers, cytokine production. Agonist activity can also be measured in vivo by various assays that measure surrogate end points such as, but not limited to the measurement of T cell proliferation or cytokine production.

As used herein the term "antagonist" refers to an antigen binding protein including, but not limited to, an antibody, that is capable of fully or partially inhibiting the biological activity of the antigen to which it binds for example by fully or partially blocking binding or neutralising activity. In one embodiment, the antagonist upon contact with a co-signalling receptor causes one or more of the following (1) attenuates, blocks or inactivates the receptor and/or blocks activation of a receptor by its natural ligand, (2) reduces, decreases or shortens the activity, function or presence of the receptor and/or (3) reduces, decreases, abrogates the expression of the receptor. Antagonist activity can be measured in vitro by various assays know in the art such as, but not limited to, measurement of an increase or decrease in cell signalling, cell proliferation, immune cell activation markers, cytokine production. Antagonist activity can also be measured in vivo by various assays that measure surrogate end points such as, but not limited to the measurement of T cell proliferation or cytokine production. In one embodiment, the PD-1 binding protein is an antagonist PD-1 binding protein. In one embodiment, the CTLA-4 binding protein is an antagonist CTLA-4 binding protein.

By "isolated" it is intended that the molecule, such as an antigen binding protein or nucleic acid, is removed from the environment in which it may be found in nature. For example, the molecule may be purified away from substances with which it would normally exist in nature. For example, the mass of the molecule in a sample may be 95% of the total mass.

The term "expression vector" as used herein means an isolated nucleic acid, which can be used to introduce a nucleic acid of interest into a cell, such as a eukaryotic cell or prokaryotic cell, or a cell free expression system, where the nucleic acid sequence of interest is expressed as a peptide chain such as a protein. Such expression vectors may be, for example, cosmids, plasmids, viral sequences, transposons, and linear nucleic acids comprising a nucleic acid of interest. Once the expression vector is introduced into a cell or cell free expression system (e.g. reticulocyte lysate) the protein encoded by the nucleic acid of interest is produced by the transcription/translation machinery. Expression vectors within the scope of the disclosure may provide necessary elements for eukaryotic or prokaryotic expression and include viral promoter driven vectors, such as CMV promoter driven vectors, e.g. pcDNA3.1, pCEP4, and their derivatives, Baculovirus expression vectors, Drosophila expression vectors, and expression vectors that are driven by mammalian gene promoters, such as human Ig gene promoters. Other examples include prokaryotic expression vectors, such as T7 promoter driven vectors, e.g. pET41, lactose promoter driven vectors and arabinose gene promoter driven vectors. Those of ordinary skill in the art will recognize many other suitable expression vectors and expression systems.

The term "recombinant host cell" as used herein means a cell that comprises a nucleic acid sequence of interest that was isolated prior to its introduction into the cell. For example, the nucleic acid sequence of interest may be in an expression vector while the cell may be prokaryotic or eukaryotic. Exemplary eukaryotic cells are mammalian cells, such as but not limited to, COS-1, COS- 7, HEK293, BHK21, CHO, BSC-1, HepG2, 653, SP2/0, NS0, 293, HeLa, myeloma, lymphoma cells or any derivative thereof. Most preferably, the eukaryotic cell is a HEK293, NS0, SP2/0, or CHO cell. E. coli is an exemplary prokaryotic cell. A recombinant cell according to the disclosure may be generated by transfection, cell fusion, immortalization, or other procedures well known in the art. A nucleic acid sequence of interest, such as an expression vector, transfected into a cell may be extrachromasomal or stably integrated into the chromosome of the cell.

As used herein, the term "effective dose" means that dose of a drug or pharmaceutical agent that will elicit the biological or medical response of a tissue, system, animal or human that is being sought, for instance, by a researcher or clinician. Furthermore, the term "therapeutically effective dose" means any dose that, as compared to a corresponding subject who has not received such dose, results in improved treatment, healing, prevention, or amelioration of a disease, disorder, or side effect, or a decrease in the rate of advancement of a disease or disorder. The term also includes within its scope doses effective to enhance normal physiological function. Therapeutically effective amounts and treatment regimes are generally determined empirically and may be dependent on factors, such as the age, weight, and health status of the patient and disease or disorder to be treated. Such factors are within the purview of the attending physician.

Ranges provided herein, of any type, include all values within a particular range described and values about an endpoint for a particular range.

The term "therapeutic agents" refers to therepeutic agents of the invention. In one embodiment, the therapeutic agents are agonist ICOS binding proteins, chemotherapeutic agents and immunomdulatory agents. In other embodiments, one or more additional agents may be administered in addition to agonist ICOS binding proteins and chemotherapeutic agents. Examples of one or more additional agents include, but are not limited to additional immunomodulators. Examples of additional immunomodulators such as a PD1 binding protein or antigen binding portion thereof, a PDL-1 binding protein or antigen binding portion thereof, a CTLA-4 binding protein or antigen binding portion thereof.

It will be understood that references to "therapeutic agents" include embodiments where the two therapeutic agents are administered in any temporal order, such as concurrently or sequentially. The terms concurrent and sequential administration of therapeutic agents are well understood in the art. The individual therapeutic agents, and pharmaceutical compositions comprising such therapeutic agents may be administered together or separately. When administered separately, this may occur concurrently or sequentially in any order (by the same or by different routes of administration). Such sequential administration may be close in time or remote in time. The dose of a therapeutic agents or pharmaceutically acceptable salt thereof and the further therapeutically active agent(s) and the relative timings of administration will be selected in order to achieve the desired combined therapeutic effect. The administration of the therapeutic agents may be advantageous over the individual therapeutic agents in that the combination of the therapeutic agents may provide one or more of the following improved properties when compared to the individual administration of a single therapeutic agent alone: i) a greater anticancer effect than the most active single agent, ii) synergistic or highly synergistic anticancer activity, iii) a dosing protocol that provides enhanced anticancer activity with reduced side effect profile, iv) a reduction in the toxic effect profile, v) an increase in the therapeutic window, and/or vi) an increase in the bioavailability of one or both of the therapeutic agents.

In one embodiment, each therapeutic agent is formulated into its own pharmaceutical composition and each of the pharmaceutical compositions are administered to treat cancer. In this embodiment, each of the pharmaceutical compositions may have the same or different carriers, diluents or excipients. For example, in one embodiment, a first pharmaceutical composition contains an agonist ICOS binding protein, a second pharmaceutical composition contains a chemotherapeutic agent, and the first and second pharmaceutical compositions are both administered to treat cancer.

In one embodiment, the combination comprising an agonist ICOS binding protein and chemotherapeutic agent is formulated together into a single pharmaceutical composition and administered to treat cancer. For example, in one embodiment, a single pharmaceutical composition contains both an agonist ICOS binding protein and a chemotherapeutic agent and is administered as a single pharmaceutical composition to treat cancer.

Antigen Binding Proteins and Antibodies that bind ICOS

Agents directed to ICOS in any of the aspects or embodiments of the present invention include a monoclonal antibody (mAb), or antigen binding fragment thereof, which specifically binds to ICOS. In some embodiments, the mAb to ICOS specifically binds to human ICOS. In one embodiment, the agonist ICOS binding protein is a monoclonal antibody or antigen binding fragment thereof. The mAb may be a human antibody, a humanized antibody or a chimeric antibody, and may include a human constant region. The human constant region is selected from the group consisting of IgGl, IgG2, IgG3 and IgG4 constant regions, and in preferred embodiments, the human constant region is an IgGl or IgG4 constant region. The antigen binding fragment may be selected from the group consisting of Fab, Fab'-SH, F(ab')2, scFv and Fv fragments.

As used herein "ICOS" means any Inducible T-cell costimulator protein. Pseudonyms for ICOS (Inducible T-cell COStimulator) include AILIM; CD278; CVIDl, JTT-1 or JTT-2, MGC39850, or 8F4. ICOS is a CD28-superfamily costimulatory molecule that is expressed on activated T cells. The protein encoded by this gene belongs to the CD28 and CTLA-4 cell-surface receptor family. It forms homodimers and plays an important role in cell-cell signaling, immune responses, and regulation of cell proliferation. The amino acid sequence of human ICOS (isoform 2) (Accession No.: UniProtKB - Q9Y6W8-2) is shown below as SEQ ID NO: 11.

The amino acid sequence of human ICOS (isoform 1) (Accession No. : UniProtKB - Q9Y6W8- 1) is shown below as SEQ ID NO: 12.

Activation of ICOS occurs through binding by ICOS-L (B7RP-1/B7-H2). Neither B7-1 nor B7-2 (ligands for CD28 and CTLA4) bind or activate ICOS. However, ICOS-L has been shown to bind weakly to both CD28 and CTLA-4 (Yao etal. "B7-H2 is a costimulatory ligand for CD28 in human", Immunity, 34(5); 729-40 (2011)). Expression of ICOS appears to be restricted to T cells. ICOS expression levels vary between different T cell subsets and on T cell activation status. ICOS expression has been shown on resting TH17, T follicular helper (TFH) and regulatory T (Treg) cells; however, unlike CD28; it is not highly expressed on naive THI and TH2 effector T cell populations (Paulos et al. "The inducible costimulator (ICOS) is critical for the development of human Thl7 cells", Sci Transl Med, 2(55); 55ra78 (2010)). ICOS expression is highly induced on CD4+ and CD8+ effector T cells following activation through TCR engagement (Wakamatsu et al. "Convergent and divergent effects of costimulatory molecules in conventional and regulatory CD4+ T cells", Proc Natl Acad Sci USA, 110(3); 1023-8 (2013)). Co-stimulatory signalling through ICOS receptor only occurs in T cells receiving a concurrent TCR activation signal (Sharpe AH and Freeman GJ. "The B7-CD28 Superfamily", Nat. Rev Immunol, 2(2); 116-26 (2002)). In activated antigen specific T cells, ICOS regulates the production of both THI and TH2 cytokines including IFN-y, TNF-a, IL-10, IL-4, IL-13 and others. ICOS also stimulates effector T cell proliferation, albeit to a lesser extent than CD28 (Sharpe AH and Freeman GJ. "The B7-CD28 Superfamily", Nat. Rev Immunol, 2(2); 116-26 (2002)).

By "agent directed to ICOS" is meant any chemical compound or biological molecule capable of binding to ICOS. In some embodiments, the agent directed to ICOS is an agonist ICOS binding protein or antigen binding portion thereof.

The term "ICOS binding protein" as used herein refers to a protein that binds to ICOS, including an antibody or an antigen binding fragment thereof, or engineered molecules that function in similar ways to antibodies that are capable of binding to ICOS. In one embodiment, the antibody is a monoclonal antibody. In some instances, the ICOS is human ICOS. The term "ICOS binding protein" can be used interchangeably with "ICOS binding protein", "ICOS binding agent", "ICOS antigen binding protein" or "ICOS antigen binding agent". Thus, as is understood in the art, anti-ICOS antibodies and/or ICOS antigen binding proteins would be considered ICOS binding proteins. This definition does not include the natural cognate ligand or receptor. References to ICOS binding proteins, in particular anti-ICOS antibodies, includes antigen binding portions or fragments thereof. As used herein "antigen binding portion" of an ICOS binding protein would include any portion of the ICOS binding protein capable of binding to ICOS, including but not limited to, an antigen binding antibody fragment.

In one embodiment, the agonist ICOS binding proteins of the present invention comprise any one or a combination of the following CDRs:

CDRH1: DYAMH (SEQ ID NO:l)

CDRH2: LISIYSDHTNYNQKFQG (SEQ ID NO:2)

CDRH3: NNYGNYGWYFDV (SEQ ID NO:3)

CDRL1: SASSSVSYMH (SEQ ID NO:4)

CDRL2: DTSKLAS (SEQ ID NO: 5)

CDRL3 : FQGSGYPYT (SEQ ID NO:6)

In one embodiment, the agonist ICOS binding protein comprises a heavy chain variable region CDR1 ("CDRFI1") comprising an amino acid sequence with one or two amino acid variation(s) ("CDR variant") to the amino acid sequence set forth in SEQ ID NO:l.

In one embodiment, the agonist ICOS binding protein comprises a heavy chain variable region CDR2 ("CDRFI2") comprising an amino acid sequence with five or fewer, such as four or fewer, three or fewer, two or fewer, or one amino acid variation(s) ("CDR variant") to the amino acid sequence set forth in SEQ ID NO:2. In a further embodiment, the CDRFI2 comprises an amino acid sequence with one or two amino acid variation(s) to the amino acid sequence set forth in SEQ ID NO:2.

In one embodiment, the agonist ICOS binding protein comprises a heavy chain variable region CDR3 ("CDRFI3") comprising an amino acid sequence with one or two amino acid variation(s) ("CDR variant") to the amino acid sequence set forth in SEQ ID NO:3.

In one embodiment, the agonist ICOS binding protein comprises a light chain variable region CDR1 ("CDRL1") comprising an amino acid sequence with three or fewer, such as one or two amino acid variation(s) ("CDR variant") to the amino acid sequence set forth in SEQ ID NO:4.

In one embodiment, the agonist ICOS binding protein comprises a light chain variable region CDR2 ("CDRL2") comprising an amino acid sequence with one or two amino acid variation(s) ("CDR variant") to the amino acid sequence set forth in SEQ ID NO: 5.

In one embodiment, the agonist ICOS binding protein comprises a light chain variable region CDR3 ("CDRL3") comprising an amino acid sequence with three or fewer, such as one or two amino acid variation(s) ("CDR variant") to the amino acid sequence set forth in SEQ ID NO:6. In one embodiment, the agonist ICOS binding protein comprises a CDRH1 comprising an amino acid sequence with up to one amino acid variation to the amino acid sequence set forth in SEQ ID NO:l; a CDRH2 comprising an amino acid sequence with up to five amino acid variations to the amino acid sequence set forth in SEQ ID NO:2; a CDRH3 comprising an amino acid sequence with up to one amino acid variation to the amino acid sequence set forth in SEQ ID NO:3; a CDRL1 comprising an amino acid sequence with up to three amino acid variations to the amino acid sequence set forth in SEQ ID NO:4; a CDRL2 comprising an amino acid sequence with up to one amino acid variation to the amino acid sequence set forth in SEQ ID NO: 5; and/or a CDRL3 comprising an amino acid sequence with up to three amino acid variations to the amino acid sequence set forth in SEQ ID NO: 6. In one embodiment, the agonist ICOS binding protein binding protein comprises any one or a combination of the CDRs of SEQ ID NO:l, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, and SEQ ID NO:6. In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof comprises one or more of : CDRH1 as set forth in SEQ ID NO:l; CDRH2 as set forth in SEQ ID NO:2; CDRH3 as set forth in SEQ ID NO:3; CDRL1 as set forth in SEQ ID NO:4; CDRL2 as set forth in SEQ ID NO:5 and/or CDRL3 as set forth in SEQ ID NO:6 or a direct equivalent of each CDR wherein a direct equivalent has no more than two amino acid substitutions in said CDR.

In one embodiment of the present invention the agonist ICOS binding protein comprises CDRH1 (SEQ ID NO:l), CDRH2 (SEQ ID NO:2), and CDRH3 (SEQ ID NO:3) in the heavy chain variable region having the amino acid sequence set forth in SEQ ID NO:7. The agonist ICOS binding proteins of the present invention comprising the humanized heavy chain variable region set forth in SEQ ID NO:7 are designated as"H2." In some embodiments, the anti-ICOS antibodies of the present invention comprise a heavy chain variable region having at least 90% sequence identity to SEQ ID NO: 7. Suitably, the agonist ICOS binding proteins of the present invention may comprise a heavy chain variable region having about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to SEQ ID NO:7. In one embodiment, the anti-ICOS antibodies of the present invention comprise a heavy chain variable region having at least 90% sequence identity to SEQ ID NO:7. In one embodiment, the anti-ICOS antibodies of the present invention comprise a heavy chain variable region as set forth in SEQ ID NO:7.

Humanized heavy chain (VH) variable region (H2):

In one embodiment, the agonist ICOS binding protein comprises a heavy chain variable region ("VH") comprising an amino acid sequence with at least about 90%, 91%, 92%, 93%, 94%, 95%,

96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 7. In one embodiment, the VH comprises an amino acid sequence with at least one amino acid variation to the amino acid sequence set forth in SEQ ID NO: 7, such as between 1 and 5, such as between 1 and 3, in particular up to 2 amino acid variations to the amino acid sequence set forth in SEQ ID NO:7.

In one embodiment of the present invention the agonist ICOS binding protein comprises CDRL1 (SEQ ID NO:4), CDRL2 (SEQ ID NO:5), and CDRL3 (SEQ ID NO:6) in the light chain variable region having the amino acid sequence set forth in SEQ ID NO:8. Agonist ICOS binding proteins of the present invention comprising the humanized light chain variable region set forth in SEQ ID NO:8 are designated as "L5." Thus, an agonist ICOS binding protein of the present invention comprising the heavy chain variable region of SEQ ID NO:7 and the light chain variable region of SEQ ID NO:8 can be designated as H2L5 herein.

In some embodiments, the agonist ICOS binding proteins of the present invention comprise a light chain variable region having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO:8. Suitably, the agonist ICOS binding proteins of the present invention may comprise a light chain variable region having about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to SEQ ID NO:8.

Humanized light chain (VL) variable region (L5):

EIVLTOSPATLSLSPGERATLSCSASSSVSYMHWYOOKPGOAPRLLIYDTSKLASGIPARFSGSGSGTDYTLTISS LEPEDFAVYYCFOGSGYPYTFGOGTKLEIK (SEQ ID NO:8; underlined amino acid residues correspond to the positions of CDRs).

In one embodiment, the agonist ICOS binding protein comprises a light chain variable region ("VL") comprising an amino acid sequence with at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO:8. In one embodiment, the VL comprises an amino acid sequence with at least one amino acid variation to the amino acid sequence set forth in SEQ ID NO: 8, such as between 1 and 5, such as between 1 and 3, in particular up to 2 amino acid variations to the amino acid sequence set forth in SEQ ID NO:8. In one embodiment, the anti-ICOS antibody comprises a light chain variable region having at least 90% sequence identity to SEQ ID NO: 8. In one embodiment, the anti-ICOS antibody comprises a light chain variable region as set forth in SEQ ID NO:8.

In one embodiment, the agonist ICOS binding protein comprises a VH domain comprising an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:7 and/or a VL domain comprising an amino acid sequence at least 90% identical to the amino acid sequence as set forth in SEQ ID NO:8. In one embodiment, the agonist ICOS binding protein comprises a VH domain comprising an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:7 and/or a VL domain comprising an amino acid sequence at least 90% identical to the amino acid sequence as set forth in SEQ ID NO:8, wherein the agonist ICOS binding protein specifically binds to human ICOS. In one embodiment, the agonist ICOS binding protein comprises a VH with the amino acid sequence set forth in SEQ ID NO:7; and a VL with the amino acid sequence set forth in SEQ ID NO:8.

In one embodiment, the agonist ICOS binding protein comprises a VH comprising an amino acid sequence of SEQ ID NO:7 and a VL comprising an amino acid sequence of SEQ ID NO:8. In one embodiment, the agonist ICOS binding protein specifically binds to human ICOS.

In one embodiment, the agonist ICOS binding protein comprises a VH comprising an amino acid sequence with at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO:7; and a VL comprising an amino acid sequence with at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO:8.

In one embodiment, the agonist ICOS binding protein comprises a VH domain comprising an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:7 and/or a VL domain comprising an amino acid sequence at least 90% identical to the amino acid sequence as set forth in SEQ ID NO: 8, and further comprises any one or a combination of the CDRs of SEQ ID NO:l, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO: 5, and SEQ ID NO:6.

In one embodiment, the agonist ICOS binding protein is a humanized monoclonal antibody comprising a heavy chain (HC) amino acid sequence having at least 90%, 91%, 92,%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO:9.

In one embodiment, the HC comprises an amino acid sequence with at least one amino acid variation to the amino acid sequence set forth in SEQ ID NO:9, such as between 1 and 10, such as between 1 and 7, in particular up to 6 amino acid variations to the amino acid sequence set forth in SEQ ID NO:9. In a further embodiment, the HC comprises one, two, three, four, five, six or seven amino acid variations to the amino acid sequence set forth in SEQ ID NO:9.

In one embodiment, the agonist ICOS binding protein is a humanized monoclonal antibody comprising a light chain (LC) amino acid sequence having at least 90%, 91%, 92,%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 10.

In one embodiment, the LC comprises an amino acid sequence with at least one amino acid variation to the amino acid sequence set forth in SEQ ID NO: 10, such as between 1 and 10, such as between 1 and 5, in particular up to 3 amino acid variations to the amino acid sequence set forth in SEQ ID NO: 10. In a further embodiment, the LC comprises one, two or three amino acid variations to the amino acid sequence set forth in SEQ ID NO: 10.

In one embodiment, the agonist ICOS binding protein comprises a HC comprising an amino acid sequence with at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO:9; and a LC comprising an amino acid sequence with at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 10. Therefore, the antibody is an antibody with a heavy chain at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO:9 and/or with a light chain at least about 90% identical to the light chain amino acid sequence of SEQ ID NO: 10.

In one embodiment, the agonist ICOS binding protein comprises a heavy chain amino acid sequence at least about 90% identical to the amino acid sequence of SEQ ID NO:9 and/or a light chain amino acid sequence at least about 90% identical to the amino acid sequence of SEQ ID NO: 10.

In one embodiment, the agonist ICOS binding protein comprises a heavy chain sequence of SEQ ID NO:9 and a light chain sequence of SEQ ID NO: 10.

In one embodiment there is provided an agonist ICOS binding protein comprising a heavy chain constant region that has reduced ADCC and/or complement activation or effector functionality as compared to IgGl. In one embodiment, the IgGl is wild type (WT) IgGl. In one such embodiment the heavy chain constant region may comprise a naturally disabled constant region of IgG2 or IgG4 isotype or a mutated or disabled IgGl constant region.

In one embodiment, the agonist ICOS binding protein comprises an IgG4 Fc region comprising the amino acid substitutions S228P and L235E or functional equivalents thereof. In one embodiment, the agonist ICOS binding protein comprises an IgG4 Fc region comprising amino acid substitutions S229P and L236E. Such embodiments may have the designation IgG4PE. Thus, an agonist ICOS binding protein having the heavy chain variable region H2 and the light chain variable region L5 and an IgG4PE Fc region will be designated as H2L5 IgG4PE or synonymously as H2L5 hIgG4PE.

Antibodies to ICOS and methods of using in the treatment of disease are described, for instance, in W02012131004, US20110243929, and US20160215059. US20160215059 is incorporated by reference herein. CDRs for murine antibodies to human ICOS having agonist activity are shown in PCT/EP2012/055735 (W02012131004). Antibodies to ICOS are also disclosed in WO2008137915, W02010056804, EP1374902, EP1374901, and EP1125585. Agonist antibodies to ICOS or ICOS binding proteins are disclosed in W02012/13004, WO2014033327, WO2016120789, US20160215059, and US20160304610. Exemplary antibodies in US20160304610 include 37A10S713. Sequences of 37A10S713 are reproduced below as SEQ ID NOS: 13-20.

37A10S713 VH CDRl: GFTFSDYWMD (SEQ ID NO: 13)

37A10S713 VH CDR2: NIDEDGSITEYSPFVKG (SEQ ID NO: 14)

37A10S713 VH CDR3: WGRFGFDS (SEQ ID NO: 15)

37A10S713 VL CDRl: KSSQSLLSGSFNYLT (SEQ ID NO: 16)

37A10S713 VL CDR2: YASTRHT (SEQ ID NO:17)

37A10S713 VL CDR3: HHHYNAPPT (SEQ ID NO: 18)

37A10S713 heavy chain variable region:

EVOLVESGGLVOPGGSLRLSCAASGFTFSDYWMDWVRQAPGKGLVWVSNIDEDGSITEYSPFVKGRFTISRDN AKNTLYLOM NSLRAEDTAVYYCTRWG RFG FDSWGOGTLVTVSS (SEQ ID NO: 19; underlined amino acid residues correspond to the positions of CDRs)

37A10S713 light chain variable region:

DIVMTOSPDSLAVSLGERATINCKSSOSLLSGSFNYLTWYOOKPGOPPKLLIFYASTRHTGVPDRFSGSGSGTDF TLTISSLOAEDVAVYYCHHHYNAPPTFGPGTKVDIK (SEQ ID NO:20; underlined amino acid residues correspond to the positions of CDRs)

In an embodiment, the agonist ICOS binding protein is vopratelimab. In one embodiment, the agonist ICOS binding protein is JTX-2011.

