Use of C / EBP-beta antagonists and immunomodulators

Abstract

Methods are provided for administering peptide antagonists of CCAAT / enhancer-binding protein beta (C / EBPβ) in combination with immunomodulatory agents.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Application No. 63 / 414,397, filed October 7, 2022.

[0002] Electronic Sequence Listing Reference The contents of the electronic sequence listing (file name: Sapience020WO1.xml, size: 4,534 bytes, and creation date: October 6, 2023) are incorporated herein by reference in their entirety. [Background technology]

[0003] CCAAT / enhancer-binding protein β (C / EBPβ) is a basic leucine zipper (bZIP) transcription factor that causes aberrant gene activation in many cancers. Upregulated or hyperactivated C / EBPβ drives carcinogenesis by promoting tumor survival and proliferation and is an essential regulator of the immunosuppressive environment (Homma 2006, Ruffell 2009). Specifically, C / EBPβ regulates macrophage differentiation, promoting the expression of M2 myeloid-derived suppressor cells (MDSCs), which are involved in suppressing antitumor immunity and correlate with poor prognosis (Marigo 2010). Remigrating tumor-associated macrophages (TAMs) from an M2 to an M1 phenotype represents a potential strategy to enhance antitumor immunity.

[0004] Although immune checkpoint inhibitors (ICIs) have shown unprecedented success in immunogenic tumors such as melanoma, many patients with advanced solid tumors respond poorly or not at all to ICI therapy. Refractory tumors are generally classified as "cold" and demonstrate either a lack of T cell infiltration into the tumor stroma or an immunosuppressive environment that eliminates both T cell infiltration and activity. Because the immunosuppressive tumor microenvironment (TME) represents the greatest obstacle to successfully advancing ICI therapy in solid tumors, the ability to convert the TME to an immunoactive state shows great promise for improving responses.

[0005] ST101 is a novel peptide antagonist that prevents C / EBPβ dimerization and inhibits C / EBPβ-dependent gene expression. The observed responses in melanoma and other tumors motivated us to evaluate the effect of ST101 on macrophage differentiation. Summary of the Invention

[0006] Some of the main aspects of the present invention are summarized below. Additional aspects are described in the detailed description, examples, drawings, and claims sections of this disclosure. The statements in each section of this disclosure are intended to be interpreted with reference to the other sections. Furthermore, the various embodiments described in each section of this disclosure can be combined in various different ways, and all such combinations are intended to be within the scope of the present invention.

[0007] We demonstrate that ST101 exposure reprograms M2-like macrophages to an immunopromoting M1-like phenotype both in vitro and in vivo. Macrophage repolarization leads to the restoration of cytotoxic T cell activity and enhances antitumor responses in vivo in concert with anti-PD-1 therapy in a triple-negative breast cancer model. These results identify ST101 as a novel approach to enhance macrophage antitumor activity and support its utility for combination strategies in cancers that respond poorly to ICIs.

[0008] Accordingly, the present disclosure provides a combination procedure for administering a peptide antagonist of CCAAT enhancer-binding protein β (C / EBPβ) and an immunomodulatory agent. In one embodiment, a method for inhibiting the growth of a solid tumor in a subject is provided, the method comprising combination therapy with (i) a pharmaceutical composition comprising an effective amount of a peptide antagonist of C / EBPβ and (ii) a pharmaceutical composition comprising an effective amount of an immunomodulatory agent, wherein the pharmaceutical composition comprising the antagonist of C / EBPβ and the pharmaceutical composition comprising the immunomodulatory agent are administered to the subject simultaneously or separately.

[0009] Another embodiment is a method for reducing the volume of a solid tumor in a subject, the method comprising combination therapy using (i) a pharmaceutical composition comprising an effective amount of a peptide antagonist of C / EBPβ, and (ii) a pharmaceutical composition comprising an effective amount of an immunomodulatory agent, wherein the pharmaceutical composition comprising the antagonist of C / EBPβ and the pharmaceutical composition comprising the immunomodulatory agent are administered to the subject simultaneously or separately.

[0010] Further embodiments provide methods of treating a solid tumor in a subject, the methods comprising combination therapy using (i) a pharmaceutical composition comprising an effective amount of a peptide antagonist of C / EBPβ, and (ii) a pharmaceutical composition comprising an effective amount of an immunomodulatory agent, wherein the pharmaceutical composition comprising the antagonist of C / EBPβ and the pharmaceutical composition comprising the immunomodulatory agent are administered to the subject simultaneously or separately.

[0011] Also provided are pharmaceutical compositions comprising an effective amount of a peptide antagonist of C / EBPβ and an effective amount of an immunomodulatory agent for use in combination therapy methods for inhibiting the growth of a solid tumor in a subject, reducing solid tumor volume in a subject, and / or treating a solid tumor in a human subject.

[0012] In a preferred embodiment, the antagonist of C / EBPβ is ST101 and the immunomodulatory agent is a PD-1 inhibitor.

[0013] In some embodiments, the solid tumor is melanoma, carcinoma, or sarcoma. In certain embodiments, the subject has been diagnosed with locally advanced or metastatic breast cancer (LA / MBC), melanoma, glioblastoma (GBM), or castration-resistant prostate cancer (CRPC).