Exemplary antibodies in US2018/0289790 include ICOS.33 IgGlf S267E. Sequences of ICOS.33 IgGlf S267E are reproduced below as SEQ ID NOS:21-22.

ICOS.33 IgGlf S267E light chain variable domain:

In one embodiment, the agonist ICOS binding protein is BMS-986226.

Exemplary antibodies in WO2018/029474 include STIM003. Sequences of STIM003 are reproduced below as SEQ ID NOS: 23-24.

STIM003 heavy chain variable domain:

STIM003 light chain variable domain:

In one embodiment, the agonist ICOS binding protein is KY1044.

Exemplary antibodies in W02018/045110 include XENP23104. Sequences of the ICOS binding Fab side ([ICOS]_H0.66_L0) of XENP23104 are reproduced below as SEQ ID NOS:25-32.

XENP23104 [ICOS]_H0.66_L0 heavy chain variable domain:

underlined amino acid residues correspond to the positions of CDRs).

correspond to the positions of CDRs).

As used herein "ICOS-L" and "ICOS Ligand" are used interchangeably and refer to the membrane bound natural ligand of human ICOS. ICOS ligand is a protein that in humans is encoded by the ICOSLG gene. ICOSLG has also been designated as CD275 (cluster of differentiation 275). Pseudonyms for ICOS-L include B7RP-1 and B7-H2.

Chemotherapeutic agent

Chemotherapeutic agents, also referred to as anti-neoplastic agents, are used to directly or indirectly inhibit the proliferation of rapidly growing cells, typically in the context of malignancy. Examples of such agents can be found in Cancer Principles and Practice of Oncology by V.T. Devita and S. Heilman (editors), 6^th edition (February 15, 2001), Lippincott Williams & Wilkins Publishers. Typical anti-neoplastic agents useful in the present invention include, but are not limited to, anti- microtubule agents such as diterpenoids and vinca alkaloids; platinum coordination complexes; alkylating agents such as nitrogen mustards, oxazaphosphorines, alkylsulfonates, nitrosoureas, and triazenes; antibiotic agents such as anthracyclins, actinomycins and bleomycins; topoisomerase II inhibitors such as epipodophyllotoxins; antimetabolites such as purine and pyrimidine analogues and anti-folate compounds; topoisomerase I inhibitors such as camptothecins; non-receptor tyrosine kinase angiogenesis inhibitors; proapoptotic agents; and cell cycle signaling inhibitors.

Examples of chemotherapeutic agents for use in combination or co-administered with the present agonist ICOS binding protein or antigen binding fragment thereof are detailed below.

Anti-microtubule or anti-mitotic agents are phase specific agents active against the microtubules of tumor cells during M or the mitosis phase of the cell cycle. Examples of anti- microtubule agents include, but are not limited to, diterpenoids and vinca alkaloids.

Diterpenoids, which are derived from natural sources, are phase specific anti -cancer agents that operate at the G2/M phases of the cell cycle. It is believed that the diterpenoids stabilize the b- tubulin subunit of the microtubules, by binding with this protein. Disassembly of the protein appears then to be inhibited with mitosis being arrested and cell death following. Examples of diterpenoids include, but are not limited to, paclitaxel and its analog docetaxel. In one embodiment, the chemotherapeutic agent is paclitaxel or docetaxel.

Paclitaxel, 5b,20-epoxy-l,2a,4,7p,10p,13a-hexa-hydroxytax-ll-en-9-one 4,10-diacetate 2- benzoate 13-ester with (2R,3S)-N-benzoyl-3-phenylisoserine; is a natural diterpene product isolated from the Pacific yew tree Taxus brevifolia and is commercially available as an injectable solution TAXOL®. It is a member of the taxane family of terpenes. It was first isolated in 1971 by Wani et al. (J. Am. Chem, Soc., 93:2325. 1971), who characterized its structure by chemical and X-ray crystallographic methods. One mechanism for its activity relates to paclitaxel's capacity to bind tubulin, thereby inhibiting cancer cell growth. Schiff et al., Proc. Natl, Acad, Sci. USA, 77:1561-1565 (1980); Schiff et al., Nature, 277:665-667 (1979); Kumar, J. Biol, Chem, 256: 10435-10441 (1981). For a review of synthesis and anticancer activity of some paclitaxel derivatives see: D. G. I. Kingston eta/., Studies in Organic Chemistry vol. 26, entitled "New trends in Natural Products Chemistry 1986", Attaur-Rahman, P.W. Le Quesne, Eds. (Elsevier, Amsterdam, 1986) pp 219-235.

Paclitaxel has been approved for clinical use in the treatment of refractory ovarian cancer in the United States (Markman et al., Yale Journal of Biology and Medicine, 64:583, 1991; McGuire et al., Ann. Intern, Med., 111:273,1989) and for the treatment of breast cancer (Holmes et al., J. Nat. Cancer Inst., 83:1797,1991.) It is a potential candidate for treatment of neoplasms in the skin (Einzig et. al., Proc. Am. Soc. Clin. Oncol., 20:46) and head and neck carcinomas (Forastire et. al., Sem. Oncol., 20:56, 1990). The compound also shows potential for the treatment of polycystic kidney disease (Woo et. al., Nature, 368:750. 1994, lung cancer and malaria. Treatment of patients with paclitaxel results in bone marrow suppression (multiple cell lineages, Ignoff, R.J. et. al, Cancer Chemotherapy Pocket Guide* 1998) related to the duration of dosing above a threshold concentration (50nM) (Kearns, C.M. et. al., Seminars in Oncology, 3(6) p.16-23, 1995). In one embodiment, the chemotherapeutic agent is paclitaxel. In another embodiment, paclitaxel is administered once every three weeks. In one embodiment, paclitaxel is administered by intravenous (IV) infusion. In one embodiment, paclitaxel is administered at a dose of 135 mg/m² or 175 mg/m². In one embodiment, paclitaxel is administered at a dose of 175 mg/m² every three weeks. In one embodiment, paclitaxel is administered at a dose of 175 mg/m² over 3 hours every three weeks. In one embodiment, paclitaxel is administered at a dose of 135 mg/m² once every three weeks. In one embodiment, paclitaxel is administered at a dose of 135 mg/m² over 24 hours, once every three weeks.

Docetaxel, (2R,3S)- N-carboxy-3-phenylisoserine,N-tert-butyl ester, 13-ester with 5b-20- epoxy-l,2a,4,7b,10b,13a-hexahydroxytax-ll-en-9-one 4-acetate 2-benzoate, trihydrate; is commercially available as an injectable solution as TAXOTERE®. Docetaxel is indicated for the treatment of breast cancer. Docetaxel is a semisynthetic derivative of paclitaxel q.v., prepared using a natural precursor, 10-deacetyl-baccatin III, extracted from the needle of the European Yew tree. The dose limiting toxicity of docetaxel is neutropenia. In one embodiment, the chemotherapeutic agent is docetaxel. In one embodiment, docetaxel is administered by IV infusion. In another embodiment, docetaxel is administered at a dose of 75 mg/m². In one embodiment, docetaxel is administered at a dose of 75 mg/m² over 1 hour. In another embodiment, docetaxel is administered once every three weeks. In another embodiment, docetaxel is administered at a dose of 60 to 100 mg/m² via IV over 1 hour every three weeks.

Vinca alkaloids are phase specific anti-neoplastic agents derived from the periwinkle plant. Vinca alkaloids act at the M phase (mitosis) of the cell cycle by binding specifically to tubulin. Consequently, the bound tubulin molecule is unable to polymerize into microtubules. Mitosis is believed to be arrested in metaphase with cell death following. Examples of vinca alkaloids include, but are not limited to, vinblastine, vincristine, and vinorelbine.

Vinblastine, vincaleukoblastine sulfate, is commercially available as VELBAN® as an injectable solution. Although, it has possible indication as a second line therapy of various solid tumors, it is primarily indicated in the treatment of testicular cancer and various lymphomas including Hodgkin's Disease; and lymphocytic and histiocytic lymphomas. Myelosuppression is the dose limiting side effect of vinblastine.

Vincristine, vincaleukoblastine, 22-oxo-, sulfate, is commercially available as ONCOVIN® as an injectable solution. Vincristine is indicated for the treatment of acute leukemias and has also found use in treatment regimens for Hodgkin's and non-Hodgkin's malignant lymphomas. Alopecia and neurologic effects are the most common side effect of vincristine and to a lesser extent myelosupression and gastrointestinal mucositis effects occur. In one embodiment, vincristine is administered at a dose of 1.4 mg/m². In one embodiment, vincristine is administered once a week.

Vinorelbine, 3',4'-didehydro-4'-deoxy-C'-norvincaleukoblastine [R-(R*,R*)-2,3- dihydroxybutanedioate (l:2)(salt)], commercially available as an injectable solution of vinorelbine tartrate (NAVELBINE®), is a semisynthetic vinca alkaloID Vinorelbine is indicated as a single agent or in combination with other chemotherapeutic agents, such as cisplatin, in the treatment of various solid tumors, particularly non-small cell lung, advanced breast, and hormone refractory prostate cancers. Myelosuppression is the most common dose limiting side effect of vinorelbine. In one embodiment, vinorelbine is administered at a dose of 30 mg/m² once a week.

Platinum coordination complexes are non-phase specific anti-cancer agents, which are interactive with DNA. The platinum complexes enter tumor cells, undergo, aquation and form intra- and interstrand crosslinks with DNA causing adverse biological effects to the tumor. Examples of platinum coordination complexes include, but are not limited to, cisplatin and carboplatin. In one embodiment, the chemotherapeutic agent is a platinum-based chemotherapy doublet. In one embodiment, the platinum-based chemotherapy doublet is 5-FU/carboplatin. In one embodiment, the platinum-based chemotherapy doublet is 5-FU/cisplatin. In another embodiment, the chemotherapeutic agent is selected from one or more of the following: pemetrexed, gemcitabine, fluorouracil (5-FU) paclitaxel, docetaxel, cisplatin or carboplatin. In one embodiment, the chemotherapeutic agent is pemetrexed/carboplatin or pemetrexed/cisplatin doublet. In one embodiment, the chemotherapeutic agent is paclitaxel/carboplatin or paclitaxel/cisplatin doublet. In one embodiment, the chemotherapeutic agent is gemcitabine/carboplatin or gemcitabine cisplatin doublet. In another embodiment, the chemotherapeutic agent is selected from one or more of the following: paclitaxel, docetaxel, or carboplatin.

Cisplatin, cis-diamminedichloroplatinum, is commercially available as PLATINOL® as an injectable solution. Cisplatin is primarily indicated in the treatment of metastatic testicular and ovarian cancer and advanced bladder cancer. The primary dose limiting side effects of cisplatin are nephrotoxicity, which may be controlled by hydration and diuresis, and ototoxicity. In one embodiment, the chemotherapeutic agent is cisplatin. In one embodiment, cisplatin is administered at a dose of 100 mg/m² once every four weeks. In one embodiment, cisplatin is administered at a dose of about 75 mg/m² to about 100 mg/m² once every four weeks. In one embodiment, cisplatin is administered at a dose of about 50 mg/m² to about 70 mg/m² once every three to four weeks.

Carboplatin, platinum, diammine [l,l-cyclobutane-dicarboxylate(2-)-0,0'], is commercially available as PARAPLATIN® as an injectable solution. Carboplatin is primarily indicated in the first and second line treatment of advanced ovarian carcinoma. Bone marrow suppression is the dose limiting toxicity of carboplatin. In one embodiment, the chemotherapeutic agent is carboplatin. In one embodiment, carboplatin is administered at a dose of about 360 mg/m² every four weeks. In one embodiment, carboplatin is administered at a dose of about 300 mg/m² every four weeks.

Alkylating agents are non-phase anti-cancer specific agents and strong electrophiles. Typically, alkylating agents form covalent linkages, by alkylation, to DNA through nucleophilic moieties of the DNA molecule such as phosphate, amino, sulfhydryl, hydroxyl, carboxyl, and imidazole groups. Such alkylation disrupts nucleic acid function leading to cell death. Examples of alkylating agents include, but are not limited to, nitrogen mustards such as cyclophosphamide, melphalan, and chlorambucil; alkyl sulfonates such as busulfan; nitrosoureas such as carmustine; and triazenes such as dacarbazine.

Cyclophosphamide, 2-[bis(2-chloroethyl)amino]tetrahydro-2H-l,3,2-oxazaphosphorine 2- oxide monohydrate, is commercially available as an injectable solution or tablets as CYTOXAN®. Cyclophosphamide is indicated as a single agent or in combination with other chemotherapeutic agents, in the treatment of malignant lymphomas, multiple myeloma, and leukemias. Alopecia, nausea, vomiting and leukopenia are the most common dose limiting side effects of cyclophosphamide. In one embodiment, the chemotherapeutic agent is cisplatin or carboplatin. In another embodiment, the chemotherapeutic agent is selected from one or more of the following: paclitaxel, docetaxel, cisplatin, carboplatin, or cyclophosphamide. In another embodiment, the chemotherapeutic agent is cyclophosphamide. In one embodiment, the chemotherapeutic agent is docetaxel, pemetrexed, gemcitabine, carboplatin, cisplatin, paclitaxel, fluorouracil or a combination thereof. In one embodiment, the chemotherapeutic agent is a platinum-based chemotherapy doublet. In one embodiment, the chemotherapeutic agent is fluorouracil and carboplatin (5-FU/carboplatin). In one embodiment the chemotherapeutic agent is fluorouracil and cisplatin (5-FU/cisplatin). A description of the chemotherapy and administration is provided in Table 10. In one embodiment, cyclophosphamide is administered at a dose of about 40 mg/m² to about 50 mg/m² every two to five days. In another embodiment, cyclophosphamide is administered at a dose of about 10 mg/kg to about 15 mg/kg every 7 to 10 days or 5 mg/kg 2 times a week. In one embodiment, cyclophosphamide is administered via IV infusion.

Melphalan, 4-[bis(2-chloroethyl)amino]-L-phenylalanine, is commercially available as an injectable solution or tablets as ALKERAN®. Melphalan is indicated for the palliative treatment of multiple myeloma and non-resectable epithelial carcinoma of the ovary. Bone marrow suppression is the most common dose limiting side effect of melphalan. In one embodiment, melphalan is administered at a dose of 2 mg or 6 mg, once daily. In one embodiment, melphalan is administered at a dose of 0.2 mg/kg daily.

Chlorambucil, 4-[bis(2-chloroethyl)amino]benzenebutanoic acid, is commercially available as LEUKERAN® tablets. Chlorambucil is indicated for the palliative treatment of chronic lymphatic leukemia, and malignant lymphomas such as lymphosarcoma, giant follicular lymphoma, and Hodgkin's disease. Bone marrow suppression is the most common dose limiting side effect of chlorambucil. In one embodiment, chlorambucil is administered at a dose of 0.1 to 0.2 mg/kg daily or 4 to 10 mg daily.

Busulfan, 1,4-butanediol dimethanesulfonate, is commercially available as MYLERAN® TABLETS. Busulfan is indicated for the palliative treatment of chronic myelogenous leukemia. Bone marrow suppression is the most common dose limiting side effects of busulfan. In one embodiment, busulfan is administered at a dose of 1.8mg/m² daily.

Carmustine, l,3-[bis(2-chloroethyl)-l-nitrosourea, is commercially available as single vials of lyophilized material as BiCNU®. Carmustine is indicated for the palliative treatment as a single agent or in combination with other agents for brain tumors, multiple myeloma, Hodgkin's disease, and non- Hodgkin's lymphomas. Delayed myelosuppression is the most common dose limiting side effects of carmustine. In one embodiment, carmustine is administered at 150 to 200 mg/m² IV every 6 weeks.

Dacarbazine, 5-(3,3-dimethyl-l-triazeno)-imidazole-4-carboxamide, is commercially available as single vials of material as DTIC-Dome®. Dacarbazine is indicated for the treatment of metastatic malignant melanoma and in combination with other agents for the second line treatment of Hodgkin's

Disease. Nausea, vomiting, and anorexia are the most common dose limiting side effects of dacarbazine. In one embodiment, dacarbazine is administered at 150 mg/m² IV once a day or 375 mg/m² IV every 15 days.

Antibiotic anti-neoplastics are non-phase specific agents, which bind or intercalate with DNA. Typically, such action results in stable DNA complexes or strand breakage, which disrupts ordinary function of the nucleic acids leading to cell death. Examples of antibiotic anti-neoplastic agents include, but are not limited to, actinomycins such as dactinomycin, anthrocyclins such as daunorubicin and doxorubicin; and bleomycins.

Dactinomycin, also known as Actinomycin D, is commercially available in injectable form as COSMEGEN®. Dactinomycin is indicated for the treatment of Wilm's tumor and rhabdomyosarcoma. Nausea, vomiting, and anorexia are the most common dose limiting side effects of dactinomycin.

Daunorubicin, (8S-cis-)-8-acetyl-10-[(3-amino-2,3,6-trideoxy-a-L-lyxo-hexopyranosyl)oxy]- 7,8,9,10-tetrahydro-6,8,ll-trihydroxy-l-methoxy-5,12 naphthacenedione hydrochloride, is commercially available as a liposomal injectable form as DAUNOXOME® or as an injectable as CERUBIDINE®. Daunorubicin is indicated for remission induction in the treatment of acute non lymphocytic leukemia and advanced HIV associated Kaposi's sarcoma. Myelosuppression is the most common dose limiting side effect of daunorubicin.

Doxorubicin, (8S, 10S)-10-[(3-amino-2,3,6-trideoxy-a-L-lyxo-hexopyranosyl)oxy]-8-glycoloyl, 7,8,9,10-tetrahydro-6,8,ll-trihydroxy-l-methoxy-5,12 naphthacenedione hydrochloride, is commercially available as an injectable form as RUBEX® or ADRIAMYCIN RDF®. Doxorubicin is primarily indicated for the treatment of acute lymphoblastic leukemia and acute myeloblastic leukemia, but is also a useful component in the treatment of some solid tumors and lymphomas. Myelosuppression is the most common dose limiting side effect of doxorubicin.

Bleomycin, a mixture of cytotoxic glycopeptide antibiotics isolated from a strain of Streptomyces verticillus, is commercially available as BLENOXANE®. Bleomycin is indicated as a palliative treatment, as a single agent or in combination with other agents, of squamous cell carcinoma, lymphomas, and testicular carcinomas. Pulmonary and cutaneous toxicities are the most common dose limiting side effects of bleomycin.

Topoisomerase II inhibitors include, but are not limited to, epipodophyllotoxins.

Epipodophyllotoxins are phase specific anti-neoplastic agents derived from the mandrake plant. Epipodophyllotoxins typically affect cells in the S and G2 phases of the cell cycle by forming a ternary complex with topoisomerase II and DNA causing DNA strand breaks. The strand breaks accumulate and cell death follows. Examples of epipodophyllotoxins include, but are not limited to, etoposide and teniposide. Etoposide, 4'-demethyl-epipodophyllotoxin 9[4,6-0-(R )-ethylidene-p-D-glucopyranoside], is commercially available as an injectable solution or capsules as VePESID® and is commonly known as VP-16. Etoposide is indicated as a single agent or in combination with other chemotherapy agents in the treatment of testicular and non-small cell lung cancers. Myelosuppression is the most common side effect of etoposide. The incidence of leucopenia tends to be more severe than thrombocytopenia.

Teniposide, 4'-demethyl-epipodophyllotoxin 9[4,6-0-(R )-thenylidene-p-D-glucopyranoside], is commercially available as an injectable solution as VUMON® and is commonly known as VM-26. Teniposide is indicated as a single agent or in combination with other chemotherapy agents in the treatment of acute leukemia in children. Myelosuppression is the most common dose limiting side effect of teniposide. Teniposide can induce both leucopenia and thrombocytopenia.

Anti metabolite neoplastic agents are phase specific anti-neoplastic agents that act at S phase (DNA synthesis) of the cell cycle by inhibiting DNA synthesis or by inhibiting purine or pyrimidine base synthesis and thereby limiting DNA synthesis. Consequently, S phase does not proceed and cell death follows. Examples of antimetabolite anti-neoplastic agents include, but are not limited to, fluorouracil, methotrexate, cytarabine, mecaptopurine, thioguanine, and gemcitabine.

5-fluorouracil, 5-fluoro-2,4- (114,31-1) pyrimidinedione, is commercially available as fluorouracil. Administration of 5-fluorouracil leads to inhibition of thymidylate synthesis and is also incorporated into both RNA and DNA. The result typically is cell death. 5-fluorouracil is indicated as a single agent or in combination with other chemotherapy agents in the treatment of carcinomas of the breast, colon, rectum, stomach and pancreas. Myelosuppression and mucositis are dose limiting side effects of 5-fluorouracil. Other fluoropyrimidine analogs include 5-fluoro deoxyuridine (floxuridine) and 5-fluorodeoxyuridine monophosphate. In one embodiment, fluorouracil is administered 1000mg/m²/day on day 1 through day 4 of every 21 day cycle (Q3W).

Cytarabine, 4-amino-1-b-D-arabinofuranosyl-2 (lH)-pyrimidinone, is commercially available as CYTOSAR-U® and is commonly known as Ara-C. It is believed that cytarabine exhibits cell phase specificity at S-phase by inhibiting DNA chain elongation by terminal incorporation of cytarabine into the growing DNA chain. Cytarabine is indicated as a single agent or in combination with other chemotherapy agents in the treatment of acute leukemia. Other cytidine analogs include 5-azacytidine and 2',2'-difluorodeoxycytidine (gemcitabine). Cytarabine induces leucopenia, thrombocytopenia, and mucositis.

Mercaptopurine, l,7-dihydro-6H-purine-6-thione monohydrate, is commercially available as PURINETHOL®. Mercaptopurine exhibits cell phase specificity at S-phase by inhibiting DNA synthesis by an as of yet unspecified mechanism. Mercaptopurine is indicated as a single agent or in combination with other chemotherapy agents in the treatment of acute leukemia. Myelosuppression and gastrointestinal mucositis are expected side effects of mercaptopurine at high doses. A useful mercaptopurine analog is azathioprine. Thioguanine, 2-amino-1,7-dihydro-6H-purine-6-thione, is commercially available as TABLOID®. Thioguanine exhibits cell phase specificity at S-phase by inhibiting DNA synthesis by an as of yet unspecified mechanism. Thioguanine is indicated as a single agent or in combination with other chemotherapy agents in the treatment of acute leukemia. Myelosuppression, including leucopenia, thrombocytopenia, and anemia, is the most common dose limiting side effect of thioguanine administration. However, gastrointestinal side effects occur and can be dose limiting. Other purine analogs include pentostatin, erythrohydroxynonyladenine, fludarabine phosphate, and cladribine.

Gemcitabine, 2'-deoxy-2', 2'-difluorocytidine monohydrochloride (b-isomer), is commercially available as GEMZAR®. Gemcitabine exhibits cell phase specificity at S-phase and by blocking progression of cells through the Gl/S boundary. Gemcitabine is indicated in combination with cisplatin in the treatment of locally advanced non-small cell lung cancer and alone in the treatment of locally advanced pancreatic cancer. Myelosuppression, including leucopenia, thrombocytopenia, and anemia, is the most common dose limiting side effect of gemcitabine administration.

Methotrexate, N-[4[[(2,4-diamino-6-pteridinyl) methyl]methylamino] benzoyl]-L-glutamic acid, is commercially available as methotrexate sodium. Methotrexate exhibits cell phase effects specifically at S-phase by inhibiting DNA synthesis, repair and/or replication through the inhibition of dyhydrofolic acid reductase which is required for synthesis of purine nucleotides and thymidylate. Methotrexate is indicated as a single agent or in combination with other chemotherapy agents in the treatment of choriocarcinoma, meningeal leukemia, non-Hodgkin's lymphoma, and carcinomas of the breast, head, neck, ovary and bladder. Myelosuppression (leucopenia, thrombocytopenia, and anemia) and mucositis are expected side effect of methotrexate administration.

Camptothecins, including, camptothecin and camptothecin derivatives are available or under development as Topoisomerase I inhibitors. Camptothecins cytotoxic activity is believed to be related to its Topoisomerase I inhibitory activity. Examples of camptothecins include, but are not limited to irinotecan, topotecan, and the various optical forms of 7-(4-methylpiperazino-methylene)-10,ll- ethylenedioxy-20-camptothecin described below.

Irinotecan HCI, (4S)-4,ll-diethyl-4-hydroxy-9-[(4-piperidinopiperidino) carbonyloxy]-lH- pyrano[3',4',6,7]indolizino[l,2-b]quinoline-3,14(4H,12H)-dione hydrochloride, is commercially available as the injectable solution CAMPTOSAR®.

Irinotecan is a derivative of camptothecin which binds, along with its active metabolite SN-38, to the topoisomerase I - DNA complex. It is believed that cytotoxicity occurs as a result of irreparable double strand breaks caused by interaction of the topoisomerase I : DNA : irintecan or SN-38 ternary complex with replication enzymes. Irinotecan is indicated for treatment of metastatic cancer of the colon or rectum. The dose limiting side effects of irinotecan HCI are myelosuppression, including neutropenia, and GI effects, including diarrhea. Topotecan HCI, (S)-10-[(dimethylamino)methyl]-4-ethyl-4,9-dihydroxy-lH- pyrano[3',4',6,7]indolizino[l,2-b]quinoline-3,14-(4H,12H)-dione monohydrochloride, is commercially available as the injectable solution HYCAMTIN®. Topotecan is a derivative of camptothecin which binds to the topoisomerase I - DNA complex and prevents religation of singles strand breaks caused by Topoisomerase I in response to torsional strain of the DNA molecule. Topotecan is indicated for second line treatment of metastatic carcinoma of the ovary and small cell lung cancer. The dose limiting side effect of topotecan HCI is myelosuppression, primarily neutropenia. mTOR inhibitors include but are not limited to rapamycin (FK506) and rapalogs, RAD001 or everolimus (Afinitor), CCI-779 or temsirolimus, AP23573, AZD8055, WYE-354, WYE-600, WYE-687 and Ppl21.

Bexarotene is sold as Targretin® and is a member of a subclass of retinoids that selectively activate retinoid X receptors (RXRs). These retinoid receptors have biologic activity distinct from that of retinoic acid receptors (RARs). The chemical name is 4-[l-(5,6,7,8-tetrahydro-3,5,5,8,8- pentamethyl-2-naphthalenyl) ethenyl] benzoic acID Bexarotene is used to treat cutaneous T-cell lymphoma CTCL, a type of skin cancer) in people whose disease could not be treated successfully with at least one other medication.

Sorafenib marketed as Nexavar® is in a class of medications called multikinase inhibitors. Its chemical name is 4-[4-[[4-chloro-3-(trifluoromethyl)phenyl]carbamoylamino] phenoxy]-/V-methyl- pyridine-2-carboxamide. Sorafenib is used to treat advanced renal cell carcinoma (a type of cancer that begins in the kidneys). Sorafenib is also used to treat unresectable hepatocellular carcinoma (a type of liver cancer that cannot be treated with surgery).

Examples of erbB inhibitors include lapatinib, erlotinib, and gefitinib. Lapatinib, A/-(3-chloro- 4-{[(3-fluorophenyl)methyl]oxy}phenyl)-6-[5-({[2-(methylsulfonyl)ethyl]amino}methyl)-2-furanyl]-4- quinazolinamine (represented by formula II, as illustrated), is a potent, oral, small-molecule, dual inhibitor of erbB-1 and erbB-2 (EGFR and HER2) tyrosine kinases that is approved in combination with capecitabine for the treatment of HER2-positive metastatic breast cancer.

The free base, HCI salts, and ditosylate salts of the compound of formula (II) may be prepared according to the procedures disclosed in WO 99/35146, published July 15, 1999; and WO 02/02552 published January 10, 2002.

Erlotinib, /V-(3-ethynylphenyl)-6,7-bis{[2-(methyloxy)ethyl]oxy}-4-quinazolinamine Commercially available under the tradename Tarceva) is represented by formula III, as illustrated:

III

The free base and HCI salt of erlotinib may be prepared, for example, according to U.S. 5,747,498, Example 20. Gefitinib, 4-quinazolinamine,N-(3-chloro-4-fluorophenyl)-7-methoxy-6-[3-4- morpholin)propoxy] is represented by formula IV, as illustrated:

Gefitinib, which is commercially available under the trade name IR.ESSA® (AstraZenenca) is an erbB-1 inhibitor that is indicated as monotherapy for the treatment of patients with locally advanced or metastatic non-small-cell lung cancer after failure of both platinum-based and docetaxel chemotherapies. The free base, HCI salts, and diHCI salts of gefitinib may be prepared according to the procedures of International Patent Application No. PCT/GB96/00961, filed April 23, 1996, and published as WO 96/33980 on October 31, 1996. Also of interest, is the camptothecin derivative of formula A following, currently under development, including the racemic mixture (R,S) form as well as the R and S enantiomers:

known by the chemical name "7-(4-methylpiperazino-methylene)-10,ll-ethylenedioxy- 20(R,S)-camptothecin (racemic mixture) or"7-(4-methylpiperazino-methylene)-10,ll-ethylenedioxy- 20(R)-camptothecin (R enantiomer) or "7-(4-methylpiperazino-methylene)-10,ll-ethylenedioxy- 20(S)-camptothecin (S enantiomer). Such compound as well as related compounds are described, including methods of making, in U.S. Patent Nos. 6,063,923; 5,342,947; 5,559,235; 5,491,237 and pending U.S. patent Application No. 08/977,217 filed November 24, 1997.

Signal transduction pathway inhibitors are those inhibitors, which block or inhibit a chemical process which evokes an intracellular change. As used herein this change is cell proliferation or differentiation. Signal transduction inhibitors useful in the present invention include inhibitors of receptor tyrosine kinases, non-receptor tyrosine kinases, SH2/SH3domain blockers, serine/threonine kinases, phosphotidyl inositol-3 kinases, myo-inositol signaling, and Ras oncogenes.

Several protein tyrosine kinases catalyse the phosphorylation of specific tyrosyl residues in various proteins involved in the regulation of cell growth. Such protein tyrosine kinases can be broadly classified as receptor or non-receptor kinases.

Receptor tyrosine kinases are transmembrane proteins having an extracellular ligand binding domain, a transmembrane domain, and a tyrosine kinase domain. Receptor tyrosine kinases are involved in the regulation of cell growth and are generally termed growth factor receptors. Inappropriate or uncontrolled activation of many of these kinases, i.e. aberrant kinase growth factor receptor activity, for example by over-expression or mutation, has been shown to result in uncontrolled cell growth. Accordingly, the aberrant activity of such kinases has been linked to malignant tissue growth. Consequently, inhibitors of such kinases could provide cancer treatment methods. Growth factor receptors include, for example, epidermal growth factor receptor (EGFr), platelet derived growth factor receptor (PDGFr), erbB2, erbB4, vascular endothelial growth factor receptor (VEGFr), tyrosine kinase with immunoglobulin-like and epidermal growth factor homology domains (TIE-2), insulin growth factor -I (IGFI) receptor, macrophage colony stimulating factor Cfms), BTK, ckit, cmet, fibroblast growth factor (FGF) receptors, Trk receptors (TrkA, TrkB, and TrkC), ephrin (eph) receptors, and the RET protooncogene. Several inhibitors of growth receptors are under development and include ligand antagonists, antibodies, tyrosine kinase inhibitors and anti-sense oligonucleotides. Growth factor receptors and agents that inhibit growth factor receptor function are described, for instance, in Kath, John C., Exp. Opin. Ther. Patents (2000) 10(6):803-818; Shawver et al DDT Vol 2, No. 2 February 1997; and Lofts, F. J. et al, "Growth factor receptors as targets", New Molecular Targets for Cancer Chemotherapy, ed. Workman, Paul and Kerr, David, CRC press 1994, London.

Tyrosine kinases, which are not growth factor receptor kinases are termed non-receptor tyrosine kinases. Non-receptor tyrosine kinases useful in the present invention, which are targets or potential targets of anti-cancer drugs, include cSrc, Lck, Fyn, Yes, Jak, cAbl, FAK (Focal adhesion kinase), Brutons tyrosine kinase, and Bcr-Abl. Such non-receptor kinases and agents which inhibit non-receptor tyrosine kinase function are described in Sinh, S. and Corey, S.J., (1999) Journal of Hematotherapy and Stem Cell Research 8 (5): 465 - 80; and Bolen, J.B., Brugge, J.S., (1997) Annual review of Immunology. 15: 371-404.

SH2/SH3 domain blockers are agents that disrupt SH2 or SH3 domain binding in a variety of enzymes or adaptor proteins including, PI3-K p85 subunit, Src family kinases, adaptor molecules (She, Crk, Nek, Grb2) and Ras-GAP. SH2/SH3 domains as targets for anti-cancer drugs are discussed in Smithgall, T.E. (1995), Journal of Pharmacological and Toxicological Methods. 34(3) 125-32.

Inhibitors of Serine/Threonine Kinases including MAP kinase cascade blockers which include blockers of Raf kinases (rafk), Mitogen or Extracellular Regulated Kinase (MEKs), and Extracellular Regulated Kinases (ERKs); and Protein kinase C family member blockers including blockers of PKCs (alpha, beta, gamma, epsilon, mu, lambda, iota, zeta). IkB kinase family (IKKa, IKKb), PKB family kinases, AKT kinase family members, and TGF beta receptor kinases. Such Serine/Threonine kinases and inhibitors thereof are described in Yamamoto, T., Taya, S., Kaibuchi, K., (1999), Journal of Biochemistry. 126 (5) 799-803; Brodt, P, Samani, A., and Navab, R. (2000), Biochemical Pharmacology, 60. 1101-1107; Massague, J., Weis-Garcia, F. (1996) Cancer Surveys. 27:41-64; Philip, P.A., and Harris, A.L. (1995), Cancer Treatment and Research. 78: 3-27, Lackey, K. et al Bioorganic and Medicinal Chemistry Letters, (10), 2000, 223-226; U.S. Patent No. 6,268,391; and Martinez-Iacaci, L, et al, Int. J. Cancer (2000), 88(1), 44-52.

Inhibitors of Phosphotidyl inositol-3 Kinase family members including blockers of PI3-kinase, ATM, DNA-PK, and Ku are also useful in the present invention. Such kinases are discussed in Abraham, R.T. (1996), Current Opinion in Immunology. 8 (3) 412-8; Canman, C.E., Lim, D.S. (1998), Oncogene 17 (25) 3301-3308; Jackson, S.P. (1997), International Journal of Biochemistry and Cell Biology. 29 (7):935-8; and Zhong, H. et al, Cancer res, (2000) 60(6), 1541-1545.

Also useful in the present invention are Myo-inositol signaling inhibitors such as phospholipase C blockers and Myoinositol analogues. Such signal inhibitors are described in Powis, G., and Kozikowski A., (1994 New Molecular Targets for Cancer Chemotherapy ed., Paul Workman and David Kerr, CRC press 1994, London. Another group of signal transduction pathway inhibitors are inhibitors of Ras Oncogene. Such inhibitors include inhibitors of farnesyltransferase, geranyl-geranyl transferase, and CAAX proteases as well as anti-sense oligonucleotides, ribozymes and immunotherapy. Such inhibitors have been shown to block ras activation in cells containing wild type mutant ras, thereby acting as antiproliferation agents. Ras oncogene inhibition is discussed in Scharovsky, O.G., Rozados, V.R., Gervasoni, S.I. Matar, P. (2000), Journal of Biomedical Science. 7(4 292-8; Ashby, M.N. (1998), Current Opinion in Lipidology. 9 (2) 99 - 102; and Bennett, C.F. and Cowsert, L.M. BioChim. Biophys. Acta, (1999) 1489(1): 19-30.

As mentioned above, antibody antagonists to receptor kinase ligand binding may also serve as signal transduction inhibitors. This group of signal transduction pathway inhibitors includes the use of humanized antibodies to the extracellular ligand binding domain of receptor tyrosine kinases. For example Imclone C225 EGFR specific antibody (see Green, M.C. et al, Monoclonal Antibody Therapy for Solid Tumors, Cancer Treat. Rev., (2000), 26(4, 269-286); Herceptin^® erbB2 antibody (see Tyrosine Kinase Signalling in Breast cancenerbB Family Receptor Tyrosine Kinases, Breast cancer Res., 2000, 2(3), 176-183); and 2CB VEGFR2 specific antibody (see Brekken, R.A. et al, Selective Inhibition of VEGFR2 Activity by a monoclonal Anti-VEGF antibody blocks tumor growth in mice, Cancer Res. (2000) 60, 5117-5124.

Non-receptor kinase angiogenesis inhibitors may also find use in the present invention. Inhibitors of angiogenesis related VEGFR and TIE2 are discussed above in regard to signal transduction inhibitors (both receptors are receptor tyrosine kinases). Angiogenesis in general is linked to erbB2/EGFR signaling since inhibitors of erbB2 and EGFR have been shown to inhibit angiogenesis, primarily VEGF expression. Thus, the combination of an erbB2/EGFR inhibitor with an inhibitor of angiogenesis makes sense. Accordingly, non-receptor tyrosine kinase inhibitors may be used in combination with the EGFR/erbB2 inhibitors of the present invention. For example, anti-VEGF antibodies, which do not recognize VEGFR (the receptor tyrosine kinase), but bind to the ligand; small molecule inhibitors of integrin (alpha_v betas) that will inhibit angiogenesis; endostatin and angiostatin (non-RTK) may also prove useful in combination with the disclosed erb family inhibitors. (See Bruns CJ et al (2000), Cancer Res., 60: 2926-2935; Schreiber AB, Winkler ME, and Derynck R. (1986), Science, 232: 1250-1253; Yen L et al. (2000), Oncogene 19: 3460-3469).

Agents used in immunotherapeutic regimens may also be useful in combination with the compounds of formula (I). There are a number of immunologic strategies to generate an immune response against erbB2 or EGFR. These strategies are generally in the realm of tumor vaccinations. The efficacy of immunologic approaches may be greatly enhanced through combined inhibition of erbB2/EGFR signaling pathways using a small molecule inhibitor. Discussion of the immunologic/tumor vaccine approach against erbB2/EGFR are found in Reilly RT et al. (2000), Cancer Res. 60: 3569-3576; and Chen Y, Hu D, Eling DJ, Robbins J, and Kipps TJ. (1998), Cancer Res. 58: 1965-1971. Agents used in proapoptotic regimens (e.g., bcl-2 antisense oligonucleotides) may also be used in the combination of the present invention. Members of the Bcl-2 family of proteins block apoptosis. Upregulation of bcl-2 has therefore been linked to chemoresistance. Studies have shown that the epidermal growth factor (EGF) stimulates anti-apoptotic members of the bcl-2 family (i.e., mcl-1). Therefore, strategies designed to downregulate the expression of bcl-2 in tumors have demonstrated clinical benefit and are now in Phase II/III trials, namely Genta's G3139 bcl-2 antisense oligonucleotide. Such proapoptotic strategies using the antisense oligonucleotide strategy for bcl-2 are discussed in Water JS et al. (2000), J. Clin. Oncol. 18: 1812-1823; and Kitada S et al. (1994, Antisense Res. Dev. 4: 71-79.

Cell cycle signalling inhibitors inhibit molecules involved in the control of the cell cycle. A family of protein kinases called cyclin dependent kinases CDKs) and their interaction with a family of proteins termed cyclins controls progression through the eukaryotic cell cycle. The coordinate activation and inactivation of different cyclin/CDK complexes is necessary for normal progression through the cell cycle. Several inhibitors of cell cycle signalling are under development. For instance, examples of cyclin dependent kinases, including CDK2, CDK4, and CDK6 and inhibitors for the same are described in, for instance, Rosania et al, Exp. Opin. Ther. Patents (2000) 10(2):215-230.

In one embodiment, the chemotherapeutic agent(s) is one or a combination of chemotherapeutic agents selected from docetaxel, pemetrexed, paclitaxel, gemcitabine, 5-FU, carboplatin and cisplatin.

Immunomodulatory agent

As used herein an "immunomodulatory agent" or"immuno-modulator" refers to any substance including monoclonal antibodies that effects the immune system. The agonist ICOS binding proteins of the present invention can be considered an immunomodulatory agent. Immuno-modulators can be used as anti-neoplastic agents for the treatment of cancer. For example, immune-modulators include, but are not limited to, anti-CTLA-4 antibodies such as ipilimumab (YERVOY), tremelimumab and anti-PD-1 antibodies (e.g. dostarlimab, OPDIVO/nivolumab, KEYTRUDA/pembrolizumab and LIBTAYO/cemiplimab). Other immuno-modulators include, but are not limited to, OX-40 antibodies, PD-L1 antibodies, such as bintrafusp alpha, LAG3 antibodies, TIM-3 antibodies, 41BB antibodies and GITR antibodies.

Yervoy (ipilimumab) is a fully human CTLA-4 antibody marketed by Bristol Myers Squibb. The protein structure of ipilimumab and methods are using are described in US Patent Nos. 6,984,720 and 7,605,238.

Opdivo/nivolumab is a fully human monoclonal antibody marketed by Bristol Myers Squibb directed against the negative immunoregulatory human cell surface receptor PD-1 (programmed death-1 or programmed cel! death- 1/PCD- 1) with immunopotentiation activity. Nivolumab binds to and blocks the activation of PD-1, an Ig superfamily transmembrane protein, by its ligands PD-L1 and PD-L2, resulting in the activation of T-cells and cell-mediated immune responses against tumor cells or pathogens. Activated PD-1 negatively regulates T-cell activation and effector function through the suppression of P13k/Akt pathway activation. Other names for nivolumab include: BMS-936558, MDX- 1106, and ONO-4538. The amino acid sequence for nivolumab and methods of using and making are disclosed in US Patent No. US 8,008,449.

KEYTRUDA/pembrolizumab is an anti-PD-1 antibodies marketed for the treatment of lung cancer by Merck. The amino acid sequence of pembrolizumab and methods of using are disclosed in US Patent No. 8,168,757.

CD134, also known as 0X40, is a member of the TNFR-superfamily of receptors which is not constitutively expressed on resting naive T cells, unlike CD28. 0X40 is a secondary costimulatory molecule, expressed after 24 to 72 hours following activation; its ligand, OX40L, is also not expressed on resting antigen presenting cells, but is following their activation. Expression of 0X40 is dependent on full activation of the T cell; without CD28, expression of 0X40 is delayed and of fourfold lower levels. OX-40 antibodies, OX-40 fusion proteins and methods of using them are disclosed in US Patent Nos: US 7,504,101; US 7,758,852; US 7,858,765; US 7,550,140; US 7,960,515; WO2012027328; WO2013028231.

Antibodies to PD-L1 (also referred to as CD274 or B7-H1) and methods for use are disclosed in US Patent No. 7,943,743; US Patent No. 8,383,796; US20130034559, WO2014055897, US Patent No. 8,168,179; and US Patent No. 7,595,048. PD-L1 antibodies are in development as immunomodulatory agents for the treatment of cancer.

As used herein "immunostimulatory agent" refers to any agent that can stimulate the immune system. As used herein immunostimulatory agents include, but are not limited to, vaccine adjuvants.

Aminoalkyl glucosaminide phosphates (AGPs) are known to be useful as vaccine adjuvants and immunostimulatory agents for stimulating cytokine production, activating macrophages, promoting innate immune response, and augmenting antibody production in immunized animals. Aminoalkyl glucosaminide phosphates (AGPs) are synthetic ligands of the Toll-like Receptor 4 (TLR4). AGPs and their immunomodulating effects via TLR4 are disclosed in patent publications such as WO 2006/016997, WO 2001/090129, and/or U.S. Patent No. 6,113,918 and have been reported in the literature. Additional AGP derivatives are disclosed in U.S. Patent No. 7,129,219, U.S. Patent No. 6,525,028 and U.S. Patent No 6,911,434. Certain AGPs act as agonists of TLR4, while others are recognized as TLR4 antagonists.

Aminoalkyl glucosaminide phosphate compounds employed in the present invention have the structure set forth in Formula 1 as follows:

(Formula 1) wherein m is 0 to 6 n is 0 to 4;

X is 0 or S, preferably 0;

Y is 0 or NH;

Z is 0 or H; each R₁, R₂, R₃ is selected independently from the group consisting of a C_1-20 acyl and a C_1-20 alkyl;

R4 is H or Me; R₅ is selected independently from the group consisting of -H, -OH, -(C_I-C₄) alkoxy, - PO3R₈R₉, -OPO₃R₈R₉, -SO₃R₈, -OSO₃R₈, -NR₈R₉, -SR₈, -CN, -NO₂, -CHO, -CO₂R₈, and -CONR₈R₉, wherein R₈ and R₉ are each independently selected from H and (C_I-C₄) alkyl; and each R₆ and R₇ is independently H or PO₃H₂.

In Formula 1 the configuration of the 3' stereogenic centers to which the normal fatty acyl residues (that is, the secondary acyloxy or alkoxy residues, e.g., R₁O, R₂O, and R₃O) are attached is R or S, preferably R (as designated by Cahn-Ingold-Prelog priority rules). Configuration of aglycon stereogenic centers to which R₄ and R₅ are attached can be R or S. All stereoisomers, both enantiomers and diastereomers, and mixtures thereof, are considered to fall within the scope of the present invention.

The number of carbon atoms between heteroatom X and the aglycon nitrogen atom is determined by the variable "n", which can be an integer from 0 to 4, preferably an integer from 0 to 2.

The chain length of normal fatty acids R₁, R₂, and R₃ can be from about 6 to about 16 carbons, preferably from about 9 to about 14 carbons. The chain lengths can be the same or different. Some preferred embodiments include chain lengths where RI, R2 and R3 are 6 or 10 or 12 or 14. Formula 1 encompasses L/D-seryl, -threonyl, -cysteinyl ether and ester lipid AGPs, both agonists and antagonists and their homologs (n=1-4), as well as various carboxylic acid bioisosteres (i.e, R₅ is an acidic group capable of salt formation; the phosphate can be either on 4- or 6- position of the glucosamine unit, but preferably is in the 4-position). In a preferred embodiment of the invention employing an AGP compound of Formula 1, n is

0, R₅ is CO₂H, R₆ is PO₃H₂, and R₇ is H. This preferred AGP compound is set forth as the structure in Formula la as follows:

(Formula la) wherein X is 0 or S; Y is 0 or NH; Z is 0 or H; each R₁, R₂, R₃ is selected independently from the group consisting of a C_1-20 acyl and a C_1-20 alkyl; and R₄ is H or methyl.

In Formula la the configuration of the 3' stereogenic centers to which the normal fatty acyl residues (that is, the secondary acyloxy or alkoxy residues, e.g., R₁O, R₂O, and R₃O) are attached as R or S, preferably R (as designated by Cahn-Ingold-Prelog priority rules). Configuration of aglycon stereogenic centers to which R₄ and CO₂H are attached can be R or S. All stereoisomers, both enantiomers and diastereomers, and mixtures thereof, are considered to fall within the scope of the present invention.

Formula la encompasses L/D-seryl, -threonyl, -cysteinyl ether or ester lipid AGPs, both agonists and antagonists.

In both Formula 1 and Formula la, Z is 0 attached by a double bond or two hydrogen atoms which are each attached by a single bond. That is, the compound is ester-linked when Z=Y=0; amide- linked when Z =0 and Y=NH; and ether-linked when Z=H/H and Y=0.

Especially preferred compounds of Formula 1 are referred to as CRX-601 and CRX-527. Their structures are set forth as follows:

5 Additionally, another preferred embodiment employs CRX 547 having the structure shown. CRX 547

Still other embodiments include AGPs such as CRX 602 or CRX 526 providing increased stability having shorter secondary acyl or alkyl chains.

CRX 602

In one embodiment, at least one second immunomodulatory agent is administered. In one embodiment said second immunomodulatory agent is selected from the group of: an anti-CTLA4 antibody, an anti-PD-1 antibody, an anti-PD-Ll antibody, an anti-OX40 antibody, an anti-BCMA antibody, an anti-GITR antibody, and anti-41BB antibody, an anti-LAG3 antibody and an anti-TIM3 antibody.

In one embodiment at least one immuno-stimulatory agent is administered. In one embodiment, the immuno-stimulatory agent is a TLR4 agonist. In one embodiment the immune- stimulatory agent is an AGP. In one embodiment, the immuno-stimulatory agent is a compound of Formula I. In one embodiment, it is a compound of Formula la. In one embodiment, the immune- stimulatory agent is selected from the group consisting of: CRX-601, CRX-547, CRX-602, CRX-527, and CRX-526.

Methods of Treatment

The therapeutic agents described herein may also be used in methods of treatment. It will be appreciated by those skilled in the art that references herein to treatment refer to the treatment of established conditions. However, compositions of the invention may, depending on the condition, also be useful in the prevention of certain diseases. The therapeutic agents described herein can be used in an effective amount for therapeutic, prophylactic or preventative treatment. A therapeutically effective amount of the therapeutic agents described herein is an amount effective to ameliorate or reduce one or more symptoms of, or to prevent or cure, the disease.