[0014] In one embodiment, the peptide antagonist comprises the D-amino acid sequence VAEAREELERLEARLGQARGEL (SEQ ID NO: 1). In another embodiment, the peptide antagonist comprises the amino acid sequence LEGRAQGLRAELRELEERAEAV (SEQ ID NO: 3). In some embodiments, the peptide antagonist is a cell-penetrating peptide. For example, the peptide antagonist can comprise a cell-penetrating sequence. In a specific embodiment, the peptide antagonist is ST101 (SEQ ID NO: 2).

[0015] In some embodiments, the peptide antagonist is administered to the subject at a dose of about 0.5-16 mg / kg. In one embodiment, the dose of ST101 is about 500 mg.

[0016] In certain aspects of the invention, the immunomodulatory agent is an antibody or antibody-drug conjugate selected from the group consisting of, for example, amivantamab, belantamab mafodotin-blmf, bevacizumab, cetuximab, denosumab, dinutuximab, enfortumab vedotin-ejfv, margetuximab, naxitamab-gqgk, necitumumab, panitumumab, pertuzumab, ramucirumab, sacituzumab govitecan-hziy, tebentafsp-tebn, tisotumab vedotin, trastuzumab, trastuzumab deruxtecan, and trastuzumab emtansine. In certain aspects, the antibody is an anti-PD-1 antibody.

[0017] In one aspect of the present invention, the immunomodulatory agent is selected from the group consisting of a checkpoint inhibitor, a cytokine, and an immunoadjuvant. The checkpoint inhibitor can target, for example, at least one of PD-1, PD-L1, CTLA-4, or LAG-3. In certain embodiments, the checkpoint inhibitor is atezolizumab, avelumab, cemiplimab, dostarlimab, durvalumab, ipilimumab, nivolumab, pembrolizumab, relatimab, or a combination thereof.

[0018] In one embodiment, the immunomodulatory agent is a cytokine that targets, for example, at least one of the IL-2 pathway, the IL-2R pathway, the IFNAR1 pathway, or the IFNAR2 pathway. Exemplary cytokines for use in the methods of the invention include aldesleukin, granulocyte-macrophage colony-stimulating factor, interferon alpha-2a, interferon alpha-2b, and pegylated interferon alpha-2b.

[0019] The immunomodulatory agent can be an immunoadjuvant. In one embodiment, the immunoadjuvant targets the Toll-like receptor 7 pathway or the Toll-like receptor 3 pathway. Specific examples of immunoadjuvants include imiquimod and poly-ICLC.

[0020] In one embodiment, the pharmaceutical composition comprising an antagonist of C / EBPβ and / or the pharmaceutical composition comprising an immunomodulatory agent is administered parenterally, for example, intravenously.

[0021] In some embodiments, the pharmaceutical composition comprising an antagonist of C / EBPβ and the pharmaceutical composition comprising an immunomodulatory agent are administered to a subject on different days.

[0022] In certain embodiments, the pharmaceutical composition comprising the antagonist of C / EBPβ is administered once weekly for at least 3 weeks, or once every two weeks for at least 4 weeks, hi certain embodiments, the pharmaceutical composition comprising the immunomodulatory agent is administered once weekly for at least 3 weeks, or once every two weeks for at least 4 weeks, or once every three weeks for at least 6 weeks. [Brief explanation of the drawings]

[0023] [Figure 1] FIG. 1 shows a schematic diagram of the protocol for the human peripheral blood mononuclear cell (hPBMC) M1 / M2 polarization model described in Example 1. [Figure 2A] Figure 1 shows that ST101 shifts the M2 program to an M1-like phenotype in cultured hPBMCs from donor 1. Cells were treated with the indicated concentrations of ST101, and CD163 and CD68 expression was measured by flow cytometry. [Figure 2B] Figure 1 shows that ST101 shifts the M2 program to an M1-like phenotype in cultured hPBMCs from donor 2. Cells were treated with the indicated concentrations of ST101, and CD163 and CD68 expression was measured by flow cytometry. [Figure 3A] Figure 1 shows that ST101 substantially reduces hPBMC-derived M2 cells in cultures from donor 1. Cells were treated with the indicated concentrations of ST101, and expression of CD163 and CD68 was measured by flow cytometry. [Figure 3B] Figure 1 shows that ST101 substantially reduces hPBMC-derived M2 cells in cultures from donor 1. The ratio of M2 to M1 cells is shown after treatment with the indicated concentrations of ST101. [Figure 3C] Figure 1 shows that ST101 substantially reduces hPBMC-derived M2 cells in cultures from donor 2. Cells were treated with the indicated concentrations of ST101, and expression of CD163 and CD68 was measured by flow cytometry. [Figure 3D]Figure 1 shows that ST101 substantially reduces hPBMC-derived M2 cells in cultures from donor 2. The ratio of M2 to M1 cells is shown after treatment with the indicated concentrations of ST101. [Figure 4] A-B show that ST101 inhibits M2 macrophage polarization in hPBMCs from donor 1. Cells were treated with the indicated concentrations of ST101, and expression of CD163 (A) and CD68 (B) was measured by flow cytometry. [Figure 5A] 1 shows that ST101 affects the tumor microenvironment. Nanostring analysis of tumor samples from human patients treated with ST101 shows a decrease in IL6 signaling. [Figure 5B] 1 shows that ST101 affects the tumor microenvironment. Nanostring analysis of tumor samples from human patients treated with ST101 shows an increase in tumor-infiltrating macrophages. [Figure 5C] 1 shows that ST101 affects the tumor microenvironment. Nanostring analysis of tumor samples from human patients treated with ST101 shows a decrease in regulatory T cells. [Figure 6] We show that ST101 restores activation of cultured human CD8+ T cells co-incubated with immunosuppressive M2 macrophages. [Figure 7] 1 shows enhanced tumor growth inhibition (TGI) when treated with a combination of a C / EBPβ antagonist (ST101) and an immune checkpoint inhibitor (anti-PD-1 antibody). DETAILED DESCRIPTION OF THE INVENTION