In one aspect, a method of treating cancer in a human in need thereof, the method comprising administering to the human an agonist ICOS binding protein or antigen binding portion thereof at a dose of about 0.08 mg to about 240 mg and administering to the human a chemotherapeutic agent.

In one aspect, there is provided a composition comprising an agonist ICOS binding protein or antigen binding portion thereof at about 0.08 mg to about 240 mg and a chemotherapeutic agent.

Routes of administration and dosages

The doses provided in the present application are suitable for mammals, in particular a human. It is to be understood that where agonist ICOS binding protein is used herein, the antigen binding portion thereof is also implied.

In some embodiments, a therapeutically effective dose of the agonist ICOS binding protein is a dose of about 0.01 - 1000 mg (e.g. a dose about 0.01 mg; a dose about 0.08 mg; a dose about 0.1 mg; a dose about 0.24 mg; a dose about 0.8 mg; a dose about 1 mg; a dose about 2.4 mg; a dose about 7.2 mg; a dose about 8 mg; a dose about 10 mg; a dose about 20 mg; a dose about 24 mg; a dose about 30 mg; a dose about 40 mg; a dose about 48 mg; a dose about 50 mg; a dose about 60 mg; a dose about 70 mg; a dose about 72 mg; a dose about 80 mg; a dose about 90 mg; a dose about 100 mg; a dose about 160 mg; a dose about 200 mg; a dose about 240 mg; a dose about 300 mg; a dose about 320 mg; a dose about 400 mg; a dose about 480 mg; a dose about 500 mg; a dose about 600 mg; a dose about 700 mg; a dose about 720 mg; a dose about 800 mg; a dose about 900 mg; or a dose about 1000 mg).

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg. In one embodiment, the agonist ICOS binding protein is administered at a dose of 0.08 mg, 0.24 mg, 0.8 mg, 2.4 mg, 8 mg, 24 mg, 48 mg, 80 mg, 160 mg or 240 mg in particular 24 mg, 48 mg, 80 mg or 160 mg.

It is to be understood that where mg/kg is used, this is mg/kg of body weight. In some embodiments, a therapeutically effective dose of the agonist ICOS binding protein is a dose of about 0.001 mg/kg to 10 mg/kg. In some embodiments, a therapeutically effective dose is about 0.001 mg/kg. In some embodiments, a therapeutically effictive dose is about 0.003 mg/kg. In some embodiments, a therapeutically effective dose is about 0.01 mg/kg. In some embodiments, a therapeutically effective dose is about 0.03 mg/kg. In some embodiments, a therapeutically effective dose is about 0.1 mg/kg. In some embodiments, a therapeutically effective dose is about 0.3 mg/kg. In some embodiments, a therapeutically effective dose is about 0.6 mg/kg. In some embodiments, a therapeutically effective dose is about 1 mg/kg. In some embodiments, a therepeutically effective dose is about 2 mg/kg. In some embodiments, a therapeutically effective dose is about 3 mg/kg. In some embodiments, a therapeutically effective dose is about 4 mg/kg; about 5 mg/kg; about 6 mg/kg; about 7 mg/kg; about 8 mg/kg; about 9 mg/kg or about 10 mg/kg.

In one embodiment, the dose of the agonist ICOS binding protein is between about 0.001 mg/kg to about 3.0 mg/kg. In another embodiment, the dose of the agonist ICOS binding protein is about 0.001 mg/kg, about 0.003 mg/kg, about 0.01 mg/kg, about 0.03 mg/kg, about 0.1 mg/kg, about 0.3 mg/kg, about 1.0 mg/kg, about 3.0 mg/kg, or about 10 mg/kg. In one embodiment, the dose of agonist ICOS binding protein is about 0.3 mg/kg. In another embodiment, the dose of the agonist ICOS binding protein is at least 3.0 mg/kg. In one embodiment, the dose of the agonist ICOS binding protein is in the range of about 0.001 mg/kg to about 10 mg/kg. In one embodiment, the dose of the agonist ICOS protein is about 0.1 mg/kg to about 3 mg/kg. In one embodiment, the dose of the ICOS binding protein is about 0.1 mg/kg to about 1.0 mg/kg. In one embodiment, the dose of the agonist ICOS binding protein is about 0.1 mg/kg. In one embodiment, the dose of the ICOS binding protein is at least 0.1 mg/kg. In another embodiment, the dose of the agonist ICOS binding protein is about 0.3 mg/kg. In another embodiment, the dose of the agonist ICOS binding protein is about 1 mg/kg. In one embodiment, the dose of the agonist ICOS binding protein is about 3 mg/kg. In one embodiment, a fixed dose of agonist ICOS binding protein may be administered, assuming a typical median weight of 80 kg.

In one embodiment, the dose of agonist ICOS binding protein is increased during the treatment regimen. In one embodiment an initial dose of about 0.001 mg/kg, about 0.003 mg/kg, about 0.01 mg/kg, about 0.03 mg/kg, about 0.1 mg/kg, about 0.3 mg/kg, about 1.0 mg/kg is increased to about 0.003 mg/kg, about 0.01 mg/kg, about 0.03 mg/kg, about 0.1 mg/kg, about 0.3 mg/kg, about 1.0 mg/kg, about 3.0 mg/kg or at least 3.0 mg/kg. In one embodiment, an initial dose of 0.1 mg/kg is increased to 1 mg/kg. In one embodiment, an initial dose of 0.3 mg/kg is increased to 1 mg/kg. In one embodiment, the initial dose of 0.6 mg/kg is increased to 2 mg/kg.

In one embodiment, the agonist ICOS binding protein is administered at 0.1 mg/kg x 3 doses then 1 mg/kg. In one embodiment, the agonist ICOS binding protein is administered at about 0.001 mg/kg, about 0.003 mg/kg, about 0.01 mg/kg, about 0.03 mg/kg, about 0.1 mg/kg, about 0.3 mg/kg, about 1.0 mg/kg, or about 3.0 mg/kg then increased to about 0.01 mg/kg, about 0.03 mg/kg, about 0.1 mg/kg, about 0.3 mg/kg, about 1.0 mg/kg, about 3.0 mg/kg or about 10 mg/kg.

The chemotherapeutic agent may be administered as a single agent or in combination with other chemotherapeutic agents. In one embodiment, the doses of chemotherapeutic agent(s) administered in combination other chemotherapeutic agents are according to Category 1 recommendations in National Comprehensive Cancer Network treatment guidelines.

In some embodiments, the dosage regimen of the chemotherapeutic agent(s) is as per local standard of care.

It is to be understood that where mg/ m² is used, this is mg/ m² of body surface.

In one embodiment, docetaxel is administered at a dose of about 30 mg/m². In one embodiment, docetaxel is administered at a dose of about 75 mg/m². In one embodiment, docetaxel is administered at a dose of about 100 mg/m². In one embodiment, docetaxel is administered via IV infusion. In one embodiment, docetaxel is administered via IV infusion over 1 hour. In one embodiment, docetaxel is administered once every three weeks. In one embodiment, docetaxel is administered once every three weeks for 1, 2, 3, 4, 5 or 6 cycles, . In one embodiment, 30 mg/m² docetaxel is administered weekly for 5 weeks in a 6 week cycle for up to 5 cycles. In one embodiment, docetaxel is administered at a dose of about 75 mg/m² once every three weeks for up to 10 cycles. In one embodiment, docetaxel is administered at a dose of about 75 mg/m² once every three weeks via IV infusion.

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg or about 0.001 mg/kg to about 3 mg/kg and the docetaxel is administered at a dose of 75 mg/m². In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg or 0.3 mg/kg and the docetaxel is administered at a dose of 75 mg/m². In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 80 mg or 1 mg/kg the docetaxel is administered at a dose of 75 mg/m².

In one embodiment, pemetrexed is administered at a dose of about 500 mg/m². In one embodiment, pemetrexed is administered via IV infusion. In one embodiment, pemetrexed is administered once every three weeks. In one embodiment, pemetrexed is administered at a dose of about 500 mg/m² once every three weeks via IV infusion.

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg or about 0.001 mg/kg to about 3 mg/kg and the pemetrexed is administered at a dose of 500 mg/m². In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg or 0.3 mg/kg and the pemetrexed is administered at a dose of 500 mg/m². In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 80 mg or 1 mg/kg and the pemetrexed is administered at a dose of 500 mg/m².

In one embodiment, paclitaxel is administered at a dose of about 135 mg/m² to about 225 mg/m². In one embodiment, paclitaxel is administered at a dose of about 175 mg/m². In one embodiment, paclitaxel is administered at a dose of about 200 mg/m². In one embodiment, paclitaxel is administered at a dose of about 225 mg/m². In one embodiment, paclitaxel is administered via IV infusion. In one embodiment, paclitaxel is administered via IV infusion over 3 hours. In one embodiment, paclitaxel is administered once every three weeks. In one embodiment, paclitaxel is administered once every three weeks for up to 8 cycles, (i.e. 1, 2, 3, 4, 5, 6, 7 or 8 cycles). In one embodiment, paclitaxel is administered at a dose of about 200 mg/m² once every three weeks via IV infusion. In one embodiment, paclitaxel is administered at a dose of about 135 mg/m² to 175 mg/m² every three weeks.

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg or about 0.001 mg/kg to about 3 mg/kg and the paclitaxel is administered at a dose of 200 mg/m². In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg or 0.3 mg/kg and the paclitaxel is administered at a dose of 200 mg/m². In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 80 mg or 1 mg/kg and the paclitaxel is administered at a dose of 200 mg/m².

In one embodiment, gemcitabine is administered at a dose of about 1000 mg/m² to about 1250 mg/m². In one embodiment, gemcitabine is administered at a dose of about 1250 mg/m². In one embodiment, gemcitabine is administered via IV infusion. In one embodiment, gemcitabine is administered on Days 1 and 8 of each 21 day cycle (Q3W). In one embodiment, gemcitabine is administered on Days 1, 8 and 15 of each 28 day cycle (Q4W).In one embodiment, gemcitabine is administered in a 4 week cycle, wherein it is administered once weekly for 3 weeks followed by a 1 week rest period. In one embodiment, gemcitabine is administered in an 8 week cycle, wherein gemcitabine is administered weekly for 7 weeks followed by a week of rest. In one embodiment, gemcitabine is administered once every three weeks. In one embodiment, gemcitabine is administered at a dose of about 1250 mg/m² once every three weeks via IV infusion. In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg or about 0.001 mg/kg to about 3 mg/kg and the gemcitabine is administered at a dose of 1250 mg/m² on Days 1 and 8 of each 21 day cycle (Q3W). In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg or 0.3 mg/kg and the gemcitabine is administered at a dose of 1250 mg/m² on Days 1 and 8 of each 21 day cycle (Q3W). In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 80 mg or 1 mg/kg and the gemcitabine is administered at a dose of 1250 mg/m² on Days 1 and 8 of each 21 day cycle (Q3W).

In one embodiment, 5-FU is administered at a dose of about 200 mg/m² to about 1200 mg/m². In one embodiment, 5-FU is administered at a dose of about 200 mg/m² to about 1000 mg/m². In one embodiment, 5-FU is administered at a dose of about 200 mg/m² to about 600 mg/m². In one embodiment, 5-FU is administered at a dose of about 200 mg/m² to about 500 mg/m². In one embodiment, 5-FU is administered at a dose of about 500 mg/m² to about 600 mg/m². In one embodiment, 5-FU is administered at a dose of about 200 mg/m². 5-FU may administered continuously over several days. In such cases, the unit mg/m²/day is used. In one embodiment, 5-FU is administered at a dose of about 1000 mg/m² to about 1200 mg/m²/day. In one embodiment, 5-FU is administered at a dose of about 1000 mg/m²/day. In one embodiment, 5-FU is administered via IV infusion. In one embodiment, 5-FU is administered on Day 1 through Day 4 of every 21-day cycle (Q3W). In one embodiment 5-FU is administered once every week. In one embodiment 5-FU is administered once every 3 weeks. In one embodiment 5-FU is administered once every 4 weeks. In one embodiment 5-FU is administered monthly or bimonthly. In one embodiment, 5-FU is administered at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of every 21-day cycle (Q3W) via IV infusion.

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg or about 0.001 mg/kg to about 3 mg/kg and the 5-FU is administered at a dose of 1000 mg/m²/day on Day 1 through Day 4 of every 21-day cycle (Q3W). In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg or 0.3 mg/kg and the 5-FU is administered at a dose of 1000 mg/m²/day on Day 1 through Day 4 of every 21-day cycle (Q3W). In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 80 mg or 1 mg/kg and the 5-FU is administered at a dose of 1000 mg/m²/day on Day 1 through Day 4 of every 21-day cycle (Q3W).

In one embodiment, carboplatin is administered at a dose of about AUC 4-7 mg/ml per min. In one embodiment, carboplatin is administered at a dose of about AUC 4-6 mg/ml per min. In one embodiment, carboplatin is administered at a dose of about AUC 5 mg/ml per min. In one embodiment, carboplatin is administered at a dose of about 300 mg/m² to about 360 mg/m². In one embodiment, carboplatin is administered via IV infusion. In one embodiment, carboplatin is administered intra peritonea I ly. In one embodiment, carboplatin is administered once every three weeks. In one embodiment, carboplatin is administered once every 4 weeks. In one embodiment, carboplatin is administered at a dose of about AUC 4-6 mg/ml per min once every three weeks via IV infusion. In one embodiment, carboplatin is administered at a dose of about 300 mg/m² to about 360 mg/m² once every 4 weeks via IV infusion.

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg or about 0.001 mg/kg to about 3 mg/kg and the carboplatin is administered at a dose of AUC 4-6 mg/ml per min. In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg or 0.3 mg/kg and the carboplatin is administered at a dose of AUC 4-6 mg/ml per min. In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 80 mg or 1 mg/kg the carboplatin is administered at a dose of AUC 4-6 mg/ml per min.

In one embodiment, cisplatin is administered at a dose of about 20 mg/m² to about 120 mg/m². In one embodiment, cisplatin is administered at a dose of about 20 mg/m². In one embodiment, cisplatin is administered at a dose of about 40 mg/m². In one embodiment, cisplatin is administered at a dose of about 100 mg/m². In one embodiment, docetaxel is administered via IV infusion. In one embodiment, cisplatin is administered once every three weeks (Q3W). In one embodiment, cisplatin is administered once every 4 weeks. In one embodiment, cisplatin is administered once every week for 6 weeks. In one embodiment, cisplatin is administered at a dose of about 75 mg/m² once every three weeks via IV infusion. In one embodiment, cisplatin is administered at a dose of 75 mg/m² to about 100 mg/m² once every four weeks or about 50 mg/m² to about 70 mg/m² once every three to four weeks once every three weeks via IV infusion.

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg or about 0.001 mg/kg to about 3 mg/kg and the cisplatin is administered at a dose of 100 mg/m². In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg or 0.3 mg/kg and the cisplatin is administered at a dose of 100 mg/m². In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 80 mg or 1 mg/kg the cisplatin is administered at a dose of 100 mg/m².

In some embodiments, a combination (e.g. doublet) of chemotherapeutic agents may be administered. In one embodiment, the combination of chemotherapeutic agents is carboplatin and one of pemetrexed or paclitaxel or gemcitabine or fluorouracil. In one embodiment, the combination of chemotherapeutic agents is carboplatin and pemetrexed. In one embodiment, the combination of chemotherapeutic agents is carboplatin and paclitaxel. In one embodiment, the combination of chemotherapeutic agents is carboplatin and gemcitabine. In one embodiment, the combination of chemotherapeutic agents is carboplatin and fluorouracil.

In one embodiment, the combination of chemotherapeutic agents is cisplatin and one of pemetrexed or paclitaxel or gemcitabine or fluorouracil. In one embodiment, the combination of chemotherapeutic agents is cisplatin and pemetrexed. In one embodiment, the combination of chemotherapeutic agents is cisplatin and paclitaxel. In one embodiment, the combination of chemotherapeutic agents is cisplatin and gemcitabine. In one embodiment, the combination of chemotherapeutic agents is cisplatin and fluorouracil.

The carboplatin chemotherapy doublets may calculate the dose of carboplatin according to Calvert formula using the target dose of about AUC of 4-6 mg/ml per min as per local standard of care in combination with one of i) pemetrexed at a dose of about 500 mg/m²; ii) gemcitabine at a dose of about 1250 mg/m²; and iii) paclitaxel at a dose of about 200 mg/m².

In one embodiment, the combination of chemotherapeutic agents is a platinum-based chemotherapy doublet. In one embodiment, the platinum-based chemotherapy doublet is 5-FU and carboplatin or cisplatin. In one embodiment, the platinum-based chemotherapy doublet is 5-FU and carboplatin. In one embodiment, the platinum-based chemotherapy doublet is 5-FU and cisplatin.

In one embodiment, the 5-FU is administered at a dose of about 1000 mg/m² and the carboplatin is administered at a dose of about AUC 5 mg/ml per min. In one embodiment, the 5-FU is administered at a dose of about 1000 mg/m² and the cisplatin is administered at a dose of about 100 mg/m².

In some embodiments, the agonist ICOS binding protein or antigen binding portion thereof is administered prior to the chemotherapeutic agent(s). In some embodiments, the chemotherapeutic agent(s) are administered within at least 30 minutes and no longer than one hour following administration of the agonist ICOS binding protein or antigen binding portion thereof.

In some embodiments, administration of the chemotherapeutic agent is started 1 hour and no more than 2 hours after the end of the administration of agonist ICOS binding protein or antigen binding portion thereof. In one embodiment, administration of agonist ICOS binding protein or antigen binding portion thereof is started 1 hour and no more than 2 hours after the end of administration of the chemotherapeutic agent.

Premedication regimens and supplementation may be administered according to the product label or standard of care of the respective chemotherapeutic agent(s). Chemotherapy premedication administered on day of dosing may be administered after administration of the agonist ICOS binding protein or antigen binding portion thereof. The chemotherapeutic agent(s) may be administered for a minimum of 4 and maximum of 6 cycles according to standard of care. Treatment with docetaxel and pemetrexed may continue beyond 6 cycles according to standard of care. The therapeutic agents disclosed herein may be administered either in separate or combined form ( e.g . as pharmaceutical formulations) by any convenient route. For some therapeutic agents (i.e. binding proteins), suitable routes include oral, rectal, nasal, topical (including buccal and sublingual), vaginal, and parenteral (including subcutaneous, intramuscular, intravenous, intradermal, intrathecal, and epidural). It will be appreciated that the preferred route may vary with, for example, the condition of the recipient of the combination and the cancer to be treated. It will also be appreciated that each of the agents administered may be administered by the same or different routes and that the therapeutic agents may be formulated together or in separate pharmaceutical compositions.

In one embodiment, the therapeutic agent(s) is administered intravenously. In a further embodiment, the therapeutic agent(s) is administered by intravenous infusion. In another embodiment, the therapeutic agent(s) administered intratu morally. In another embodiment, the therapeutic agent(s) is administered orally. In another embodiment, the therapeutic agent(s) is administered systemically, e.g. intravenously, and one or more other therapeutic agents of the invention are administered intratu morally. In another embodiment, all of the therapeutic agents are administered systemically, e.g. intravenously. In an alternative embodiment, all of the therapeutic agents are administered intratumorally. In any of the embodiments, e.g. in this paragraph, the therapeutic agents of the invention may be administered as one or more pharmaceutical compositions.

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered via intravenous (IV) infusion. In one embodiment, the chemotherapeutic agent is administered via IV infusion.

In one embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered concurrently and/or sequentially with a chemotherapeutic agent. In another embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered concurrently with a chemotherapeutic agent. In another embodiment, the agonist ICOS binding protein or antigen binding portion thereof is administered sequentially with a chemotherapeutic agent.

In one embodiment, the therapeutic agent(s) are administered once every 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 15 days, 16 days, 17 days, 18 days, 19 days, 20 days, 21 days, 22 days, 23 days, 24 days, 25 days, 26 days, 27 days, 28 days, 29 days, 30 days, 31 days, 32 days, 33 days, 34 days, 35 days, 36 days, 37 days, 38 days, 39 days, or 40 days.

In one embodiment, the therapeutic agent(s) are administered once every 1-6 weeks. In one embodiment, the therapeutic agent(s) are administered once every 1 week, once every 2 weeks, once every 3 weeks, once every 4 weeks, once every 5 weeks or once every 6 weeks. In one embodiment, the therapeutic agent(s) are administered once every 3 weeks. In one embodiment, the therapeutic agent(s) are administered once every 6 weeks. In one embodiment, the combination is administered once every 3 weeks for 2-6 dosing cycles (e.g. the first 3, 4, or 5 dosing cycles, in particular, the first 4 dosing cycles). In one embodiment, the therapeutic agent(s) are administered for up to 2 years or unacceptable toxcity. In one embodiment, the therapeutic agent(s) are administered every three weeks up to 35 cycles or unacceptable toxicity.

If desired, the effective daily dose of a (therapeutic) combination may be administered as two, three, four, five, six or more doses administered separately at appropriate intervals throughout the day, optionally, in unit dosage forms.

The present disclosure provides methods of treating cancer comprising administering to a patient in need of treatment one or both of the therapeutic agents at a first dose at a first interval for a first period; and administering to the patient one or both of the therapeutic agents at a second dose at a second interval for a second period. There may be a rest period between the first and second periods in which one or both of the binding proteins in the combination are not administered to the patient. In some embodiments, there is a rest period between the first period and second period. In some embodiments, the rest period is between 1 and 30 days. In some embodiments, the rest period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28,

29, 30 or 31 days. In some embodiments, the rest period is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 13 weeks, 14 weeks or 15 weeks.

In some embodiments, the first dose and second dose are the same. In some embodiments, the first interval and second interval are the same. In some embodiments, the first interval and the second interval are once every three weeks. In some embodiments, the first interval and the second interval are once every six weeks. In some embodiments, the first interval and the second interval are different. In some embodiments, the first interval is once every three weeks and the second interval is once every six weeks.

In some embodiments, the first interval and the second interval are different. In some embodiments, the first interval is once every three weeks and the second interval is once every six weeks. In some embodiments, combination is administered at the first dose of 24 mg once every three weeks for the first period of 2-6 dosing cycles (e.g. the first 3, 4, or 5 dosing cycles, in particular, the first 4 dosing cycles), and at the second dose of 80 mg once every six weeks until therapy is discontinued (e.g. due to disease progression, an adverse event, or as determined by a physician).

In some embodiments, the therapeutic agent(s) described herein are administered according to dosing regimens demonstrated to achieve a clinical benefit for the patient. In some embodiments, a clinical benefit is stable disease ("SD"), a partial response ("PR") and/or a complete response ("CR"). In some embodiments, a clinical benefit is stable disease ("SD"). In some embodiments, a clinical benefit is a partial response ("PR"). In some embodiments, a clinical benefit is a complete response ("CR"). In some embodiments, PR or CR is determined in accordance with Response Evaluation Criteria in Solid Tumors (RECIST). In some embodiments, the combination is administered for a longer period to maintain clinical benefit. In one aspect of the invention, there is provided a pharmaceutical kit comprising about 0.08 mg to about 1000 mg of an agonist ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent.

In one embodiment, the pharmaceutical kit comprises about 0.08 mg to about 240 mg of the agonist ICOS binding protein or antigen binding portion thereof and the chemotherapeutic agent.

In some embodiments, the pharmaceutical kit comprises a further immunomodulatory agent. In one embodiment, the further immunomodulatory agent is a PD-1 binding protein or an antigen binding portion thereof, a PD-L1 binding protein or an antigen binding portion thereof, or a CTLA-4 binding protein or an antigen binding portion thereof.

In one embodiment, the pharmaceutical kit comprises the agonist ICOS binding protein at a concentration of 10 mg/mL.

In one embodiment, the pharmaceutical kit comprises cisplatin at a concentration of 1 mg/ml. In one embodiment, the pharmaceutical kit comprises carboplatin at a concentration of 10 mg/ml. In one embodiment, the pharmaceutical kit comprises paclitaxel at a concentration of 6 mg/ml. In one embodiment, the pharmaceutical kit comprises pemetrexed as a 100 mg or 500 mg powder for solution. In one embodiment, the pharmaceutical kit comprises pemetrexed at a concentration of 25 mg/ml. In one embodiment, the pharmaceutical kit comprises gemcitabine at a concentration of 10 mg/ml, 38 mg/ml or 100 mg/ml. In one embodiment, the pharmaceutical kit comprises gemcitabine as a 200 mg, 1000 mg or 2g powder for solution. In one embodiment, the pharmaceutical kit comprises docetaxel at a concentration of 10 mg/ml or 20 mg/ml. In one embodiment, the pharmaceutical kit comprises 5-FU at a concentration of 25 mg/ml or 50 mg/ml.

In one embodiment, the pharmaceutical kit comprises the PD-1 binding protein at a concentration of about 20 mg/mL to about 125 mg/mL. In a further embodiment, the pharmaceutical kit comprises the PD-1 binding protein at a concentration of 20 mg/mL to 50 mg/mL. In one embodiment, the PD-1 binding protein is at a concentration of 20 mg/mL. In another embodiment, the PD-1 binding protein is at a concentration of 50 mg/mL.

Cancer

The therapeutic agent(s) and methods of the invention may be used in the treatment of cancer.

By the term "treating" and grammatical variations thereof as used herein, is meant therapeutic therapy. In reference to a particular condition, treating means: (1) to ameliorate, or lessen the severity of, the condition of one or more of the biological manifestations of the condition, (2) to interfere with (a) one or more points in the biological cascade that leads to or is responsible for the condition or (b) one or more of the biological manifestations of the condition, (3) to alleviate one or more of the symptoms or signs, effects or side effects associated with the condition or treatment thereof, (4) to slow the progression of the condition, that is to say prolong survival, or one or more of the biological manifestations of the condition and/or (5) to cure said condition or one or more of the biological manifestations of the condition by eliminating or reducing to undetectable levels one or more of the biological manifestations of the condition for a period of time considered to be a state of remission for that manifestation without additional treatment over the period of remission. One skilled in the art will understand the duration of time considered to be remission for a particular disease or condition. Prophylactic therapy is also contemplated thereby. The skilled artisan will appreciate that "prevention" is not an absolute term. In medicine, "prevention" is understood to refer to the prophylactic administration of a drug to substantially diminish the likelihood or severity of a condition or biological manifestation thereof, or to delay the onset of such condition or biological manifestation thereof. Prophylactic therapy is appropriate, for example, when a subject is considered at high risk for developing cancer, such as when a subject has a strong family history of cancer or when a subject has been exposed to a carcinogen.

As used herein, the terms "cancer", "neoplasm", "malignancy", and "tumor" are used interchangeably and, in either the singular or plural form, refer to cells that have undergone a malignant transformation that makes them pathological to the host organism. Primary cancer cells can be readily distinguished from non-cancerous cells by well-established techniques, particularly histological examination. The definition of a cancer cell, as used herein, includes not only a primary cancer cell, but any cell derived from a cancer cell ancestor. This includes metastasized cancer cells, and in vitro cultures and cell lines derived from cancer cells. When referring to a type of cancer that normally manifests as a solid tumor, a "clinically detectable" tumor is one that is detectable on the basis of tumor mass; e.g. by procedures such as computed tomography (CT) scan, magnetic resonance imaging (MRI), X-ray, ultrasound or palpation on physical examination, and/or which is detectable because of the expression of one or more cancer-specific antigens in a sample obtainable from a patient. Tumors may be a hematopoietic (or hematologic or hematological or blood-related) cancer, for example, cancers derived from blood cells or immune cells, which may be referred to as "liquid tumors." Specific examples of clinical conditions based on hematologic tumors include leukemias such as chronic myelocytic leukemia, acute myelocytic leukemia, chronic lymphocytic leukemia and acute lymphocytic leukemia; plasma cell malignancies such as multiple myeloma, MGUS and Waldenstrom's macroglobulinemia; lymphomas such as non-Hodgkin's lymphoma, Hodgkin's lymphoma; and the like.

The cancer may be any cancer in which an abnormal number of blast cells or unwanted cell proliferation is present or that is diagnosed as a hematological cancer, including both lymphoid and myeloid malignancies. Myeloid malignancies include, but are not limited to, acute myeloid (or myelocytic or myelogenous or myeloblastic) leukemia (undifferentiated or differentiated), acute promyeloid (or promyelocytic or promyelogenous or promyeloblastic) leukemia, acute myelomonocytic (or myelomonoblastic) leukemia, acute monocytic (or monoblastic) leukemia, erythroleukemia and megakaryocytic (or megakaryoblastic) leukemia. These leukemias may be referred together as acute myeloid (or myelocytic or myelogenous) leukemia (AML). Myeloid malignancies also include myeloproliferative disorders (MPD) which include, but are not limited to, chronic myelogenous (or myeloid) leukemia (CML), chronic myelomonocytic leukemia (CMML), essential thrombocythemia (or thrombocytosis), and polcythemia vera (PCV). Myeloid malignancies also include myelodysplasia (or myelodysplastic syndrome or MDS), which may be referred to as refractory anemia (RA), refractory anemia with excess blasts (RAEB), and refractory anemia with excess blasts in transformation (RAEBT); as well as myelofibrosis (MFS) with or without angiogenic myeloid metaplasia.

Hematopoietic cancers also include lymphoid malignancies, which may affect the lymph nodes, spleens, bone marrow, peripheral blood, and/or extranodal sites. Lymphoid cancers include B-cell malignancies, which include, but are not limited to, B-cell non-Hodgkin's lymphomas (B-NHLs). B- NHLs may be indolent (or low-grade), intermediate-grade (or aggressive) or high-grade (very aggressive). Indolent Bcell lymphomas include follicular lymphoma (FL); small lymphocytic lymphoma (SLL); marginal zone lymphoma (MZL) including nodal MZL, extranodal MZL, splenic MZL and splenic MZL with villous lymphocytes; lymphoplasmacytic lymphoma (LPL); and mucosa-associated-lymphoid tissue (MALT or extranodal marginal zone) lymphoma. Intermediate-grade B-NHLs include mantle cell lymphoma (MCL) with or without leukemic involvement, diffuse large cell lymphoma (DLBCL), follicular large cell (or grade 3 or grade 3B) lymphoma, and primary mediastinal lymphoma (PML). High-grade B-NHLs include Burkitt's lymphoma (BL), Burkitt-like lymphoma, small non-cleaved cell lymphoma (SNCCL) and lymphoblastic lymphoma. Other B-NHLs include immunoblastic lymphoma (or immunocytoma), primary effusion lymphoma, HIV associated (or AIDS related) lymphomas, and post-transplant lymphoproliferative disorder (PTLD) or lymphoma. B-cell malignancies also include, but are not limited to, chronic lymphocytic leukemia (CLL), prolymphocytic leukemia (PLL), Waldenstrom's macroglobulinemia (WM), hairy cell leukemia (HCL), large granular lymphocyte (LGL) leukemia, acute lymphoid (or lymphocytic or lymphoblastic) leukemia, and Castleman's disease. NHL may also include T-cell non-Hodgkin's lymphoma s(T-NHLs), which include, but are not limited to T- cell non-Hodgkin's lymphoma not otherwise specified (NOS), peripheral T-cell lymphoma (PTCL), anaplastic large cell lymphoma (ALCL), angioimmunoblastic lymphoid disorder (AILD), nasal natural killer (NK) cell / T-cell lymphoma, gamma/delta lymphoma, cutaneous T cell lymphoma, mycosis fungoides, and Sezary syndrome.

Hematopoietic cancers also include Hodgkin's lymphoma (or disease) including classical Hodgkin's lymphoma, nodular sclerosing Hodgkin's lymphoma, mixed cellularity Hodgkin's lymphoma, lymphocyte predominant (LP) Hodgkin's lymphoma, nodular LP Hodgkin's lymphoma, and lymphocyte depleted Hodgkin's lymphoma. Hematopoietic cancers also include plasma cell diseases or cancers such as multiple myeloma (MM) including smoldering MM, monoclonal gammopathy of undetermined (or unknown or unclear) significance (MGUS), plasmacytoma (bone, extramedullary), lymphoplasmacytic lymphoma (LPL), Waldenstrom's Macroglobulinemia, plasma cell leukemia, and primary amyloidosis (AL). Hematopoietic cancers may also include other cancers of additional hematopoietic cells, including polymorphonuclear leukocytes (or neutrophils), basophils, eosinophils, dendritic cells, platelets, erythrocytes and natural killer cells. Tissues which include hematopoietic cells referred herein to as "hematopoietic cell tissues" include bone marrow; peripheral blood; thymus; and peripheral lymphoid tissues, such as spleen, lymph nodes, lymphoid tissues associated with mucosa (such as the gut-associated lymphoid tissues), tonsils, Peyer's patches and appendix, and lymphoid tissues associated with other mucosa, for example, the bronchial linings.

In some embodiments, the treatment of cancer is first-line treatment of cancer. In one embodiment, the treatment of cancer is second-line treatment of cancer. In some embodiments, the treatment is third-line treatment of cancer. In some embodiments, the treatment is fourth-line treatment of cancer. In some embodiments, the treatment is fifth-line treatment of cancer. In some embodiments, prior treatment to said second-line, third-line, fourth-line or fifth-line treatment of cancer comprises one or more of radiotherapy, chemotherapy, surgery or radiochemotherapy.

In one embodiment, the cancer is selected from: brain cancer, glioblastomas, glioma (such as diffuse intrinsic pontine glioma), Bannayan-Zonana syndrome, Cowden disease, Lhermitte-Duclos disease, breast cancer ( e.g . inflammatory breast cancer), Wilm's tumor, ependymoma, medulloblastoma, cardiac tumors, colon cancer, colorectal cancer, head and neck cancer (e.g. squamous cell carcinoma of the head and neck, cancer of the mouth (/.e. oral cancer), salivary gland cancer, buccal cancer, pharyngeal cancer, oropharyngeal cancer, nasopharangeal cancer, hypopharyngeal cancer, laryngeal cancer), eye cancer (e.g. retinoblastoma), lung cancer (e.g. nonsmall cell lung cancer, small cell cancer), liver cancer (/.e. hepatocellular cancer), skin cancer (e.g. basal cell carcinoma, merkel cell carcinoma, squamous cell carcinoma), melanoma, ovarian cancer, pancreatic cancer, bile duct cancer, gallbladder cancer, prostate cancer, sarcoma (e.g. soft tissue sarcoma, Ewing's sarcoma, Kaposi sarcoma, rhabdomyosarcoma), bone cancer, osteosarcoma, giant cell tumor of bone, thyroid cancer, parathyroid cancer, thymoma, blood cancer (which may be broadly categorised as leukemias, lymphomas or myelomas, and include examples such as lymphoblastic T- cell leukemia, chronic myelogenous leukemia, chronic lymphocytic leukemia, hairy-cell leukemia, acute lymphoblastic leukemia, acute myelogenous leukemia, chronic neutrophilic leukemia, acute lymphoblastic T-cell leukemia, plasmacytoma, immunoblastic large cell leukemia, mantle cell leukemia, multiple myeloma megakaryoblastic leukemia, multiple myeloma, acute megakaryocytic leukemia, promyelocytic leukemia, erythroleukemia, malignant lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymphoblastic T cell lymphoma, Burkitt's lymphoma, and follicular lymphoma), neuroblastoma, pituitary tumor, adrenocortical cancer, anal cancer (/.e. rectal cancer), bladder cancer, urothelial cancer, urethral cancer, vaginal cancer, vulvar cancer, cervical cancer, endometrial cancer, uterine cancer, fallopian tube cancer, renal cancer (/.e. kidney cancer, e.g. renal cell carcinoma), mesothelioma (e.g. malignant pleural mesothelioma), esophageal cancer (e.g. esophageal squamous cell carcinoma), gastric cancer ( i.e . stomach cancer), gastrointestinal carcinoid tumor, GIST (gastrointestinal stromal tumor), appendiceal cancer, penile cancer, testicular cancer, germ cell tumors.

In one embodiment, the cancer exhibits microsatellite instability (MSI). Microsatellite instability ("MSI") is or comprises a change that in the DNA of certain cells (such as tumor cells) in which the number of repeats of microsatellites (short, repeated sequences of DNA) is different than the number of repeats that was contained in the DNA from which it was inherited. Microsatellite instability arises from a failure to repair replication-associated errors due to a defective DNA mismatch repair (MMR) system. This failure allows persistence of mismatch mutations all over the genome, but especially in regions of repetitive DNA known as microsatellites, leading to increased mutational load. It has been demonstrated that at least some tumors characterized by MSI-H have improved responses to certain anti-PD-1 agents (Le etal. (2015) N. Engl. J. Med. 372(26):2509-2520; Westdorp etal. (2016) Cancer Immunol. Immunother. 65(10): 1249-1259).

In some embodiments, a cancer has a microsatellite instability status of high microsatellite instability (e.g. MSI-H status). In some embodiments, a cancer has a microsatellite instability status of low microsatellite instability (e.g. MSI-L status). In some embodiments, a cancer has a microsatellite instability status of microsatellite stable (e.g. MSS status). In some embodiments microsatellite instability status is assessed by a next generation sequencing (NGS)-based assay, an immunohistochemistry (IHC)-based assay, and/or a PCR-based assay. In some embodiments, microsatellite instability is detected by NGS. In some embodiments, microsatellite instability is detected by IHC. In some embodiments, microsatellite instability is detected by PCR.

In some embodiments, the cancer is associated with a high tumor mutation burden (TMB). In some embodiments, the cancer is associated with high TMB and MSI-H. In some embodiments, the cancer is associated with high TMB and MSI-L or MSS. In some embodiments, the cancer is endometrial cancer associated with high TMB. In some related embodiments, the endometrial cancer is associated with high TMB and MSI-H. In some related embodiments, the endometrial cancer is associated with high TMB and MSI-L or MSS.

In some embodiments, a cancer is a mismatch repair deficient (dMMR) cancer. Microsatellite instability may arise from a failure to repair replication-associated errors due to a defective DNA mismatch repair (MMR) system. This failure allows persistence of mismatch mutations all over the genome, but especially in regions of repetitive DNA known as microsatellites, leading to increased mutational load that may improve responses to certain therapeutic agents.

In some embodiments, a cancer is a hypermutated cancer. In some embodiments, a cancer harbors a mutation in polymerase epsilon (POLE). In some embodiments, a cancer harbors a mutation in polymerase delta (POLD). In some embodiments, the cancer is an advanced cancer. In some embodiments, the cancer is a metastatic cancer. In some embodiments, the cancer is a recurrent cancer ( e.g . a recurrent gynaecological cancer such as recurrent epithelial ovarian cancer, recurrent fallopian tube cancer, recurrent primary peritoneal cancer, or recurrent endometrial cancer). In one embodiment, the cancer is recurrent or advanced.

In some embodiments, the cancer is current/metastatic (R/M). In some embodiments, the cancer is recurring/refectory (R/R).

In another embodiment the human has a liquid tumor such as diffuse large B cell lymphoma (DLBCL), multiple myeloma, chronic lymphoblastic leukemia, follicular lymphoma, acute myeloid leukemia and chronic myelogenous leukemia.

In one embodiment, the human has a solid tumor. In one embodiment, the solid tumor is advanced solid tumor. In one embodiment, the cancer is selected from head and neck cancer, squamous cell carcinoma of the head and neck (SCCHN or HNSCC), gastric cancer, melanoma, mesothelioma, renal cell carcinoma (RCC), esophageal cancer, non-small cell lung carcinoma (NSCLC), prostate cancer, esophageal cancer, esophageal squamous cell carcinoma, colorectal cancer, cervical cancer, bladder cancer, urothelial cancer, ovarian cancer and pancreatic cancer. In one embodiment, the human has one or more of the following: HNSCC, colorectal cancer, esophageal cancer, cervical cancer, bladder cancer, breast cancer, head and neck cancer, ovarian cancer, melanoma, renal cell carcinoma (RCC), esophageal squamous cell carcinoma, non-small cell lung carcinoma, mesothelioma (e.g. pleural malignant mesothelioma), urothelial cancer and prostate cancer.

In one embodiment, the cancer is head and neck cancer. In one embodiment, the cancer is HNSCC. Squamous cell carcinoma is a cancer that arises from particular cells called squamous cells. Squamous cells are found in the outer layer of skin and in the mucous membranes, which are the moist tissues that line body cavities such as the airways and intestines. Head and neck squamous cell carcinoma (HNSCC) develops in the mucous membranes of the mouth, nose, and throat. HNSCC can occur in the mouth (oral cavity), the middle part of the throat near the mouth (oropharynx), the space behind the nose (nasal cavity and paranasal sinuses), the upper part of the throat near the nasal cavity (nasopharynx), the voicebox (larynx), or the lower part of the throat near the larynx (hypopharynx). Depending on the location, the cancer can cause abnormal patches or open sores (ulcers) in the mouth and throat, unusual bleeding or pain in the mouth, sinus congestion that does not clear, sore throat, earache, pain when swallowing or difficulty swallowing, a hoarse voice, difficulty breathing, or enlarged lymph nodes. HNSCC can metastasize to other parts of the body, such as the lymph nodes, lungs or liver.

Tobacco use and alcohol consumption are the two most important risk factors for the development of HNSCC, and their contributions to risk are synergistic. In addition, the human papillomavirus (HPV), especially HPV-16, is now a well-established independent risk factor. Patients with HNSCC have a relatively poor prognosis. Recurrent/metastatic (R/M) HNSCC is especially challenging, regardless of human papillomavirus (HPV) status, and currently, few effective treatment options are available in the art. HPV-negative HNSCC is associated with a locoregional relapse rate of 19-35% and a distant metastatic rate of 14-22% following standard of care, compared with rates of

9-18% and 5-12%, respectively, for HPV-positive HNSCC. The median overall survival for patients with R/M disease is 10-13 months in the setting of first-line chemotherapy and 6 months in the second-line setting. The current standard of care is platinum-based doublet chemotherapy with or without cetuximab. Second-line standard of care options include cetuximab, methotrexate, and taxanes. All of these chemotherapeutic agents are associated with significant side effects, and only

10-13% of patients respond to treatment. HNSCC regressions from existing systemic therapies are transient and do not add significantly increased longevity, and virtually all patients succumb to their malignancy.

In one embodiment, the cancer is recurrent/metastatic (R/M) HNSCC. In one embodiment, the cancer is recurring/refractory (R/R) HNSCC. In one embodiment, the cancer is HPV-negative or HPV-positive HNSCC. In one embodiment, the cancer is a locally advanced HNSCC. In one embodiment, the cancer is (R/M) HNSCC in PD-L1 CPS (Combined Positive Score) positive (CPS ³1) patients. The combined positive score is as determined by an FDA-approved test. PD-L1 CPS is the number of PD-L1 staining cells (tumor cells, lymphocytes, macrophages) divided by the total number of viable tumor cells, multiplied by 100. In one embodiment, PD-L1 CPS is determined using PharmDx 22C3. In one embodiment, the cancer is HNSCC in PD-1 binding protein/PD-Ll binding protein experienced or PD-1 binding protein/PD-Ll binding protein nna^'ive patients. In one embodiment, the cancer is HNSCC in PD-1 binding protein/PD-Ll binding protein experienced or PD-1 binding protein/PD-Ll binding protein na^'ive patients.

In one embodiment, the head and neck cancer is oropharyngeal cancer. In one embodiment, the head and neck cancer is an oral cancer {i.e. a mouth cancer).

In one embodiment, the treatment is first-line or second line treatment of HNSCC. In one embodiment, the treatment is first-line or second line treatment of recurrent/metastatic HNSCC. In one embodiment the treatment is first line treatment of recurrent/ metastatic (1L R/M) HNSCC. In one embodiment, the treatment is first line treatment of 1L R/M HNSCC in a PD-L1 CPS (combined positive score) positive (CPS ³1) patients. In one embodiment the treatment is second line treatment of recurrent/metastatic (2L R/M) HNSCC.

In one embodiment, the treatment is first-line, second-line, third-line, fourth-line or fifth-line treatment of PD-l/PD-Ll-naTve HNSCC. In one embodiment, the treatment first-line, second-line, third-line, fourth-line or fifth-line treatment of PD-1/PD-L1 experienced HNSCC.

In one embodiment, the cancer is lung cancer. In some embodiments, the lung cancer is a squamous cell carcinoma of the lung. In some embodiments, the lung cancer is small cell lung cancer (SCLC). In some embodiments, the lung cancer is non-small cell lung cancer (NSCLC), such as squamous NSCLC. In some embodiments, the lung cancer is an ALK-translocated lung cancer ( e.g . ALK-translocated NSCLC). In some embodiments, the cancer is NSCLC with an identified ALK translocation. In some embodiments, the lung cancer is an EGFR-mutant lung cancer ( e.g . EGFR- mutant NSCLC). In some embodiments, the cancer is NSCLC with an identified EGFR mutation. In one embodiment, the cancer is advanced NSCLC. In another embodiment, the cancer is relapsed/refractory advanced NSCLC.

In one embodiment, the cancer is melanoma. In some embodiments, the melanoma is an advanced melanoma. In some embodiments, the melanoma is a metastatic melanoma. In some embodiments, the melanoma is an MSI-H melanoma. In some embodiments, the melanoma is a MSS melanoma. In some embodiments, the melanoma is a POLE-mutant melanoma. In some embodiments, the melanoma is a POLD-mutant melanoma. In some embodiments, the melanoma is a high TMB melanoma.

In one embodiment, the cancer is urothelial cancer. In some embodiments, the urothelial cancer is an advanced urothelial cancer. In some embodiments, the urothelial cancer is a metastatic urothelial cancer. In some embodiments, the urothelial cancer is a MSI-H urothelial cancer. In some embodiments, the urothelial cancer is a MSS urothelial cancer. In some embodiments, the urothelial cancer is a POLE-mutant urothelial cancer. In some embodiments, the urothelial cancer is a POLD- mutant urothelial cancer. In some embodiments, the urothelial cancer is a high TMB urothelial cancer.

In one embodiment of the method of treating cancer in a human in need thereof, the method comprises administering to the human an agonist ICOS binding protein or antigen binding portion thereof at a dose of about 24 mg once every 3 weeks and administering to the human a fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and carboplatin at a dose of about AUC 4-6 mg/ml once every 3 weeks.

In one embodiment of the method of treating cancer in a human in need thereof, the method comprises administering to the human an agonist ICOS binding protein or antigen binding portion thereof at a dose of about 24 mg once every 3 weeks and administering to the human a fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and cisplatin at a dose of about 100 mg/m² once every 3 weeks.

In one embodiment of the method of treating cancer in a human in need thereof, the method comprises administering to the human an agonist ICOS binding protein or antigen binding portion thereof at a dose of about 24 mg once every 3 weeks and administering to the human a chemotherapeutic agent, wherein the cancer is HNSCC that expresses PD-L1 at a combined positive score (CPS) ³1, and wherein the chemotherapeutic agent is fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and cisplatin at a dose of about 100 mg/m² once every 3 weeks, or fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and carboplatin at a dose of about AUC 4-6 mg/ml once every 3 weeks. In one embodiment of the agonist ICOS binding protein or antigen binding portion thereof for use in treating cancer, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg once every 3 weeks and is administered with fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and carboplatin at a dose of about AUC 4-6 mg/ml once every 3 weeks.

In one embodiment of the agonist ICOS binding protein or antigen binding portion thereof for use in treating cancer, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg once every 3 weeks and is administered with fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and cisplatin at a dose of about 100 mg/m² once every 3 weeks.

In one embodiment of the agonist ICOS binding protein or antigen binding portion thereof for use in treating cancer, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg once every 3 weeks and is administered with a chemotherapeutic agent, wherein the cancer is HNSCC that expresses PD-L1 at a combined positive score (CPS) ³1, and wherein the chemotherapeutic agent is fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and cisplatin at a dose of about 100 mg/m² once every 3 weeks, or fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and carboplatin at a dose of about AUC 4-6 mg/ml once every 3 weeks.

In one embodiment of the combination of an agonist ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent for use in treating cancer, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg once every 3 weeks and is administered with fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and carboplatin at a dose of about AUC 4-6 mg/ml once every 3 weeks.

In one embodiment of the combination of an agonist ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent for use in treating cancer, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg once every 3 weeks and is administered with fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and cisplatin at a dose of about 100 mg/m² once every 3 weeks.