[0024] In order to facilitate an understanding of the present invention, certain terms are first defined. Further definitions are set forth throughout this disclosure. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0025] Any headings provided herein are to be understood with reference to the specification as a whole and are not limitations of the various aspects or embodiments of the present invention, and accordingly, the terms defined immediately below are more clearly defined with reference to the specification as a whole.

[0026] All references cited in this disclosure are incorporated herein by reference in their entirety. Additionally, any manufacturer's instructions or catalogs for any products cited or mentioned herein are incorporated by reference. Any document incorporated by reference herein, or any teaching therein, may be used in the practice of the present invention. Any document incorporated by reference herein is not admitted to be prior art.

[0027] I. Definition The terms or terminology of this disclosure are for purposes of description and not limitation, and the terms or terms herein will be interpreted by one of ordinary skill in the art in light of the teachings and guidance.

[0028] As used in this specification and claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. The term "a" (or "an") and the terms "one or more" and "at least one" can be used interchangeably.

[0029] Furthermore, "and / or" is to be interpreted as a specific disclosure of each of the two specified features or elements without regard to the presence or absence of the other. Thus, the term "and / or" used in phrases such as "A and / or B" is intended to include A and B, A or B, A alone, and B alone. Similarly, the term "and / or" used in phrases such as "A, B, and / or C" is intended to include A, B, and C, A, B, or C, A or B, A or C, B or C, A and B, A and C, B and C, A alone, B alone, and C alone.

[0030] Whenever an embodiment is described using the term "comprising," similar embodiments are included except those described using the terms "consisting of" and / or "consisting essentially of."

[0031] Units, prefixes, and symbols are shown in the format accepted by the International System of Units (SI). Numerical ranges are inclusive of the numbers defining the range, and any individual value provided herein can serve as an endpoint of a range that includes other individual values ​​provided herein. For example, a set of values ​​such as 1, 2, 3, 8, 9, and 10 is also a disclosure of numerical ranges such as 1 to 10, 1 to 8, 3 to 9, etc. Similarly, a disclosed range is a disclosure of each individual value (i.e., intermediate value) encompassed by the range, including integers and fractions. For example, the recited range 5 to 10 is also a disclosure of 5, 6, 7, 8, 9, and 10 individually, as well as 5.2, 7.5, 8.7, etc.

[0032] Unless otherwise specified, the phrase "at least" or "about" preceding a series of elements is understood to refer to every element of the series. The term "about" preceding a numerical value includes ±10% of the stated value. For example, a concentration of about 1 mg / mL includes 0.9 mg / mL to 1.1 mg / mL. Similarly, a concentration range of about 1% to 10% (w / v) includes 0.9% (w / v) to 11% (w / v).

[0033] The terms "polypeptide," "peptide," and "protein" are used interchangeably and refer to polymers of amino acids of any length, and salts thereof. The polymers can be linear or branched, can include modified amino acids, and can be truncated with non-amino acids. Except where otherwise noted, for example, for abbreviations of unusual or unnatural amino acids described herein, the three-letter and one-letter abbreviations used in the art are used herein to represent amino acid residues. Amino acids are L-amino acids unless preceded by a "D" or in lower case. Groups or strings of amino acid abbreviations are used to represent peptides. Except where otherwise noted, peptides are depicted with the N-terminus on the left, and sequences are written from the N-terminus to the C-terminus.

[0034] "Retro-inverso" peptides have an amino acid sequence that is reversed relative to a reference L-amino acid sequence and are composed of all D-amino acids (inverting the α-center chirality of the amino acid subunits) to help maintain a side chain topology similar to that of the original L-amino acid peptide.

[0035] An "isolated" molecule is a molecule in a form not found in nature, including a purified molecule.

[0036] An "active agent" is a component intended to provide a biological activity. The active agent can be associated with one or more other components. An active agent that is a peptide can also be referred to as an "active peptide."

[0037] An "effective amount" of an active agent is an amount sufficient to accomplish a specific stated purpose.

[0038] The term "pharmaceutical composition" refers to a preparation that is in a form that allows the biological activity of the active ingredient to be effective and does not contain additional components that exhibit unacceptable toxicity to the subject to which the formulation is administered. Such compositions can contain a pharmaceutically acceptable carrier, such as saline, which can be sterile. Suitable pharmaceutical compositions can contain one or more buffers (e.g., acetate buffer, phosphate buffer, or citrate buffer), surfactants (e.g., polysorbates), stabilizers (e.g., polyols or amino acids), preservatives (e.g., sodium benzoate), and / or other conventional solubilizers or dispersants.