In one embodiment of the combination of an agonist ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent for use in treating cancer, the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg once every 3 weeks and is administered with a chemotherapeutic agent, wherein the cancer is HNSCC that expresses PD-L1 at a combined positive score (CPS) ³1, and wherein the chemotherapeutic agent is fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and cisplatin at a dose of about 100 mg/m² once every 3 weeks, or fluorouracil at a dose of about 1000 mg/m²/day on Day 1 through Day 4 of a 21 day cycle and carboplatin at a dose of about AUC 4-6 mg/ml once every 3 weeks. In one embodiment of the composition comprising an agonist ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent, the agonist ICOS binding protein or antigen binding portion thereof is comprised at about 0.08 mg to about 240 mg. In one embodiment, the agonist ICOS binding protein or antigen binding protein thereof is comprised at a dose of 0.08 mg, 0.24 mg, 0.8 mg, 2.4 mg, 8 mg, 24 mg, 48 mg, 80 mg, 160 mg or 240 mg in particular 24 mg, 48 mg, 80 mg or 160 mg. In one embodiment, In one embodiment, the chemotherapeutic agent is one or more of docetaxel comprised at a dose of about 30 to about 100 mg/m² or about 75 mg/m²; carboplatin comprised at a dose of about AUC 4-7 mg/ml per min or about AUC 5 mg/ml per min; cisplatin comprised at a dose of about 20 mg/m² to about 120 mg/m² or 100 mg/m²; paclitaxel comprised at a dose of about 135 mg/m² to about 225 mg/m² or 200 mg/m²; fluorouracil comprised at a dose of about 200 mg/m² to about 1200 mg/m² or 1000 mg/m²; pemetrexed comprised at a dose of about 500 mg/m²; or gemcitabine comprised at a dose of about 1000 mg/m² to about 1250 mg/m² or 1250 mg/m². In some embodiments, the agonist ICOS binding protein is comprised at about 0.001 mg/kg to 10 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 0.001 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 0.003 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 0.01 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 0.03 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 0.1 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 0.3 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 0.6 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 1 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 2 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 3 mg/kg. In some embodiments, the agonist ICOS binding protein or antigen binding protein thereof is comprised at about 4 mg/kg; about 5 mg/kg; about 6 mg/kg; about 7 mg/kg; about 8 mg/kg; about 9 mg/kg or about 10 mg/kg. In one embodiment, In one embodiment, the chemotherapeutic agent is one or more of docetaxel comprised at a dose of about 30 to about 100 mg/m² or about 75 mg/m²; carboplatin comprised at a dose of about AUC 4-7 mg/ml per min or about AUC 5 mg/ml per min; cisplatin comprised at a dose of about 20 mg/m² to about 120 mg/m² or 100 mg/m²; paclitaxel comprised at a dose of about 135 mg/m² to about 225 mg/m² or 200 mg/m²; fluorouracil comprised at a dose of about 200 mg/m² to about 1200 mg/m² or 1000 mg/m²; pemetrexed comprised at a dose of about 500 mg/m²; or gemcitabine comprised at a dose of about 1000 mg/m² to about 1250 mg/m² or 1250 mg/m². The following examples are intended for illustration only, and are not intended to limit the scope of the invention in any way.

EXAMPLES Example 1

An experimental study was carried out to determine the efficacy of anti-mouse ICOS agonist antibody alone and in combination with carboplatin in CT26 murine colon carcinoma model in female BALB/c mice. The study was conducted under a protocol which was approved by the GSK Institutional Animal Care and Use Committee prior to commencement of the study.

Materials and Methods Mice

Female BALB/c mice (BALB/cAnNCrl, Charles River) were seven weeks old and had a body weight range of 15.2 g to 20.1 g on Day 1.

Tumour Cell Culture

The CT26 murine colon carcinoma cell line was obtained from ATCC (CRL-2638) and maintained in RPMI-1640 medium containing 10% fetal bovine serum, 2 mM glutamine, 100 units/mL penicillin G sodium, lOOmg/mL streptomycin sulfate, and 25 mg/mL gentamicin. Cell cultures were maintained in tissue culture flasks in a humidified incubator at 37°C, in an atmosphere of 5% CO2 and 95% air.

Tumour Implantation and Measurement

The CT26 tumor cells were harvested during exponential growth, and resuspended in cold PBS. Each mouse was inoculated subcutaneously in the right flank with 3xl0⁵ cells (0.1 mL in suspension). Tumors were calipered in two dimensions to monitor growth as their mean volume approached the desired 80-120 mm³ range. Tumor size was calculated using the formula:

Tumour Volume (mm³) = (w² x 1)12 where w = width and / = length, in mm, of the tumor. Tumor weight may be estimated with the assumption that 1 mg is equivalent to 1 mm³ of tumor volume.

Ten days after tumor implantation, which was designated as Day 1 of the study, animals with individual tumor volumes ranging from 63-196 mm³ were sorted into fourteen groups (n=10/group) with group mean tumor volume of 109-113 mm³. Test agents

Rat IgG2b isotype and Rat anti-mouse ICOS IgG2b antibody (clone 7E.17G9) were stored at 4°C protected from light.

Carboplatin (Teva Pharmaceuticals) was stored at room temperature.

Anti-mouse ICOS (GSK) antibody dosing solution was prepared by diluting an aliquot of the stock (5.94 mg/mL) to 0.005, 0.025, 0.1, 0.5 and 1 mg/mL in sterile PBS, which provided 1, 5, 20, 100 and 200 mg/mouse, respectively, in a dosing volume of 200 pL/ mouse.

Rat IgG2b antibody (isotype) dosing solution was prepared by diluting an aliquot of the stock (7.32 mg/mL) to 0.5 and 1 mg/mL in sterile PBS, which provided 100 and 200 mg/mouse, respectively, in a dosing volume of 200 pL/mouse.

Carboplatin stock solution (10 mg/mL) was diluted on each day of dosing with 5% dextrose in water (D5W) to yield a 7.5 mg/mL solution, providing a 75 mg/kg dosage in a 10 mL/kg dosing volume. Solutions were stored at room temperature and doses were adjusted to the body weight of the individual animals.

Treatment

On day 1 of the study, BALB/c mice bearing established CT26 tumors were sorted into fourteen groups. Dosing was initiated according to the treatment plan summarized in Table 2. All agents were administered intra peritonea lly (i.p.). On Days 1 and 8 carboplatin therapy was immediately followed by the antibody regimen.

Group 1 mice served as control group for tumor growth and statistical analysis and received no treatment.

Groups 2 and 3 received vehicle (D5W), once a week for two weeks (qwk x 2) and rat IgG2b at 100 and 200 mg/mouse on Days 1, 4 and 8.

Group 4 received carboplatin at 75 mg/kg and rat IgG2b-GSK at 100 mg/mouse on Days 1, 4 and 8.

Groups 5 - 9 received vehicle, qwk x 2 and anti-mouse ICOS, clone GSK, at 1, 5, 20, 100 and 200 mg/mouse, respectively, on Days 1, 4 and 8.

Groups 10 - 14 received carboplatin at 75 mg/kg, qwk x 2 in combination with anti-mouse ICOS at 1, 5, 20, 100 and 200 mg/mouse, respectively, on Days 1, 4 and 8. Table 2

Table 2 displays the study design as of Day 1 of the study.

Vehicle = D5W ^a - mg/ kg

Tumor Growth Delay

Animals were monitored individually for tumor growth until Day 43. Each animal was euthanized for tumor progression (TP) when its tumor reached the 1500 mm³ volume endpoint. The time to endpoint (TTE) for each mouse was calculated with the following equation:

TTE = (logio (endpoint volume)-b)/m where b is the intercept and m is the slope of the line obtained by linear regression of a log- transformed tumor growth data set. The data set is comprised of the first observation that exceeded the study endpoint volume and the three consecutive observations that immediately preceded the attainment of the endpoint volume. Any animal that did not reach end point was euthanized at the end of the study and assigned a TTE equal to the last study day (Day 43). In instances in which the log-transformed calculated TTE preceded the day prior to reaching endpoint or exceeded the day of reaching tumor volume endpoint, a linear interpolation was performed to approximate TTE. Any animal determined to have died from treatment related (TR) causes was to be assigned a TTE value equal to the day of death. Any animal that died from non-treatment-related (NTR) causes was to be excluded from the analysis.

Treatment outcome was evaluated from tumor growth delay (TGD), which was defined as the increase in the median TTE for a treatment group compared to the control group:

TGD = T - C expressed in days, or as a percentage of the median TTE of the control group:

% TGD = ((T - C) /C) x 100 where

T = median TTE for a treatment group,

C = median TTE for the control group.

MTV and Criteria for Regression Responses

Treatment efficacy was also determined from the tumor volumes of animals remaining in the study on the last day and from the number of regression responses. The MTV(n) (Median Tumor Volume (mm³) for the number of animals on the Day of TGD analysis (excludes animals with tumor volume at the endpoint)) is defined as the median tumor volume on Day 43 in the number of animals remaining, n, whose tumors have not attained the endpoint volume.

Treatment may cause partial regression (PR) or complete regression (CR) of the tumor in an animal. In a PR response, the tumor volume is 50% or less of its Day 1 volume for three consecutive measurements during the course of the study, and equal to or greater than 13.5 mm³ for one or more of these three measurements during the course of the study. Animals were scored only once during the study for a PR or CR event and only as CR if both PR and CR criteria were satisfied. All animals were monitored for regression responses. An animal with a CR response at the termination of the study is additionally classified as tumor-free survivor (TFS).

Toxicity

The mice were observed frequently for heath and overt signs of any adverse treatment-related (TR) side effects, and noteworthy clinical observations were recorded. Individual body weight loss was monitored and any animal whose weight exceeded the limits for acceptable body weight loss was euthanized. If group mean body weight recovered, dosing may resume in that group, but at a lower dose or less frequent dosing schedule. Acceptable toxicity was defined as a group mean BW loss of less than 20% during the study and not more than one TR death among ten treated animals, or 10%. Any dosing regimen resulting in greater toxicity is considered above the maximum tolerated does (MTD). A death was to be classified as TR if it was attributable to treatment side effects as evidenced by clinical signs and/or necropsy, or may also be classified as TR if due to unknown causes during the dosing period or within 14 days of the last dose. A death was classified as NTR if there was evidence that the death was related to the tumor model, rather than treatment-related. NTR deaths are further categorized as NTRa (due to accident or human error), NTRm (due to necropsy-confirmed tumor dissemination by invasion or metastasis), and NTRu (due to unknown causes).

Statistical and Graphical Analyses

Survival was analyzed by the Kaplan-Meier method. The logrank (Mantel-Cox) and Gehan- Breslow-Wilcoxon tests determined the significance of the difference between the overall survival experiences (survival curves) of the two groups, based on TTE values. The two-tailed statistical analyses were not adjusted for multiple comparisons, and were conducted at P = 0.05.

Prism 7.02 (GraphPad) was employed for statistical and graphical analyses. Prims reports results as non-significant (ns) at P = ³0.05, significant (symbolized by "*") at 0.01 £ P £ 0.05, very significant ("**") at 0.001 < P £ 0.01 and extremely significant ("***") at P £ 0.001.

Results

A dose titration of 1, 5, 20, 100, and 200 mg per mouse of rat anti-mouse ICOS surrogate antibody clone 7E.17G9 was assessed in combination with 75 mg/kg Carboplatin for tumor growth inhibition efficacy in the CT-26 murine syngeneic model. Both agents were administered intra- peritoneally (IP). At 100 mg per mouse, rat anti-mouse ICOS in combination with Carboplatin demonstrated a significant increase in survivability by Kaplan-Meier analysis when compared to the 100 mg/mouse ICOS monotherapy (Figure 1). There were no unexpected mouse deaths in this study.

This study demonstrates that the rat anti-mouse ICOS surrogate antibody clone 7E.17G9 dosed at 100 mg/mouse in combination with carboplatin results in statistically significant survival advantage compared to ICOS 100 mg/mouse monotherapy in the CT-26 syngeneic tumor model, indicating ICOS and carboplatin combinations may be beneficial for anti-tumor efficacy.

Example 2

An experimental study was carried out to determine the efficacy of anti-mouse ICOS (anti- mlCOS) agonist antibody alone and in combination with paclitaxel in CT26 murine colon carcinoma model in female BALB/c mice. The study was conducted under a protocol which was approved by the GSK Institutional Animal Care and Use Committee prior to commencement of the study.

Details of the Mice, Tumor Cell Culture, Tumor Implantation and Measurement, Tumor Growth Delay, MTV and Criteria for Regression Responses, Toxicity, and Statistical and Graphical Analyses are as Example 1.

Test agents

Rat anti-mouse ICOS IgG2b antibody dosing solution was prepared by diluting an aliquot of the stock (5.94 mg/mL) to 0.005, 0.025, 0.1, 0.5 and 1 mg/mL in sterile PBS, which provided 1, 5, 20, 100 and 200 mg/mouse, respectively, in a dosing volume of 200 pL/mouse.

Rat IgG2a antibody (isotype) dosing solution was prepared by diluting an aliquot of the stock (7.32 mg/mL) to 0.5 and 1 mg/mL in sterile PBS, which provided 100 and 200 mg/mouse, respectively, in a dosing volume of 200 pL/mouse.

Paclitaxel (Phyton Biotech, LLC) obtained as a dry powder was made into a stock solution of 10 mg/mL in 50% ethanol: 50% Cremophor EL (stock diluent) and was stored at room temperature protected from light during the dosing period. On each day of dosing, an aliquot of the paclitaxel was diluted into 10X stock solution diluent to a concentration of 30.0 mg/mL then further diluted with 10X stock solution diluent to a concentration of 24 mg/mL. This 24 mg/mL solution was further diluted in 5% dextrose in water (5DW) to yield 2.4 mg/mL paclitaxel dosing solution in a vehicle consisting of 5% ethanol:5% Cremophor EL:90% D5W (vehicle). The 2.4 mg/mL dosing solution provided the 24 mg/kg dosage in a dosing volume of 10 mL/kg, adjusted to the body weight of the animal.

Treatment

On day 1 of the study, BALB/c mice bearing established CT26 tumors were sorted into fourteen groups. Dosing was initiated according to the treatment plan summarized in Table 3.

Groups 2 and 3 received vehicle (5% ethanol:5% Cremophor EL in D5W) intravenously (i.v.), every four days for three doses (q4d x3) and rat IgG2a-GSK intra peritonea I ly (i.p.) at 100 and 200 mg/mouse, respectively, q4d x3 starting on Day 2.

Group 4 received paclitaxel i.v., at 24 mg/kg, q4d x3 and rat IgG2a-GSK i.p. at 100 mg/mouse q4d x3 (starting on Day 2).

Groups 5 - 9 received vehicle i.v., q4d x3 and anti-mouse ICOS, clone GSK, i.p., at 1, 5, 20, 100 and 200 mg/mouse, respectively, q4d x3 (starting on Day 2). Groups 10 - 14 received paclitaxel i.v., at 24 mg/kg, q4d x 3 in combination with anti-mouse ICOS, clone GSK, at 1, 5, 20, 100 and 200 mg/mouse, respectively, q4d x 3 (starting on Day 2).

Table 3

Table 3 displays the study design.

Vehicle = 5% ethanol: 5% cremophor in D5W ^a - mg/ kg * - start on Day 2

Results

Rat anti-mouse ICOS surrogate antibody clone 7E.17G9 was assessed in combination with intravenous (IV) administration of Paclitaxel for tumor growth inhibition efficacy in the CT-26 murine syngeneic model. A dose titration of 1, 5, 20, 100, and 200 mg per mouse of mICOS administered intra-peritoneally (IP) was evaluated alongside a fixed dose of Paclitaxel (24 mg/kg) or its vehicle. As a monotherapy, mICOS demonstrated a significant increase in survivability relative to isotype control at 100 mg per mouse. Paclitaxel as a monotherapy demonstrated no ability to significantly increase survivability when compared to control in this model. At 5 mg per mouse, mICOS in combination with Paclitaxel demonstrated a significant increase in survivability when compared to both 5 mg/mouse mICOS and Paclitaxel monotherapies. Both 20 mg/mouse and 100 mg/mouse doses of mICOS with Paclitaxel led to a significant increase in survivability relative to the Paclitaxel monotherapy, but not when compared to respective mICOS 20 mg/mouse and 100 mg/mouse monotherapies (Figure 2).

There were no unexpected mouse deaths in this study. There were instances of ulceration and weight loss observed during this study. However, neither was considered severe enough or met with predetermined criteria in order to warrant removal from study.

This study demonstrates the rat anti-mouse ICOS surrogate antibody clone 7E.17G9 dosed at 5 mg/mouse has statistically significant tumor growth inhibition efficacy in combination with 24 mg/kg Paclitaxel relative to monotherapy treatments of each as measured by the Kaplan-Meier method in the CT-26 model.

Example 3

H2L5 IgG4PE is a humanized IgG4 antibody selected for its potent binding, agonist activity against human ICOS and low/no depleting effects. The unique mechanistic profile of H2L5 IgG4PE offers an opportunity to investigate the antitumor potential of targeting a T cell co-stimulator alone and in combination with standard-of-care (SoC) agents. H2L5 IgG4PE comprises CDR sequences as shown in SEQ ID NOS: 1-6, variable heavy chain and variable light chain sequences as shown in SEQ ID NO:7 and SEQ ID NO: 8, respectively, and heavy chain and light chain sequences as shown in SEQ ID NO:9 and SEQ ID NO:9, respectively.

Described herein is a first-time in human study evaluating the safety, pharmacokinetics (PK), pharmacodynamics (PD), and antitumor activity of H2L5 IgG4PE in selected solid tumours. The study consists of dose escalation and cohort expansion phases; cohort expansion phases are ongoing in several tumor types.

The objectives of the study are as follows:

Primary

• Determine safety, tolerability, and maximum tolerated/administered dose of H2L5 IgG4PE. Secondary

• Determine recommended H2L5 IgG4PE dose(s) for further exploration.

• Evaluate preliminary antitumor activity; characterize PK; evaluate immunogenicity. Exploratory

• Evaluate PD effects.

• Explore associations between antitumor activity, PK and biomarkers in tissue and blood. Methods

The Study is a dose escalation (DE) and ongoing expansion phase study of H2L5 IgG4PE. Modified toxicity probability interval informed DE decisions with ³ 3 patients enrolled per dose level (DL). H2L5 IgG4PE is administered as intravenous infusion every 3 weeks (Q3W); treatment continues up to 2 years or until progression or unacceptable toxicity. Patients must have metastatic or relapsed invasive malignancy, measurable disease, received £ 5 lines of prior therapy in the advanced setting, adequate organ function, and no active autoimmune disease requiring treatment; PK/PD cohorts require pre- treatment and Day 43 on-treatment tumor biopsies. Primary objective is to determine safety, tolerability, and maximum tolerated (MTD) H2L5 IgG4PE dose.

Patients - key inclusion criteria

• Histological or cytological documentation of advanced/metastatic or relapsed/refractory invasive malignancy and is one of the following tumour types: · Bladder/urothelial cancer of the upper and lower urinary tract

• Cervical

• Colorectal (includes appendix) (microsatellite stability (MSS)/ high microsatellite instability (MSI-H))

• Esophageal squamous cell carcinoma · Head and Neck Carcinoma (any histology)

• Melanoma (cutaneous/ocular)

. MPM

• NSCLC (including targeted mutations)

• Prostate · MSI-H/dMMR tumor (Part IB)

• HPV-positive or EBV-positive tumor (Part IB)

• Malignant pleural mesothelioma

• Disease that has progressed after standard therapy for the specific tumor type, or for which standard therapy has proven to be ineffective, is intolerable, or is considered inappropriate, or if no further standard therapy exists

• £5 prior lines of therapy for advanced disease including both standard of care and investigational therapies.

• Measurable disease according to RECIST vl.l guidelines; Eastern Cooperative Oncology Group performance status 0-1; adequate organ function.

• Agree to undergo a pre-treatment and on-treatment biopsy and have disease amenable to biopsy required in PK/PD dose expansion cohorts.

Patients - key exclusion criteria

• Prior anticancer or investigational therapy within 30 days or five half-lives, whichever is shorter. . ³Grade 3 toxicity related to prior immunotherapy and led to treatment discontinuation. . History of invasive malignancy other than disease under study unless disease-free for ³2 years.

• Central nervous system (CNS) metastasis; exceptions include previously treated CNS metastasis that is asymptomatic and has no requirement for steroids at least 14 days prior to first dose of study treatment.

• Active autoimmune disease that required systemic treatment within the last 2 years.

• History of idiopathic pulmonary fibrosis, pneumonitis requiring steroids, interstitial lung disease, or organizing pneumonia.

Study Design

The study design involves

• Accelerated titration design for the first 3 dose levels; 1 patient enrolled at each dose level.

• Modified toxicity probability interval method informed subsequent dose escalation decisions (minimum 3 patients per dose level).

• Starting dose of 0.001 mg/kg: the projected human dose based on the minimally anticipated biologic effect observed in preclinical studies.

The study consists of a dose escalation phase followed by a cohort expansion phase.

This is a FTIH, open-label, multicenter study designed to investigate the safety, tolerability, pharmacology, PK, preliminary clinical activity, and establish a recommended dose of H2L5 IgG4PE for further exploration.

The study consists of a dose escalation (Part 1A) phase followed by a cohort expansion phase (Part IB).

The dose escalation phase evaluates escalating weight-based dose levels of H2L5 IgG4PE administered intravenously once every three weeks (Q3W) to subjects with selected relapsed and/or refractory solid tumors. Based on safety and tolerability, and the PK/pharmacodynamic characteristics of the molecule, recommended monotherapy dose level or dose levels may be further investigated in expansion cohorts.

While expansion cohorts may initiate with H2L5 IgG4PE weight-based dosing, a transition to fixed dosing may be made.

Seamless design is implemented to combine dose escalation with dose expansion, based on toxicity and efficacy (Pan H, Fang X, Liu P, et al. A phase I/II seamless dose escalation/expansion with adaptive randomization scheme (SEARS). Clinical Trials. 2013; 0:1-11). The dose expansion phase may start before the dose escalation phase is completed. All available safety and tolerability data from subjects in dose expansion is incorporated into dose escalation decision making. The basis of the decision to initiate expansion of a dose level/dose will consider following graduation rules:

• Established safety and tolerability;

• Preliminary PK/pharmacodynamic characteristics (i.e., measures of target engagement and functional effects such as receptor occupancy and cytokine release) and/or

• Preliminary antitumor activity.

Once a dose level(s) passes the graduation rules the selected dose(s) may enter into the expansion phase for further investigation; alternate H2L5 IgG4PE schedules may be investigated in the expansion phase. In addition, dose levels under investigation in the ongoing dose escalation phase may incorporate information, such as safety data, from subjects who were accrued to the expansion phase. Randomization and/or futility rules may be incorporated if appropriate in expansion phase to optimize the dose allocation based on evaluations of safety and antitumor activity. The details of randomization schema for expansion cohorts will be documented before the initiation of expansion cohort; details of the futility rules will be documented in the RAP before initiation of interim analyses (Pan H, Fang X, Liu P, et al. A phase I/II seamless dose escalation/expansion with adaptive randomization scheme (SEARS). Clinical Trials. 2013; 0: 1-11).

The study will enroll subjects diagnosed with solid tumor malignancies.

• In the dose escalation phases of the study, and in the PK/pharmacodynamic expansion cohorts the solid tumor types selected for inclusion include bladder/urothelial cancer, cervical cancer, colorectal cancer (CRC), esophageal cancer with squamous cell histology, head and neck (HN) cancer, melanoma, malignant pleural mesothelioma (MPM), non-small-cell lung cancer (NSCLC), and prostate cancer.

• In the cohort expansion phases of the study, several expansion cohorts have been defined by tumor histology or by a specific characteristic such as tumors exhibiting high microsatellite instability (MSI-H), deficiency in DNA mismatch repair (dMMR) processes, or viral -mediated pathology; enrollment in these cohorts is not limited to the tumor types/histologies in the aforementioned list (defined as tumor agnostic).

• Additional expansion cohorts may enroll subjects with a specific tumor type selected from the aforementioned list or from a tumor type/histology not protocol-defined; the basis for the selection will be evidence-based and by an amendment to the protocol to define the cohorts. The overall study size may extend beyond 500 by a protocol amendment if data from expansion phases support extended enrollment or additional combinations are investigated. Assessment of disease status will be performed by the Investigator in accordance with Response Evaluation Criteria In Solid Tumors (RECIST) vl.l and Immune Related (ir) RECIST. A decision to discontinue treatment due to disease progression will be based upon irRECIST; primary efficacy endpoint analysis will use irRECIST. Scans will be collected centrally and stored to allow for the option of central review.

Table 4: Study Treatment

H2L5 IqG4PE Fixed Dose Rationale H2L5 IgG4PE was administered on body weight-based dosing. Fixed doses may be tested in the expansion cohorts and in the safety run-in phase with chemotherapy combinations, assuming a typical median weight of 80 kg.

Therapeutic monoclonal antibodies are often dosed based on body-size due to the concept that this reduces inter-subject variability in drug exposure. However, body-weight dependency of PK parameters does not always explain the observed variability in the exposure of monoclonal antibodies (Zhao X, Suryawanshi, S; Hruska, M. Assessment of nivolumab benefit-risk profile of a 240-mg flat dose relative to a 3 mg/kg dosing regimen in patient with advanced tumors. Annals of Oncology. 2017;28:2002-2008). The advantage of body-weight based versus fixed dosing in this study was evaluated through population PK modelling and simulation efforts. A preliminary population PK model was developed from monotherapy dose escalation (data up to doses of 1 mg/kg; n=19 subjects). Simulations were performed by considering body weight distribution in the simulations based on the observed distribution in the preliminary dataset. At the 5th percentile of body weight (40-47 kg), there was a 70-100% increase in median steady-state AUC(O-); H2L5 IgG4PE exposures higher than these increases have been evaluated in the current Phase 1 study with the 3 mg/kg dose regimen. At the 95th percentile of body weight (107-118 kg), there was a 23-32% decrease in median steady- state AUC(O-) as compared to the median 80 kg exposure providing adequate RO with the minimal lowering of exposure. A similar outcome is expected for steady-state Cmax and trough concentrations between body weight-based and fixed dosing.

Overall, these preliminary population PK simulations indicate that using fixed dosing would result in a similar range of exposures as that of body weight-based dosing. Also, fixed dosing offers the advantage of reduced dosing errors, reduced drug wastage, shorten preparation time, and improve ease of administration. Thus, switching to a fixed dose based on a reference body weight of 80 kg is appropriate.

The fixed dose equivalents of the weight-based H2L5 IgG4PE dose levels using 80 kg weight are presented in Table 5.

Table 5 H2L5 IgG4PE Fixed Dose Calculations

Results

In the DE phase and the PK/PD cohort, 62 patients enrolled: Part 1: 22 in DE and 40 in the PK/PD cohort. 22 patients (35%) had at least one treatment-related adverse event (TR-AE). The most frequent TR-AEs (³3 patients) were fatigue (15%), AST elevations (5%) and diarrhea (3%); AST elevations were the most frequent Grade 3/4 TR-AE (N=2 [3%]). Approximate dose proportional increases in systemic H2L5 IgG4PE concentrations over 0.01 - 3mg/kg DLs were observed. At DLs ³ 0.3 mg/kg, ICOS receptor occupancy was ³75% across the dosing interval. On-target PD effects in tumor infiltrating lymphocytes and clinical activity were observed in Part 1; including in anti-PD-l/Ll experienced patients. Table 6 shows patient disposition by cohort and dose.

Table 6

Table 7 shows patient and disease characteristics.

Table 7

‘MONOTHERAPY DOSE ESCALATION & PK /PD; 10» MSS; COMBINATION DOSE ESCALATION; 84% MSS, 36% MSI-H iSS, MICROSATELLITE STABILITY; MSI-H, HIGH MICROSATELLITE INSTABILITY; NSCLC, NON-SMALL-CELL LUNG CARCINOMA; PD-11, PROGRAMMED DEATH-LIGAND 1; SCC, SQUAMOUS CELL CARCINOMA Table 8 shows treatment-related AEs (in ³3 patients).

Table 8

AE - Adverse Event; AST - Aspartate Aminotransferase; TR-AE - Treatment-related AE.

Treatment-related safety

• 1 patient in monotherapy dose escalation cohort experienced Grade 3/4 elevations in alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, bilirubin, gamma-glutamyl transpeptidase, impaired liver function (serious) and Grade 1 amylase and G3 lipase.

• Serious adverse events (SAEs) in monotherapy group: 1 patient (3 mg/kg) had impaired liver function (Grade 3).

• Confounded by progression in liver metastases and biliary tract obstruction that required stenting.

Patient 1: H2L5 IaG4PE monotherapy fFIG. 61 History:

• 53Y Male; Stage IIIc nodular melanoma [BRAF/cKIT mutation negative].

• Prior regimens: ipilimumab/nivolumab ~ 2 months; nivolumab ~ 1 year, SD best response.

• Disease Burden: 5 target lesions (LN, Lung, SubQ): SoD=225 mm. Multiple non target lesions.

Study treatment:

• H2L5 IgG4PE monotherapy to Week 48; 0.1 mg/kg x 3 doses, then 1 mg/kg. Tumor biopsies collected after 43 days on-treatment showed greater number of T cells, granzyme-B expressing CD8 Tc cells, PD1 expressing T cells and proliferating T cells while fewer proliferating tumor cells (data not shown).

Patient 2: H2L5 IaG4PE monotherapy History:

• BRAF negative, N/KRAS mutation positive; Stage lb superficial spreading melanoma.

• Prior regimens: nivolumab (advanced/metatstatic) ~ 10 months; Electrochemotherapy.

Study treatment:

• H2L5 IgG4PE monotherapy at 1 mg/kg.

The post treatment sample (data not shown) showed:

• Higher TIL including cytotoxic, helper T cells and NK cells

• More Granzyme B+ T cells and less proliferating tumour cells

• Increase in activated T cells as observed with greater 0X40 and HLADR expression

• Upregulation of PD1 and PD-L1 upon H2L5 IgG4PE treatment

Conclusions

• H2L5 IgG4PE was well tolerated in patients with advanced solid tumours at the 0.001-3 mg/kg dose range.

• Maximum tolerated dose was not reached; maximum administered dose was 3 mg/kg H2L5 IgG4PE.

• Majority of AEs were Grade 1/2 and not attributed to study treatment.

• AEs leading to discontinuation occurred in 1 patient (n=62 patients) at the highest dose level.

• Dose proportional increases in H2L5 IgG4PE concentrations.

• PK/PD analysis showed ³75% total ICOS receptor saturation across dosing interval at H2L5 IgG4PE dose levels ³0.3 mg/kg.

• A range of doses (³0.1-1 mg/kg) have shown biological and clinical activity (including in patients with prior anti-PD-l/Ll exposure). These doses are being investigated further in expansion cohorts to establish the recommended H2L5 IgG4PE dose.

• Preliminary biological and clinical data support the mechanism of action of a non-depleting ICOS agonist as a clinical target. • Doses over 0.1 mg/kg are being investigated further in expansion cohorts to establish the recommended H2L5 IgG4PE dose.

Example 4

Example 4 describes pharmacokinetics/pharmacodynamics (PK/PD) exposure-response characterization of H2L5 IgG4PE from the study described in Example 3. H2L5 IgG4PE is an agonist IgG4PE antibody against inducible co-stimulatory receptor (ICOS) with immune stimulating and antineoplastic activity. The study described Example 4 is the first in human study investigating H2L5 IgG4PE.

Methods

Safety, PK, PD, and preliminary antitumor activity of H2L5 IgG4PE were evaluated at doses from 0.001 to 3 mg/kg every 3 weeks (Q3W). Blood samples collected prior to dosing and select time points on-study were evaluated for PK and PD effects on lymphocytes and ICOS receptor occupancy (RO). Tumor tissue at Screening and Week 6 were evaluated for changes in tumor immune infiltrates (TIL) by a multiplexed immuno-fluorescence platform.

PK Analysis

• A preliminary population PK data set was constructed with all pooled concentration-time data

• Serial plasma samples were collected throughout; PK samples were assayed by validated ELISA assay and concentration-time data was modelled using nonlinear mixed effects, as implemented in NONMEM.

Pharmacodynamic (PD) Analysis

• Flow cytometry was performed instream throughout the study to evaluate ICOS receptor occupancy (RO) with H2L5 IgG4PE.

• For PK/PD and expansion cohorts, tumor tissue was collected at pre-dose and at Week 6 for evaluation of overall TIL, changes in activation, proliferation and gene expression changes.

• Exposure measure for PK/PD analyses defined as Week 6 pre-dose trough concentration derived from population PK model.

• Evaluation of gene expression changes in the tumour micro environment (TME) were performed using the Nanostring nCounter™ platform.

• Multiomyx™ multiplexed immunofluorescence was used to characterize the immune phenotype of the TIL. Results

The PK disposition of H2L5 IgG4PE is consistent with that of other humanized mAbs, with low clearance and limited central volume of distribution. Dose and concentration-receptor occupancy (RO) analyses suggest >~0.1 mg/kg H2L5 IgG4PE maintains high RO on CD4+ and CD8+ T cells. Quantitative TIL evaluation of paired tumor biopsies demonstrates potentially favorable immune microenvironment in the tumor at exposures observed in subjects treated with 0.3mg/kg dose. TIL and gene expression data from tumor RNA demonstrate non-linear, dose-dependent changes in select markers of immune activation. Clinical exposure-response assessments reveal no difference in baseline-to-Week 9 target lesion change across exposures in the 1L R/M HNSCC expansion cohort. Likewise, cross-cohort pooled exposure-response analysis of AEs of ³Grade 2 severity demonstrates similar safety outcomes across the exposures/doses. Population PK modeling suggests fixed doses maintain exposures within established safety bounds.

Pharmacokinetics and Target Engagement

• PK and target engagement characteristics of H2L5 IgG4PE are similar to prior reports, with a population clearance estimate of ~0.27 L/day and central volume estimate of ~3.6 L, and limited impact of bodyweight on systemic exposure.

• Plasma concentrations of H2L5 IgG4PE increase in a dose-proportional manner (Figure 4A), while ICOS RO was maintained above ~70% with H2L5 IgG4PE doses of 0.1 mg/kg and higher (Figure 4B).

• Minimal differences in RO are observed for CD4⁺ with H2L5 IgG4PE doses of 0.3 mg/kg and 1.0 mg/kg (Figure 4C), with similar results for CD8⁺ (data not shown). However, there was large variability in RO for doses <1.0 mg/kg (Figure 4D)

MultiOmyx and Gene Expression Data

• Quantitative evaluation of TILs in paired tumor biopsies demonstrates on-study changes in TILs follow a non-linear, exposure/dose-dependent pattern.

• Changes in select immune activation markers favors a greater cytotoxic T cell to regulatory T cell ratio with H2L5 IgG4PE exposures of 1000-10000 ng/ml at Ctrough which corresponds to doses between ~0.3 mg/kg to 1 mg/kg (Figure 5A).

• Non-monotone dose-dependent changes in total TIL as well as other activation and proliferating T cell phenotypes were detected in on-treatment biopsies when compared to baseline in MultiOmyx™ immunofluorescence data with H2L5 IgG4PE 0.3 mg/kg and higher doses (Figure 5B).

• Gene expression changes in the tumor show a non-linear dose response trend with the highest increases at ³0.1mg/kg and greatest reductions at <lmg/kg (data not shown). • The ratio of cytotoxic T cell proliferation (CD3+CD8+Ki67+) over regulatory T cells proliferation (CD3+CD4+FOXP3+Ki67+) was higher in Week 6 on-treatment biopsies when compared to pre-treatment tumor samples for subjects at 0.3-1 mg/kg doses of H2L5 IgG4PE who experienced disease control (DC) benefit when compared to subjects that did not experience disease control (Figure 5C).

Conclusion

The current data provide preliminary evidence of H2L5 IgG4PE target engagement and biological activity at clinically tolerable doses and support further exploration of a 24 mg Q3W fixed dosage in R/M HNSCC.

Example 5

The first time in human studies described in Examples 3 and 4, was also expanded to evaluate the safety, tolerability and anti-tumour effects of H2L5 IgG4PE in combination with chemotherapy.

The dose of H2L5 IgG4PE in each chemotherapy combination is 24 mg based upon evidence of activity in other cohorts at 0.3 mg/kg and comparability of exposure at 24 mg. The passage below describes the initial dose escalation design:

H2L5 IgG4PE in combination with docetaxel or platinum-based (cisplatin or carboplatin) chemotherapy doublets consist of safety run-in cohorts only. The mTPI (modified Toxicity Probability Interval) (Ji 2010) design will guide dose escalation/de-escalation decisions to support determining the H2L5 IgG4PE recommended phase 2 (RP2) dose in combination with standard doses of chemotherapy. An initial three subjects will receive 80 mg of H2L5 IgG4PE in combination with chemotherapy; if no dose limiting toxicities (DLTs) are observed, an additional 6 to 9 subjects may receive this dose or de- escalate/escalate to a lower/higher dose in combination with chemotherapy.

A minimum of nine subjects will receive H2L5 IgG4PE at the dose recommended in combination with each chemotherapy regimen. Safety, tolerability, PK, pharmacodynamic measures, and antitumor activity will be considered in determining RP2D (recommended phase 2 dose) of H2L5 IgG4PE for each chemotherapy combination.

Study Population

The study population in H2L5 IgG4PE plus chemotherapy safety cohorts will be adults with advanced/recurrent solid tumor malignancies, who either:

• have disease that has progressed on prior systemic therapy

• are not candidates for standard of care therapy, or

• have disease for which no further standard of care therapy exists. The expansion cohort populations may be comprised of subjects with advanced/recurrent solid tumors who have not received treatment for advanced disease.

Study Treatment and Duration

The study is composed of two phases, dose escalation/safety run in; and dose expansion. Each phase of the study includes a screening period, a treatment period, and a follow-up period. For subjects who meet all eligibility criteria and register into the study, the maximum duration of treatment with H2L5 IgG4PE is expected to be two years, up to 35 cycles; in those subjects who receive combination therapy, the maximum duration of treatment with H2L5 IgG4PE in combination with chemotherapy is expected to be two years, up to 35 cycles. The duration of chemotherapy treatment will be according to institutional practice but should be a minimum of 4 cycles (cycle=21 days). The maximum follow-up period for safety assessments will be 90 days from the date of the last dose of study treatment. The expected maximum follow-up period for survival and subsequent anti-cancer therapy will be two years from the date of the last dose of study treatment. Subjects who discontinue study treatment due to achieving confirmed complete response (CR) will be followed for progression.

Subjects participating in chemotherapy combination cohorts will receive H2L5 IgG4PE 24 mg or 80 mg dose (refer to Table 5 for fixed doses) in combination with chemotherapy at doses and schedules based on standard of care practice (Table 10).

Chemotherapy dose rationale

Docetaxel is a semisynthetic taxane approved in different tumor indications. The dosage of docetaxel as a single-agent and in combination for several tumor indications, including NSCLC and HNSCC, is 75mg/m2, every three weeks; thus, this dose and schedule was selected in combination with H2L5 IgG4PE.

The carboplatin chemotherapy doublets will calculate the dose of carboplatin according to Calvert formula using the target AUC of 4-6 mg/ml per min as per local standard of care in combination with:

1. Pemetrexed at 500 mg/m2

2. Gemcitabine at 1250 mg/m2

3. Paclitaxel at 200 mg/m2

The doses selected in combination are according to Category 1 recommendations in National Comprehensive Cancer Network treatment guidelines.

Carboplatin (AUC 5) or cisplatin at 100 mg/m2 (per GSK assignment) will be combined with 5-FU at 1000 mg/m2 (see Table 10). The doses selected were in accordance with standard doses given for first-line treatment of recurrent or metastatic HNSCC [Vermorken, 2008]. Study Treatment

The study treatment is outlined in Table 9

Table 9

Chemotherapy regimens will be administered to subjects starting at least 30 minutes and no more than one hour following the end of the H2L5 IgG4PE infusion. The sequence in which chemotherapy doublets are administered is per standard practice. The date and time of administration will be documented in the source documents and reported in the eCRF. Subjects should receive the indicated premedication regimens and supplementation according to the approved product label or standard practice (i.e., corticosteroids, folic acid, vitamin B12, and diphenhydramine). Chemotherapy premedication indicated on the day of dosing should be administered after H2L5 IgG4PE EOI (end of infusion).

Chemotherapies will be administered for a minimum of 4 and maximum of 6 cycles according to standard practice; treatment with docetaxel and pemetrexed may continue beyond 6 cycles according to standard practice.

Gemcitabine will be administered on Day 1 and Day 8 of every 3-week/21-day cycle.

Fluorouracil (5-FU) will be administered continuous on Day 1 through Day 4 of every 3- week/21-day cycle. Table 10

Abbreviations: AUC=area under the curve; FU= Fluorouracil; IV=intravenous; Q3W= every three weeks H2L5 IgG4PE and 5-FU/platinum combination treatment

Part 2A includes safety run-in cohorts evaluating H2L5 IgG4PE combinations with chemotherapy. Preliminary results of combination including 5-FU/platinum chemotherapy are reported below.

Patients eligible for H2L5 IgG4PE + 5-FU/platinum (cisplatin or carboplatin) had a diagnosis of advanced selected solid tumors and £5 prior lines of systemic therapy.

5-FU (1000 mg/m²)/platinum (cisplatin 100 mg/m² or carboplatin are under the curve (AUC) 4-6 mg/ml/min) were administered every 3 weeks for 4-6 cycles (Burtness, 2019) and H2L5 IgG4PE was administered at 24 or 80 mg every 3 weeks for up to 2 years/35 cycles or until disease progression or unacceptable toxicity.

Tumour Response to H2L5 IgG4PE and 5-FU/platinum combination treatment The tumour response of the subjects treated with H2L5 IgG4PE+Fluorouracil+Cisplatin ( i.e . H2L5 IgG4PE and 5-FU/cisplatin) and IgG4PE+Fluorouracil+Carboplatin {i.e. H2L5 IgG4PE and 5- FU/carboplatin) at 24 mg or 80 mg is shown in Figure 7. A corresponding plot showing percentage change from baseline in tumour measurement (irRECIST) is shown in Figure 8. Figure 9 shows a plot of most common treatment related adverse events (AE) observed for combination treatment of H2L5 IgG4PE and 5-FU/cisplatin or H2L5 IgG4PE and 5-FU/carboplatin.

Figure 8 represents the best change from baseline of target lesion sum of diameters in patients who had at least 1 on-treatment disease assessment, defined as the evaluable population for this analysis. In the H2L5 IgG4PE combination with fluorouracil platinum-based chemotherapy (cisplatin or carboplatin) cohort, 8 patients were evaluable. One patient who received H2L5 IgG4PE 24 mg in combination with standard doses of fluorouracil platinum-based chemotherapy attained a partial response. In the 2 patients who attained a stable disease, 1 patient received H2L5 IgG4PE 24 mg in combination with standard doses of fluorouracil platinum-based chemotherapy and the other patient received H2L5 IgG4PE 80 mg in combination with standard doses of fluorouracil platinum-based chemotherapy. While 3 patients were not evaluable for an overall immune-related RECIST defined response assessment, the best change from baseline in target lesions supports a target lesion response of stable disease; all 3 of these patients received H2L5 IgG4PE 80 mg in combination with fluorouracil platinum-based chemotherapy. In the 2 patients with a best overall of progressive disease, 1 patient who received H2L5 IgG4PE 80 mg in combination with fluorouracil platinum-based chemotherapy had a target lesion response of stable disease while the other patient who received H2L5 IgG4PE 24 mg in combination with fluorouracil platinum-based chemotherapy had a target lesion response of progressive disease.

Example 6

Described herein is a randomized, Phase II open-label platform trial utilizing a master protocol designed to investigate the clinical activity of novel regimens consisting of immuno-oncology agents compared with standard of care (SoC) regimens in participants with relapsed/refractory advanced non-small cell lung cancer (NSCLC) who have failed prior platinum-containing chemotherapy regimen and an immuno-oncology agent, such as anti-programmed cell death protein 1 (PD1) / PD-Ligand 1 (PD-L1) - either in combination or as separate lines.

NSCLC is considered intrinsically resistant to immuno-oncology agents owing in part to its broad immune escape and suppressive features that include low antigenicity, despite having one of the highest frequencies of somatic mutations, and a high presence of regulatory T cells (Tregs). However, as shown by the single-agent response rates of anti-PD-1 inhibitors in NSCLC, a subset of tumors are susceptible to T cell-mediated antitumor effects, suggesting those tumors have some degree of prior T-cell immunity. Since effective anticancer immune response involves stepwise multistep processes, lung cancers may possess or acquire features that enable them to evade immune surveillance, suppress immune reactivity, proliferate, and survive within an inflammatory microenvironment, thereby rendering an immune response ineffectual. Therefore, treatment modalities that incorporate combinations with agents targeting different processes within the immune cascade have the potential to reinstate immunosurveillance; these may include regimens containing chemotherapy that possess advantageous immunological effects to improve clinical efficacy.

The objectives of the study are as follows:

Primary

• Determine whether H2L5 IgG4PE in combination with docetaxel (experimental regimen) provide evidence for improved survival over SoC therapy.

Secondary

• Evaluate milestone survival in participants treated experimental regimen versus SoC therapy for NSCLC.

• Evaluate other measures of antitumor activity of the experimental regimens compared with SoC therapy for NSCLC (RECIST (Response Evaluation Criteria In Solid Tumors) 1.1 and iRECIST).

• Evaluate the safety and tolerability of the experimental regimens compared with SoC therapy for NSCLC

• Characterize the pharmacokinetic properties of H2L5 IgG4PE when given in combination with chemotherapy and/or other immunotherapies

Overall Design

This is a randomized Phase II, open-label, platform trial utilizing a master protocol to study novel immunotherapy drug combinations compared with the current SoC in the treatment of patients with advanced NSCLC who have progressed on prior anti-PD(L)l and platinum-based combination chemotherapies. As shown in Figure 10, the study will initially evaluate 2 treatment regimens and additional regimens (Arm 3, Arm 4, etc.) will be added based on emerging nonclinical and clinical data, via future protocol amendment(s). No treatment crossover is allowed in this study.

Each additional treatment arm/regimen will be analyzed relative to the SoC treatment. The data generated from each experimental regimen and associated control arm data is considered a substudy within the overall platform study, as shown in Figure 10.

The study will initially evaluate the efficacy of H2L5 IgG4PE in combination with SoC (docetaxel) compared with SoC alone as the standard subsequent-line chemotherapy (substudy 1) in NSCLC. Within each substudy, patients will be randomized to receive either SoC or the experimental treatment. Additional treatment arms will be added as substudies (e.g., substudy 2, substudy 3, etc.) via future protocol amendments, and ongoing treatment arms may be dropped for futility, based on prespecified futility rules at interim analyses. Participants will be randomized 1:2 to Arm 1 (SoC) and Arm 2, i.e., 33% and 67%, respectively.

Patients key inclusion criteria

• Histologically or cytologically confirmed diagnosis of NSCLC with known histology (squamous or non-squamous) and: a. Documented disease progression (for example, based on radiographic imaging) during or after a maximum of 2 lines of systemic treatment for locally/regionally advanced recurrent, Stage Illb/Stage IV or metastatic disease: i. A maximum of 1 line of platinum-containing chemotherapy regimen in the metastatic setting, and ii. A maximum of 1 line of PD(L)1 mAb containing regimen. b. Participants with known BRAF molecular alterations must have had disease progression after receiving the locally available SoC treatment for the molecular alteration. Participants with this alteration could have received up to 3 lines of systemic anticancer therapy.

• Measurable disease, presenting with at least 1 measurable lesion per RECIST 1.1 guidelines; Eastern Cooperative Oncology Group performance status score of 0-1; adequate organ function.

Patients key exclusion criteria

• Received prior treatment with the following therapies (calculation is based on date of last therapy to date of first dose of study treatment): a. Docetaxel at any time b. Any of the investigational agents being tested in the current study, including experimental ICOS agonist

• Received ³4 prior lines of therapy for NSCLC, including participants with BRAF molecular alternations. Patients with known EGFR/ALK/ROS1 molecular alterations are excluded from participation in this study.

Study Treatments Standard of Care Arm: Docetaxel Alone

Table 11 Description and Administration of Docetaxel

a - TAXOTERE PI, 2015; TAXOTERE SmPC 2015. b - Docetaxel will be sourced locally from commercial stock, except in countries where regulatory authorities mandate that the Sponsor supply all study treatment(s) required for the conduct of a clinical trial.

All participants randomly assigned to docetaxel-containing arms are premedicated with oral corticosteroids such as dexamethasone 16 mg per day or its equivalent per local standards (e.g. 8 mg twice daily) for 3 days starting 1 day prior to docetaxel administration to reduce the incidence and severity of fluid retention as well as the severity of hypersensitivity reactions [TAXOTERE PI, 2015; TAXOTERE SmPC 2015.]. Intravenous corticosteroid premedication may also be utilized per local standard and at the discretion of the investigator.

Docetaxel Dose Justification

The dosage of docetaxel for this study, as a single agent and in combination, will be 75mg/m² Q3W, as described in the labels [TAXOTERE PI, 2015; TAXOTERE SmPC 2015.] which is approved (a) as a single-agent for patients with locally advanced or metastatic NSCLC after platinum-based chemotherapy, and (b) in combination with cisplatin for unresectable, locally advanced or metastatic NSCLC for patients who have not received prior chemotherapy.

Substudv 1 (Experimental Arm 21: H2L5 IgG4PE and Docetaxel Combination

Table 12 Description and Administration of Arm 2 Study Treatments

a - TAXOTERE PI, 2015; TAXOTERE SimPC 2015.