[0039] A "subject" or "individual" or "animal" or "patient" or "mammal" is any subject for which diagnosis, prognosis, or treatment is desired, particularly a mammalian subject. Mammalian subjects include, for example, humans, non-human primates, dogs, cats, pigs, cows, horses, rodents including rats and mice, rabbits, and other humans, domestic animals, livestock animals, sport animals, and laboratory animals.

[0040] A "control patient" is a subject who has not received the treatment of the present invention. A "control population" or "control patient population" is a group of subjects who have not received the treatment of the present invention. The control patients or subjects in the control population have the same disease or disorder as the subjects being compared to the control patients or control population. For example, the clinical outcomes of cancer patients who have used the compositions or methods of the present invention are compared to the average (median) outcomes of subjects with the same type of cancer without the use of the pharmaceutical compositions or methods of the present invention. In some embodiments, the control patients or patients in the control population have received treatment other than the treatment of the present invention, e.g., standard treatment.

[0041] Terms such as "treating" or "treatment" or "to treat" or "alleviating" or "to alleviate" refer to therapeutic measures that cure, delay, reduce the symptoms of, and / or halt the progression of a diagnosed pathological condition or disorder. In certain embodiments, a subject has been successfully "treated" for a disease or disorder if the subject experiences a complete, partial, or transient alleviation or elimination of at least one symptom or measurable physical parameter associated with the disease or disorder.

[0042] An "antagonist" is a substance that prevents, blocks, inhibits, neutralizes, or reduces the biological activity or effect of another molecule, e.g., a receptor or ligand.

[0043] The terms "inhibit," "block," and "suppress" can be used interchangeably and refer to any statistically significant decrease in occurrence or activity, including a complete block of occurrence or activity. For example, "inhibition" can refer to about a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% decrease in activity or frequency of occurrence. An "inhibitor" is a molecule, factor, or substance that statistically significantly decreases the occurrence or activity of a process, pathway, or molecule.

[0044] A "tumor" or "solid tumor" is a mass of neoplastic cells, such as cancer cells. The terms "progressive," "metastatic," and "progressive / metastatic" are used interchangeably to describe cancers in which malignant cells have migrated from the original tumor to another location in the patient's body, e.g., another organ.

[0045] "Tumor cells" or "neoplasms" are usually mutated / transformed in some way, resulting in abnormal growth compared to normal cells or tissues of the same type. Neoplasms include morphological irregularities as well as pathological growth. Tumor cells can be benign or malignant. Malignant neoplasms, i.e., cancers, are distinguished from benign neoplasms in that they demonstrate a loss of cellular differentiation and orientation and possess properties of invasion and metastasis.

[0046] II. Peptides and Compositions C / EBPβ C / EBPβ is a prime target for the development of peptide antagonists because it relies on the interaction of cofactors with a basic leucine zipper (bZIP). To associate with DNA and transactivate gene expression, C / EBPβ dimerizes with binding partners through interactions between their bZIP domains. In addition to homodimerization, C / EBPβ heterodimerizes with bZIP-containing transcription factors such as Jun / Fos, C / EBPγ (Huggins 2013), Delta-interacting protein A (Bezy 2005), and the CREB / ATF family (Zhao 2014).

[0047] Activating transcription factor 5 (ATF5) is a CREB / ATF factor that has been shown to associate with and activate C / EBPβ in HEK293 and HCT116 cancer cells, transactivating a pro-survival phenotype (Zhang 2015). ATF5 is highly expressed in many cancers, including gliomas, and contributes to the oncogenic phenotype by driving the overexpression of Bcl-2 family proteins and survivin, but is rarely observed in differentiated cell types (Sheng 2010). Overexpression of a truncated bZIP domain of ATF5 lacking the DNA-binding domain in gliomas and other tumor cells resulted in cytotoxicity of cancer cells (Angelastro 2006), and administration of a peptide containing the truncated bZIP domain produced similar results (Cates 2016, Karpel-Massler 2016).

[0048] C / EBPβ antagonist peptide In some embodiments, the methods of the invention involve treating a patient with a solid tumor with a combination therapy comprising an effective amount of a peptide antagonist of C / EBPβ and an immunomodulatory agent. In one embodiment, the peptide antagonist of C / EBPβ comprises the D-amino acid sequence VAEAREELERLEARLGQARGEL (SEQ ID NO: 1), a retro-inverso variant of the wild-type ATF5 bZIP domain. Peptide antagonists of C / EBPβ can be designed, for example, as described in Example 1 of WO 2021 / 262604.

[0049] The peptide antagonist of C / EBPβ can be a cell-penetrating peptide. In one embodiment, the peptide comprises a cell-penetrating domain. Numerous cell-penetrating peptide sequences have been described and characterized in the literature (see WO 2019 / 136125). In one embodiment, the peptide is a cyclic peptide. Cyclic peptides can enter cells via passive diffusion, endocytosis / endosomal escape, or other mechanisms, for example, using hydrocarbon staples (Bernal 2007, Bird 2016) or other cyclization methods known in the art (Dougherty 2019). Peptides can also be delivered to cells via mechanisms that utilize cellular receptors, for example, integrin-targeting RGD-like sequences. Alternatively, peptides can be delivered to cells via vesicles such as exosomes or liposomes, or micelles.

[0050] The ability of peptides based on the native ATF5 bZIP domain to antagonize the activity of C / EBPβ can be measured by methods described herein, for example, as described in Example 2 of WO 2021 / 262604. The cytotoxic activity of peptide antagonists of C / EBPβ can be measured in vitro by known assays and / or in vivo using known tumor models, for example, as described in WO 2019 / 136125.

[0051] ST101 is an all-D amino acid peptide that exhibits potent antitumor activity in vitro and in vivo, as well as resistance to proteolysis. In particular, we have previously demonstrated the cytotoxicity of ST101 in HL60 (promyelocytic leukemia), AML14 (acute myeloid leukemia), SET2 (megakaryoblastic leukemia), A375 (melanoma), MCF7 (breast cancer), U87 (glioblastoma), U251 (glioblastoma), DU145 (prostate cancer), A549 (lung cancer), peripheral blood mononuclear cells (PBMC), and bone marrow mononuclear cells (BMMC) (see WO 2019 / 136125). In addition, subcutaneous administration of ST101 significantly reduced tumor volume in xenograft mouse models using A375, HL60, MCF7, and U251 cells (see WO 2019 / 136125).

[0052] ST101 is composed of a modified domain based on the ATF5 bZIP domain and the Antennapedia penetratin domain, allowing cell permeability. The D-amino acid sequence of ST101 is: [ka] where the cell-penetrating region is italicized. ST101 promotes cytotoxic activity in tumor cells by preventing the association of C / EBPβ with anti-apoptotic transcription factors (see WO 2021 / 262604).

[0053] We demonstrate for the first time that ST101 alters the tumor microenvironment and reprograms macrophage differentiation toward an M1 phenotype. The potential effects of ST101 on antitumor immunity led us to test whether combining a C / EBPβ antagonist, such as ST101, with an immunomodulatory agent, such as an immune checkpoint inhibitor, would enhance antitumor activity. The combination of ST101 and PD-1 unexpectedly reduced tumor volume in a mouse model of triple-negative breast cancer, demonstrating an approximately 1.8- to 2.0-fold improvement in activity compared to either agent alone (see Example 3, Figure 7).

[0054] IV. Methods for preparing C / EBPβ antagonists Peptide antagonists of C / EBPβ can be chemically synthesized, for example, using solid-phase peptide synthesis or solution-phase peptide synthesis, or a combination of both. Synthesis can optionally be performed as peptide fragments that are then combined chemically or enzymatically.

[0055] Alternatively, peptide antagonists of C / EBPβ can be expressed using recombinant methods. For example, a nucleic acid molecule encoding ST101 can be constructed by chemical synthesis using an oligonucleotide synthesizer. The nucleic acid molecule can be designed based on the amino acid sequence of ST101 and the selection of appropriate codons in the host cell in which recombinant ST101 will be produced. Standard methods can be used to synthesize nucleic acid molecules encoding peptide antagonists of C / EBPβ, such as ST101.

[0056] Once prepared, the nucleic acid encoding the peptide can be inserted into an expression vector and operably linked to appropriate expression control sequences to express the peptide in a desired host. To obtain high levels of expression of the peptide, the nucleic acid can be operably linked to or associated with transcriptional and translational expression control sequences that are functional in the selected expression host.

[0057] Those skilled in the art will recognize a wide variety of expression host / vector combinations. Useful expression vectors for eukaryotic hosts include, for example, vectors containing expression control sequences from SV40, bovine papillomavirus, adenovirus, and cytomegalovirus. Useful expression vectors for bacterial hosts include known bacterial plasmids, e.g., E. coli-derived plasmids, including pCR1, pBR322, pMB9, and their derivatives, broad-host-range plasmids, e.g., M13, and filamentous single-stranded DNA phages.

[0058] Suitable host cells include prokaryotes, yeast, insects, or higher eukaryotic cells under the control of a suitable promoter. Prokaryotes include gram-negative or gram-positive bacteria, such as E. coli or Bacillus. Higher eukaryotic cells can be established, or cell lines of mammalian origin, such as Pichia pastoris, 293 cells, COS-7 cells, L cells, C127 cells, 3T3 cells, Chinese hamster ovary (CHO) cells, HeLa cells, and BHK cells. Cell-free translation systems can also be used.

[0059] Peptides can be purified using methods such as, for example, reverse-phase high performance liquid chromatography (RP-HPLC), multicolumn countercurrent solvent gradient purification (MCSGP), and ion exchange chromatography.

[0060] immunomodulators The methods of the present invention involve combination therapy comprising the administration of a C / EBPβ antagonist and an immunomodulatory agent. The terms "immunomodulator," "immunomodulating agent," and "immunotherapeutic agent" are used interchangeably and refer to active agents that stimulate or suppress a subject's immune system to combat disease or infection. In the context of the present invention, the disease is a neoplasm or cancer.