In H2L5 IgG4PE-containing arms, H2L5 IgG4PE is administered first as a 30-minute IV infusion (infusion time may be adjusted based on infusion related reactions). The administration of the second agent in these arms must be started 1 hour and no more than 2 hours after the end of H2L5 IgG4PE infusion.

Rationale for ICOS Agonist/ Docetaxel Combination

Cancer immunity is described as a multistep process that elicits an effective antitumor response [Chen DS, Mellman I. Oncology meets immunology: the cancer-immunity cycle. Immunity. 2013; 39:1-10., 2013]. Each step can be negatively regulated, thus providing the tumor with redundant mechanisms by which to block an antitumor immune response. In some cases, tumors are highly dependent on a single mechanism, and in these cases, there is the potential to achieve significant clinical activity with a single agent immunomodulatory therapy. Robust antitumor responses including complete cure have been achieved in some cancers by modulating the patient's immune system. Antibodies targeting the checkpoint receptors or their cognate ligands engaged in negative regulation of T cell responses, such as CTLA-4 and PD-1/PD-L1, have demonstrated efficacy as anticancer immunotherapies in a broad range of tumors including some solid tumors otherwise considered poorly immunogenic.

However, a majority of tumors are non-responsive to this class of agents. One reason for the lack of response could be the existence of multiple mechanisms of immune suppression in the tumor microenvironment which prohibits effective antitumor immune responses. In these instances, combination therapies will likely be required. The clinical data generated by the combination of ipilimumab (anti-CTLA-4) and nivolumab (anti-PD-1) in patients with metastatic melanoma is an example of the practice changing clinical benefit of such combinations [Wolchok JD, Kluger H, Callahan MK, et al. Nivolumab plus ipilimumab in advanced melanoma. N Engl J Med. 2013; 369:122-133., 2013]. In some patients, inhibition of negative immune checkpoint pathways alone may not elicit an effective antitumor response, and additional co-stimulatory signals may be necessary to mount an effective response. Immunomodulatory agents that target other components of the cancer immunity cycle are needed to expand the population of patients and range of tumor types that may respond to immunotherapy as well as enhance the magnitude and duration of antitumor responses in patients whose tumors are already sensitive to current immunotherapy approaches. The ultimate aim is to improve the survival outcome in all disease settings including the advanced setting.

H2L5 IgG4PE is a humanized IgG4 anti-ICOS monoclonal antibody selected for its nanomolar (nM) binding to and agonist activity in ICOS-expressing CD4+ and CD8+ effector T cells. H2L5 IgG4PE is specifically engineered as an Immunoglobulin (Ig)G4 hinge-stabilized isotype, IgG4PE, to markedly decrease binding affinity of the Fc (Fragment crystallizable) region of the mAb to activating Fey receptors and Clq, and thereby diminish the cytotoxic potential of H2L5 IgG4PE that would result in depletion of ICOS-positive T cells through antibody-dependent or complement-dependent cell mediated mechanisms, respectively. Moreover, the IgG4PE isotype retains functional binding to the Fey inhibitor receptor, FcyRIIb, a feature described as critical for modulating antibody agonist activity [Li, 2011], which also may be essential for optimal ICOS agonist activity and its associated antitumor effects in humans.

ICOS is a co-stimulatory receptor of the CD28/CTLA immunoglobulin super family with expression restricted to T cells. ICOS is weakly expressed on resting TH17, follicular helper T and regulatory T (Treg) cells and yet is highly induced on CD4+ and CD8+ T cells upon T cell receptor (TCR) engagement and activation [Paulos CM, Carpenito C, Plesa G, et al. The inducible costimulator (ICOS) is critical for the development of human Thl7 cells. Sci Transl Med. 2010; 2:55ra78., 2010; Wakamatsu El, Mathis D, Benoist C. Convergent and divergent effects of costimulatory molecules in conventional and regulatory CD4+ T cells. PNAS. 2013; 110: 1023-1028., 2013]. Upregulation of ICOS leads to both Thl and Th2 cytokine secretion and sustained effector T cell proliferation and function [Sharpe AH, Freeman GJ. The B7-CD28 Superfamily. Nat Rev Immunol. 2002; 2:116-126., 2002]. A growing body of evidence supports the concept that activating ICOS on CD4+ and CD8+ effector T cells has antitumor potential.

The rationale for targeting ICOS in cancer has been established by multiple lines of nonclinical and clinical evidence. Engagement of the ICOS pathway with an ICOS-L-Fc fusion protein is shown to have potent antitumor activity in multiple syngeneic mouse tumor models [Ara G, Baher A, Storm N, et al. Potent activity of soluble B7RP-1-Fc in therapy of murine tumors in syngeneic hosts. Int J Cancer. 2003; 103:501-507., 2003]. Emerging data from patients treated with anti-CTLA-4 antibodies suggest a positive role of ICOS+ effector T cells in mediating an antitumor immune response. Patients with metastatic melanoma, urothelial [Carthon BC, Wolchok JD, Yuan J, et al. Preoperative CTLA-4 blockade: Tolerability and immune monitoring in the setting of a presurgical clinical trial. Clin Cancer Res. 2010; 16:2861-2871., 2010], breast [Vonderheide RH, LoRusso PM, Khalil M, et al. Tremelimumab in combination with exemestane in patients with advanced breast cancer and treatment-associated modulation of inducible costimulator expression on patient T cells. Clin Cancer Res. 2010; 16:3485-3494., 2010] and prostate cancer [Chen H, Liakou Cl, Kamat A, et al. Anti-CTLA- 4 therapy results in higher CD4+ICOShi T cell frequency and IFN-gamma levels in both nonmalignant and malignant prostate tissues. Proc Natl Sci USA. 2009;106:2729-2734., 2009] who have increased absolute counts of circulating and tumor infiltrating CD4+ICOS+ and CD8+ICOS+ T cells after ipilimumab treatment have significantly better treatment related outcomes than patients where little or no increases are observed. Importantly, it was shown that ipilimumab changes the ICOS+ T effector to Treg ratio, reversing an abundance of Tregs pre-treatment to a significant abundance of T effectors vs. Tregs following treatment [Liakou Cl, Kamat A, Tang D, et al. CTLA-4 blockade increases IFN-gamma producing CD4+ICOShi cells to shift the ratio of effector to regulatory T cells in cancer patients. Proc Natl Acad Sci USA. 2008; 105:14987-14992., 2008; Vonderheide RH, LoRusso PM, Khalil M, et al. Tremelimumab in combination with exemestane in patients with advanced breast cancer and treatment-associated modulation of inducible costimulator expression on patient T cells. Clin Cancer Res. 2010; 16:3485-3494., 2010]. As evidenced by the clinical data, ICOS+ T effector cells may be a positive predictive biomarker of ipilimumab response, and activation of this population of cells with an ICOS agonist antibody may confer an advantage by mounting a more robust immune antitumor response.

Similar to the combination of platinum-containing chemotherapy with immuno-oncology (IO) agents (anti-PD-1) in metastatic non-squamous disease, i.e., the incorporation of the anti-PD-1 inhibitor, pembrolizumab, to the pemetrexed/carboplatin backbone in the first-line metastatic non- squamous disease is an example for the IO agents to provide a higher degree of benefit including a prolonged benefit combined with the immediate cytotoxic effects of the chemotherapy. The ICOS agonist/docetaxel combination has the potential to deliver a similar promise to later line NSCLC participants building on the existing docetaxel standard of care.

Chemotherapy can promote tumor immunity by inducing immunogenic cell death as part of its intended therapeutic effect, as well as modulating distinct features of tumor immunobiology [Emens LA, Middleton G. The interplay of immunotherapy and chemotherapy: harnessing potential synergies. Cancer Immunology Research. 2015 May l;3(5):436-43., 2015]. In preclinical models, combinations with various chemotherapy agents including docetaxel and platinum based treatments with anti-PD- L1 treatment showed increased efficacy associated with increased frequency of intratumoral subsets without antagonizing functional changes mediated by anti-PD-Ll [Cubas R, Moskalenko M, Cheung J, et al. Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; September 25-28, 2016; New York, NY DOI: 10.1158/2326-6066.IMM2016- A114 Published November 2016., 2016]. Combination of anti-ICOS surrogate antibody with carboplatin and paclitaxel showed increase efficacy in the CT26 tumor model (see Examples 1 and 2 above) Dose Justification

H2L5 IgG4PE Dose Rationale

H2L5 IgG4PE is being currently investigated in a Phase 1 study with doses ranging from 0.001 mg/kg to 3 mg/kg as monotherapy and from 0.01 mg/kg to 3 mg/kg in combination with a PD-1 inhibitor (Example 4). Preliminary PK data from the study was utilized to develop a population PK model and estimate median steady-state peak and trough exposures at different fixed doses as listed in Table 13. The 80-mg dose corresponds to an approximate 1 mg/kg dose assuming median body weight of 80 kg as described.

The functional effect of H2L5 IgG4PE has been characterized in several in vitro experiments yielding different activity coefficients depending on cell type, co-stimulation status, and cytokines analyzed. The ICOS receptor occupancy (RO) based on CD4+ or CD8+ T-cells at any given systemic exposure of H2L5 IgG4PE can be predicted by employing the in vitro potency values generated from different binding/activation assays in the range of 0.09 to 4.14 mg/mL as listed in Table 13.

Sufficiently high CD4+ RO is expected at peak exposures (89% to >99% RO) as well as at trough exposures (69% to >99% RO) at steady-state with the proposed 80 mg dose.

Collectively, based on the safety and exposure data from the Phase 1 study and the predicted target engagement, the 80-mg dose is proposed to be evaluated in combination with docetaxel in this study. No drug-drug interaction related changes are expected in H2L5 IgG4PE PK with docetaxel coadministration. The currently planned 80 mg H2L5 IgG4PE dose may be adjusted lower to 24 mg or increased to 240 mg based on any emerging safety, exposure and/or pharmacodynamic data.

Table 13 Projected CD4+ Receptor Occupancy from Population PK Predicted Median Steady-State Peak and Trough Exposures of H2L5 IgG4PE Based on In Vitro Potency Estimates.

If H2L5 IgG4PE is included in any other arm/regimen of the study, the 80-mg dose may be adjusted to a lower 24 mg dose corresponding to an approximate 0.3 mg/kg dose or to a higher 240 mg dose corresponding to an approximate 3 mg/kg dose based on any emerging data or depending on the other drug included in the combination in that study arm.

H2L5 IgG4PE Dosing Frequency

The systemic half-life of H2L5 IgG4PE is approximately 25 days based on the preliminary population PK analysis of exposure data from ongoing study. The existing H2L5 IgG4PE Q3W regimen in the ongoing clinical study is also consistent with the Q3W dosing regimen typical with IgG4 based monoclonal antibody therapies. The docetaxel label prescribes a Q3W regimen. Thus, H2L5 IgG4PE will be dosed Q3W in combination with docetaxel. Combination of H2L5 IgG4PE with any other treatment in other arms of this study may have a different dosing regimen as deemed appropriate.

Rationale for Fixed Dose

Therapeutic monoclonal antibodies are often dosed based on body-size due to the concept that this reduces inter-participant variability in drug exposure. However, body-weight dependency of PK parameters does not always explain the observed variability in the exposure of monoclonal antibodies. The advantage of body-weight based versus fixed dosing in this study was evaluated through population PK modelling and simulation efforts. A preliminary population PK model was developed from monotherapy dose escalation (data up to doses of 1 mg/kg; n=19 participants).

Simulations were performed by considering body weight distribution similar to that observed in the preliminary dataset. At the 5th percentile of body weight (40-47 kg), there was a 70-100% increase in median steady-state AUC(O-x); H2L5 IgG4PE exposures higher than these increases have been evaluated in the Phase 1 study 204691 with the 3 mg/kg dose regimen. At the 95th percentile of body weight (107-118 kg), there was a 23-32% decrease in median steady-state AUC(O-T) as compared to the median 80 kg exposure providing adequate RO with the minimal lowering of exposure. A similar outcome is expected for steady-state Cmax and trough concentrations between body weight-based and fixed dosing.

Overall, these preliminary population PK simulations indicate that using fixed dosing would result in a similar range of exposures as that of body weight-based dosing. Also, fixed dosing offers the advantage of reduced dosing errors, reduced drug wastage, shorten preparation time, and improve ease of administration. Thus, switching to a fixed dose based on a reference body weight of 80 kg is reasonable and appropriate.

Docetaxel Dose Rationale Docetaxel is a semisynthetic taxane approved in different tumor indications. The dosage of docetaxel as a single agent and in combination for several tumor indications, including NSCLC and HNSCC, is 75mg/m², every three weeks; thus, this dose and schedule was selected in combination with H2L5 IgG4PE. There are no drug-drug interaction related changes expected in docetaxel PK on co-administration with H2L5 IgG4PE.

Treatment Groups and Duration

After a screening period of up to 28 days, eligible participants for each substudy will be randomly assigned to a treatment arm (SoC or experimental) and receive study treatment (Day 1).

Unless otherwise specified, investigational combination study treatment will continue at the indicated schedule for a maximum duration of 2 years or 35 treatment visits, whichever comes first, or until disease progression, death, unacceptable toxicity, or withdrawal of consent. Single agent SoC treatment (i.e., docetaxel) may continue until disease progression, death, unacceptable toxicity, withdrawal of consent, or per institutional standard for docetaxel. After the study treatment is permanently discontinued, participants will be followed, via telephone contact, for survival and subsequent anticancer therapy every 12 weeks until death or the participant's withdrawal from further contact. Adverse events will be collected for 90 days after the last dose of study treatment or until the start of new anticancer therapy, whichever comes first. Participants permanently discontinuing study treatment prior to documented disease progression by iRECIST will also be followed every 12 weeks for disease progression or participant's withdrawal from further contact.

The study will be considered 'finished' once the last participant from all open treatment arms has completed their last survival follow-up contact.

Number of Participants

As the study uses a master protocol design, the sample size is not fixed. The initial number of participants is estimated to be at least 105 in substudy 1 (SoC arm: 35; experimental arm: 70). Additional experimental regimens may be added via protocol amendments and will be considered as another substudy. Each additional experimental arm will enroll a maximum of 70 participants. The minimum sample size for the SoC arm is 35 with additional subjects randomized to SoC concurrently with additional experimental arms/substudies. Further randomization to SoC will be minimized once 35 patients are enrolled in the first substudy.

Guidelines for Evaluation of Disease Evaluation of Anti-Cancer Activity

• RECIST version 1.1 guidelines will be used to determine the overall tumor burden at Screening, select target and non-target lesions, and in the disease assessments through the duration of the study [Eisenhauer EA, Therasse P, Bogaerts J, et al. New response evaluation criteria in solid tumours: revised RECIST guidelines (version 1.1). Eur J Cancer. 2009; 45:228-47. , 2009].

• As indicated in RECIST version 1.1 guidelines: o Lymph nodes that have a short axis of <10 mm are considered non-pathological and must not be recorded or followed. o Pathological lymph nodes with <15 mm, but ³10 mm short axis are considered non- measurable. o Pathological lymph nodes with ³15 mm short axis are considered measurable and can be selected as target lesions; however, lymph nodes should not be selected as target lesions when other suitable target lesions are available, o Measurable lesions up to a maximum of 2 lesions per organ and 5 lesions in total, representative of all involved organs, should be identified as target lesions, and recorded and measured at baseline. These lesions should be selected based on their size (lesions with the longest diameter) and their suitability for accurate repeated measurements (either by imaging techniques or clinically). iRECIST Guidelines iRECIST is based on RECIST 1.1, but adapted to account for the unique tumor response seen with immunotherapeutic drugs. iRECIST will be used to assess tumor response and progression, and make treatment decisions.

REFERENCES

Chen DS, Mellman I. Oncology meets immunology: the cancer-immunity cycle. Immunity. 2013; 39:1- 10.

Wolchok JD, Kluger H, Callahan MK, et al. Nivolumab plus ipilimumab in advanced melanoma. N Engl J Med. 2013; 369:122-133.

Li F, Ravetch JV. Inhibitory FCy receptor engagement drives adjuvant and anti-tumor activities of agonistic CD40 antibodies. Science 2011; 333: 1030-1034. Paulos CM, Carpenito C, Plesa G, et al. The inducible costimulator (ICOS) is critical for the development of human Thl7 cells. Sci Transl Med. 2010; 2:55ra78.

Wakamatsu El, Mathis D, Benoist C. Convergent and divergent effects of costimulatory molecules in conventional and regulatory CD4+ T cells. PNAS. 2013; 110: 1023-1028.

Sharpe AH, Freeman GJ. The B7-CD28 Superfamily. Nat Rev Immunol. 2002; 2:116-126.

Ara G, Baher A, Storm N, et al. Potent activity of soluble B7RP-1-Fc in therapy of murine tumors in syngeneic hosts. Int J Cancer. 2003; 103:501-507.

Carthon BC, Wolchok JD, Yuan J, et al. Preoperative CTLA-4 blockade: Tolerability and immune monitoring in the setting of a presurgical clinical trial. Clin Cancer Res. 2010; 16:2861-2871.

Chen H, Liakou Cl, Kamat A, et al. Anti-CTLA-4 therapy results in higher CD4+ICOShi T cell frequency and IFN-gamma levels in both nonmalignant and malignant prostate tissues. Proc Natl Sci USA. 2009;106:2729-2734.

Cubas R, Moskalenko M, Cheung J, et al. Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; September 25-28, 2016; New York, NY DOI: 10.1158/2326-6066. IMM2016-A114 Published November 2016.

Emens LA, Middleton G. The interplay of immunotherapy and chemotherapy: harnessing potential synergies. Cancer Immunology Research. 2015 May l;3(5):436-43.

Liakou Cl, Kamat A, Tang D, et al. CTLA-4 blockade increases IFN-gamma producing CD4+ICOShi cells to shift the ratio of effector to regulatory T cells in cancer patients. Proc Natl Acad Sci USA. 2008; 105:14987-14992.

Vonderheide RH, LoRusso PM, Khalil M, et al. Tremelimumab in combination with exemestane in patients with advanced breast cancer and treatment-associated modulation of inducible costimulator expression on patient T cells. Clin Cancer Res. 2010; 16:3485-3494.

Eisenhauer EA, Therasse P, Bogaerts J, et al. New response evaluation criteria in solid tumours: revised RECIST guidelines (version 1.1). Eur J Cancer. 2009; 45:228-47. Burtness B, Harrington Id, Greil R, Soulieres D, Tahara M, de Castro Jr G, Psyrri A, Baste N, Neupane P, Bratland A, Fuereder T, Hughes BGM, Mesia R, Ngamphaiboon N, Rordorf, Wan Ishak WZ, Hong RL, Gonzalez Mendoza R, Roy A, Zhang Y, Gumuscu B, Cheng JD, Jin F, Rischin D, on behalf of the KEYNOTE-048 Investigators*. Pembrolizumab alone or with chemotherapy versus cetuximab with chemotherapy for recurrent or metastatic squamous cell carcinoma of the head and neck (KEYNOTE- 048): a randomised, open-label, phase 3 study. Lancet 394:1915-28, 2019.

Ji Y, Liu P, Li Y, et al. A modified toxicity probability interval method for dose-finding trials. Clin Trials. 2010; 7:653-663.

Claims

1. A method of treating cancer in a human in need thereof, the method comprising administering to the human an agonist ICOS binding protein or antigen binding portion thereof at a dose of about 0.08 mg to about 240 mg and administering to the human a chemotherapeutic agent.

2. An agonist ICOS binding protein or antigen binding portion thereof for use in treating cancer, wherein the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg and is administered with a chemotherapeutic agent.

3. A combination comprising an agonist ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent for use in treating cancer, wherein the agonist ICOS binding protein or antigen binding portion thereof is to be administered at a dose of about 0.08 mg to about 240 mg.

4. Use of an agonist ICOS binding protein or antigen binding portion thereof in the manufacture of a medicament for treating cancer, wherein the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg to about 240 mg and is administered with a chemotherapeutic agent.

5. A pharmaceutical kit comprising about 0.08 mg to about 1000 mg of an agonist ICOS binding protein or antigen binding portion thereof and a chemotherapeutic agent.

6. The method, agonist ICOS binding protein, comination, use, or pharmaceutical kit of any one of claims 1-5, wherein the agonist ICOS binding protein or antigen binding portion thereof comprises one or more of: CDRH1 as set forth in SEQ ID NO:l; CDRH2 as set forth in SEQ ID NO:2; CDRH3 as set forth in SEQ ID NO:3; CDRL1 as set forth in SEQ ID NO:4; CDRL2 as set forth in SEQ ID NO:5 and/or CDRL3 as set forth in SEQ ID NO:6 or a direct equivalent of each CDR wherein a direct equivalent has no more than two amino acid substitutions in said CDR.

7. The method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of any one of claims 1-6, wherein the agonist ICOS binding protein or antigen binding portion thereof comprises a VH domain comprising an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:7 and/or a VL domain comprising an amino acid sequence at least 90% identical to the amino acid sequence as set forth in SEQ ID NO:8 wherein said agonist ICOS binding protein specifically binds to human ICOS.

8. The method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of any one of claims 1-7, wherein the agonist ICOS binding protein is a monoclonal antibody.

9. The method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of any one of claims 1-8, wherein the agonist ICOS binding protein is a humanized or fully human monoclonal antibody.

10. The method, agonist ICOS binding protein, combination, use, or pharmaceutical kit of any one of claims 1-9, wherein the agonist ICOS binding protein comprises an hIgG4PE scaffold.

11. The method, agonist ICOS binding protein, combination or use, any one of claims 1-4 and 6- 10, wherein the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 0.08 mg, about 0.24 mg, about 0.8 mg, about 2.4 mg, about 8 mg, about 24 mg, about 80 mg, or about 240 mg.

12. The method, agonist ICOS binding protein, combination or use of any one of claims 1-4 and 6-11, wherein the agonist ICOS binding protein or antigen binding portion thereof is administered at a dose of about 24 mg.

13. The method, agonist ICOS binding protein, acombination or use of any one of claims 1-4 and 6-12, wherein the agonist ICOS binding protein or antigen binding portion thereof is administered once every three weeks or every 6 weeks.

14. The method, agonist ICOS binding protein, combination or use of any one of claims 1-4 and 6-13, wherein the agonist ICOS binding protein or antigen binding portion thereof and/or the chemotherapeutic agent is administered via IV infusion.

15. The method, agonist ICOS binding protein, combination or use of any one of claims 1-4 and 6-14, wherein the cancer is a solid tumor.

16. The method, agonist ICOS binding protein, combination or use of any one of claims 1-4 and 6-15, wherein the cancer is selected from NSCLC, HNSCC, urothelial cancer, cervical cancer and melanoma.

17. The method, agonist ICOS binding protein, combination, use, or kit of any one of claims 1-16, wherein the chemotherapeutic agent is docetaxel, carboplatin, cisplatin, paclitaxel, fluorouracil, pemetrexed, gemcitabine or a combination thereof.

18. The method, agonist ICOS binding protein, combination or use of claim 17, wherein docetaxel is administered at a dose of about 30 to about 100 mg/m² or about 75 mg/m²; wherein carboplatin is administered at a dose of about AUC 4-7 mg/ml per min or about AUC 5 mg/ml per min; wherein cisplatin is administered at a dose of about 20 mg/m² to about 120 mg/m² or 100 mg/m²; wherein paclitaxel is administered at a dose of about 135 mg/m² to about 225 mg/m² or 200 mg/m²; wherein fluorouracil is administered at a dose of about 200 mg/m² to about 1200 mg/m² or 1000 mg/m²; wherein pemetrexed is administered at a dose of about 500 mg/m²; or wherein gemcitabine is administered at a dose of about 1000 mg/m² to about 1250 mg/m² or 1250 mg/m².

19. The method, agonist ICOS binding protein, combination, use, or kit of any one of claims 1-18, wherein the chemotherapeutic agent is a platinum-based chemotherapy doublet.

20. The method, agonist ICOS binding protein, combination, use, or kit of any one of claims 1-19, wherein the chemotherapeutic agent is a doublet of pemetrexed and carboplatin, paclitaxel and carboplatin or gemcitabine and carboplatin.

21. The method, agonist ICOS binding protein, combination, use, or kit of any one of claims 1-20, wherein the chemotherapeutic agent is a doublet of fluorouracil and carboplatin or fluorouracil and cisplatin.

22. The method, agonist ICOS binding protein, combination or use of any one of claims 1-4 and 6-21, wherein the agonist ICOS binding protein or antigen binding portion thereof and the chemotherapeutic agent is administered concurrently and/or sequentially.