[0061] Immunomodulatory agents include, for example, immune checkpoint inhibitors, cytokines, targeted antibodies and drug-antibody conjugates, adjuvants such as imiquimod and polyinosinic-polycytidylic acid (poly-ICLC), oncolytic viruses such as T-VEC, and small molecule drugs such as thalidomide, lenalidomide, or pomalidomide.

[0062] Immune checkpoint inhibitors target immune checkpoint proteins or their ligand(s). Immune checkpoint proteins include, but are not limited to, cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4), also known as CD152, programmed cell death protein 1 (PD-1), also known as CD279, lymphocyte activation gene 3 (LAG-3), also known as CD223, T-cell immunoglobulin mucin (TIM-3), also known as HAVcr2, and T-cell immunoreceptor with Ig and ITIM domains (TIGIT). Examples of immune checkpoint inhibitors include atezolizumab, avelumab, cemiplimab, dostarlimab, durvalumab, ipilimumab, nivolumab, pembrolizumab, and relatolimab.

[0063] Some cytokines, such as interferons, can disrupt cancer cell division and slow tumor growth. Other cytokines, such as interleukins (ILs), stimulate the growth and proliferation of immune cells. Examples of immunomodulatory cytokines include aldesleukin, granulocyte-macrophage colony-stimulating factor, interferon alpha-2a, interferon alpha-2b, and pegylated interferon alpha-2b.

[0064] Targeting antibodies can be customized to target antigens on the surface of cancer cells to disrupt cancer activity, particularly uncontrolled growth. Some targeting antibodies are conjugated to anti-cancer drugs. Other targeting antibodies are bispecific, binding to both cancer cells and T cells, for example, to enhance anti-cancer immune responses. Examples of targeted antibodies and drug-antibody conjugates include amivantamab, belantamab mafodotin-BLMF, bevacizumab, cetuximab, denosumab, dinutuximab, enfortumab vedotin-EJFV, margetuximab, naxitamab-GQGK, necitumumab, panitumumab, pertuzumab, ramucirumab, sacituzumab govitecan-HZIY, tebentafsp-TEBN, tisotumab vedotin, trastuzumab, trastuzumab deruxtecan, and trastuzumab emtansine.

[0065] Compositions and Administration In certain aspects, the present invention provides combination methods that include administering a composition, e.g., a pharmaceutical composition, comprising an effective amount of a peptide antagonist of C / EBPβ, such as ST101, and administering a composition, e.g., a pharmaceutical composition, comprising an effective amount of an immunomodulatory agent, such as a checkpoint inhibitor. Methods of administering peptide antagonists of C / EBPβ are described, for example, in WO 2021 / 262604.

[0066] Because the combination therapies of the invention demonstrate synergistic effects between a C / EBPβ antagonist and an immunomodulatory agent, the effective amount of each active agent can, in some embodiments, be less than the effective amount of each active agent when administered individually. For example, the effective amount of each of the peptide antagonist of C / EBPβ and the immunomodulatory agent can be a sub-therapeutic dose.

[0067] The compositions are preferably administered parenterally. Parenteral routes include intravenous (IV), intramuscular, intraperitoneal, intrathecal, and subcutaneous. In certain embodiments, the compositions comprising the peptide antagonist of C / EBPβ and / or the compositions comprising the immunomodulatory agent are administered to a subject by intravenous administration.

[0068] The composition comprising the peptide antagonist of C / EBPβ and the composition comprising the immunomodulatory agent can be the same composition or can be different compositions. When the peptide antagonist and the immunomodulatory agent are contained in two separate compositions, the two compositions can be administered to a subject at the same time or at different times, including on different days.

[0069] The composition comprising the peptide antagonist and the composition comprising the immunomodulator are each administered multiple times to a subject. In one embodiment, the administration of the peptide antagonist of C / EBPβ can occur once per week for a period of at least 3 weeks (i.e., 3 doses), 6 weeks (i.e., 6 doses), 9 weeks, 12 weeks, 3 months, 6 months, 9 months, or 12 months. In another embodiment, the administration can occur once every two weeks for a period of at least 4 weeks (i.e., 2 doses), 8 weeks (i.e., 4 doses), 12 weeks, 3 months, 6 months, 9 months, or 12 months. In some embodiments, a patient may receive a peptide antagonist of C / EBPβ once a week for a period of at least 3 weeks, 6 weeks, 9 weeks, 12 weeks, 3 months, 6 months, 9 months, or 12 months, followed by once every two weeks for a period of at least 4 weeks, 8 weeks, 12 weeks, 3 months, 6 months, 9 months, or 12 months.

[0070] In one embodiment, administration of the immunomodulatory agent can occur once per week for a period of at least 3 weeks (i.e., 3 doses), 6 weeks (i.e., 6 doses), 9 weeks, 12 weeks, 3 months, 6 months, 9 months, or 12 months. In another embodiment, administration can occur once every two weeks for a period of at least 4 weeks (i.e., 2 doses), 8 weeks (i.e., 4 doses), 12 weeks, 3 months, 6 months, 9 months, or 12 months. In a further embodiment, administration of the immunomodulatory agent can occur once every three weeks for a period of at least 6 weeks (i.e., 2 doses), 9 weeks (i.e., 3 doses), 12 weeks, 3 months, 6 months, 9 months, or 12 months. In a further embodiment, administration of the immunomodulatory agent can occur once every four weeks for a period of at least 8 weeks (i.e., 2 doses), 12 weeks (i.e., 3 doses), 3 months, 6 months, 9 months, or 12 months. Similarly, administration of the immunomodulatory agent can occur monthly for a period of at least 2 months (i.e., 2 doses), 3 months (i.e., 3 doses), 6 months, 9 months, or 12 months.

[0071] For purposes of this disclosure, "combination therapy" means that a period of treatment involving administration of a peptide antagonist of C / EBPβ overlaps with a period of treatment involving administration of an immunomodulatory agent.

[0072] III. Method of Use The subject in need of the method of the present invention is a patient diagnosed with a solid tumor. For example, the subject may have a locally advanced solid tumor or a metastatic, inoperable tumor. In some embodiments, the subject is afflicted with basal cell carcinoma, bladder cancer, cervical cancer, bile duct cancer, colorectal cancer, endometrial cancer, esophageal cancer, gastric cancer, hepatocellular carcinoma, head and neck cancer, Merkel cell carcinoma, melanoma, renal cell carcinoma, squamous cell carcinoma, triple-negative breast cancer, or urothelial cell carcinoma.

[0073] In certain embodiments, the subject has melanoma, carcinoma, or sarcoma. In one embodiment, the melanoma is cutaneous melanoma or mucosal melanoma. In one embodiment, the carcinoma is an adenocarcinoma, such as bladder adenocarcinoma, colon adenocarcinoma, pancreatic adenocarcinoma, gastric / signet ring adenocarcinoma, or small intestine adenocarcinoma. In one embodiment, the sarcoma is abdominal sarcoma or myofibroblastoma. In certain embodiments of the invention, combination therapy with a peptide antagonist of C / EBPβ, such as ST101, and an immunomodulatory agent can inhibit tumor growth, reduce tumor volume, or a combination thereof.

[0074] The effectiveness of treatment can be assessed by one or more known measures. For example, patients treated with the methods of the present invention may experience outcomes such as prolonged survival, improved progression-free survival, improved duration of response, prolonged remission, reduced risk of recurrence, and / or improved tumor response to treatment, compared to the same outcome(s) in patients not treated with the methods of the present invention, i.e., control patients. The outcome in patients treated with the methods of the present invention can be compared, for example, to the median outcome in a control patient population. The control patient population can be administered a regimen selected from the group consisting of, for example, placebo, surgery, radiation, chemotherapy, immunotherapy, hormone-based therapy, or targeted therapy. In another embodiment, patients treated with the combination therapy of the present invention can be compared to a control patient population treated with either a peptide antagonist of C / EBPβ or an immunomodulatory agent alone. The comparison can be statistically analyzed, for example, using the Wilcoxon signed-rank test or the Kaplan-Meier method.

[0075] The response to treatment is compared with one or more measures of efficacy after treatment regimen, compared with baseline, for example, before treatment with combination therapy.Baseline assessment is preferably carried out within 24, 48 or 72 hours, or within 1, 2, 3 or 4 weeks before first treatment.In one preferred embodiment, baseline assessment is carried out within 24 hours after first treatment.

[0076] "Tumor burden" refers to the total mass or overall size of cancerous tissue in a patient's body. Tumor response can be assessed using measures such as objective remission rate, disease control rate, and duration of response. Depending on the tumor type, these parameters can be determined, for example, by the revised Response Evaluation Criteria in Solid Tumors (RECIST 1.1) (Eisenhauer 2009), the modified Response Evaluation in Neuro-Oncology (mRANO) (Ellingson 2017), or the PCWG3 guidelines (Scher 2016).

[0077] The objective remission rate evaluates the degree of reduction in tumor size, e.g., tumor diameter, which can be determined by clinical examination and / or imaging. If a patient has multiple tumors, tumor size can optionally be expressed as the average diameter of all tumors or the sum of the diameters of all tumors. Surface tumors can be measured clinically, for example, using calipers or by photography and ruler measurement. Imaging methods typically include computed tomography (CT) with contrast agents, X-rays, magnetic resonance imaging (MRI), and positron emission tomography (PET), such as (18)F-fluorodeoxyglucose PET. In one preferred embodiment, CT is used to evaluate tumor response, for example, in LA / MBC patients or melanoma patients. In another preferred embodiment, MRI, such as gadolinium-enhanced MRI, is used to evaluate tumor response, for example, in GBM patients. Thus, in one aspect, the invention provides a method of reducing tumor burden, i.e., tumor mass and / or tumor size, in a patient, the method comprising administering to the patient a combination therapy comprising a peptide antagonist of C / EBPβ, such as ST101, and an immunomodulatory agent, and measuring the reduction in tumor burden compared to baseline.

[0078] In certain embodiments, particularly those in which assessment is performed by RECIST 1.1, the disease control rate defines the following levels of tumor response: complete response (CR), which is the disappearance of the tumor(s); partial response (PR), which indicates at least a 30% reduction in the size of the tumor(s); stable disease, which indicates no change in the size of the tumor(s); or progressive disease, which indicates at least a 20% increase in tumor size and / or new lesions.

[0079] Duration of response is the length of time between the achievement of remission and disease progression, i.e., the period during which the tumor does not grow or expand, or until death. Duration of response in a patient receiving a combination therapy of the invention can be, for example, at least 4, 6, 8, 10, or 12 weeks, at least 4, 6, 8, 10, 12, 16, 18, or 24 months, or at least 3, 4, or 5 years. Thus, in one aspect, the invention provides a method for extending the duration of response in a patient, comprising administering to the patient a combination therapy comprising a peptide antagonist of C / EBPβ and an immunomodulatory agent. The extension of duration of response is measured relative to the median duration of response in a control population.

[0080] Survival can be assessed as overall survival, i.e., the length of time a patient survives, or progression-free survival, i.e., the length of time a patient receives treatment without their disease progressing or worsening. Survival can be measured from the date of diagnosis or the date treatment began. Overall survival, median overall survival, progression-free survival, and median progression-free survival can be calculated, for example, by Kaplan-Meier analysis based on response to treatment. Thus, in one aspect, the present invention provides a method for increasing overall survival in a patient, comprising administering to the patient a combination therapy comprising a peptide antagonist of C / EBPβ and an immunomodulatory agent. The increase in overall survival is measured relative to the median overall survival in a control population. In another aspect, the present invention provides a method for increasing progression-free survival in a patient, comprising administering to the patient a combination therapy comprising a peptide antagonist of C / EBPβ and an immunomodulatory agent. The increase in progression-free survival is measured relative to the median progression-free survival in a control population.

[0081] A patient is effectively treated according to the methods of the invention if the patient experiences or exhibits at least one of the following outcomes after administering the combination therapy: - Failure to detect a tumor (or at least one tumor if multiple tumors were present at baseline), - A reduction in tumor size of at least approximately 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% compared to baseline; - No significant (e.g., less than 20%) increase in tumor size compared to baseline; - optionally, a significantly increased duration of response compared to the median duration of response in a control patient population; - optionally, a significantly increased progression-free survival compared to the median progression-free survival of a control patient population; -Optionally, a significantly improved median overall survival compared to the median overall survival of a control patient population. [Example]

[0082] Embodiments of the present disclosure can be further defined with reference to the following non-limiting examples. It will be apparent to those skilled in the art that many modifications, both to materials and methods, can be practiced without departing from the scope of the present disclosure.

[0083] Example 1. Antagonists of C / EBPβ reprogram myeloid-derived suppressor cell polarization and reduce tumor-associated Tregs Myeloid-derived suppressor cells (MDSCs) play a role in cancer progression and other related diseases by suppressing both innate and adaptive immune responses. For example, depletion of MDSCs by conditional depletion of C / EBPβ in hematopoietic cells inhibits the immunosuppressive activity of MDSCs and significantly enhances antitumor immunity.

[0084] Primary human macrophages were cultured with peripheral blood mononuclear cells (hPBMCs) and activated toward the M1 or M2 phenotype with lipopolysaccharide (LPS) and TNFα (M1) or IL-4 (M2), respectively, in the presence of the C / EBPβ antagonist ST101 (Figure 1). Macrophage M1 (CD80, CD86) and M2 (CD163, CD206) expression was analyzed by flow cytometry and rtPCR (Figure 1). Paired biopsy tissues from a phase 1-2 clinical trial of ST101 in patients with advanced, unresectable, and metastatic solid tumors were collected at screening (before ST101 exposure) and within 24 hours of ST101 administration during treatment cycle 2. Nanostring gene expression analysis was performed to identify differential gene expression and the impact of ST101 on the tumor microenvironment.

[0085] Treatment with pharmacologically relevant concentrations of ST101 (2.5, 5, or 10 μM) resulted in a dose-dependent decrease in M2+ macrophages and a corresponding induction of M1+ macrophages (Figures 2A and 2B, 3A and 3C, and 4A-4B). At the highest ST101 concentration, a 12-fold decrease in the M2 to M1 ratio was observed without substantially affecting cell viability (Figures 3B and 3D).

[0086] Paired patient biopsies from the ST101 Phase 1-2 clinical trial demonstrate reduced C / EBPβ target gene IL-6 signaling, a key driver of the M2 macrophage phenotype. Reduced IL-6 signaling is associated with increased tumor-infiltrating macrophage-to-tumor-infiltrating lymphocyte (TIL) ratios and decreased regulatory T cell (Treg)-to-TIL ratios in treated patient tumor samples (Figures 5A-5C).

[0087] Overall, these results confirm the potential of ST101 to reprogram M2 macrophages into pro-inflammatory M1 macrophages, support a novel macrophage-driven mechanism of action for ST101 as an anticancer agent, and support further exploration of ST101 in immuno-oncological therapeutic strategies. Importantly, the effects of ST101 on macrophage polarization may act in concert with direct cytotoxicity in C / EBPβ-driven cancers, raising the possibility that the ST101 target population may expand beyond C / EBPβ-driven cancer types.

[0088] Example 2. C / EBPβ antagonist potently activated CD8+ T cells Cultured human T cells (matched donor) were co-incubated with M1 or M2 macrophage cultures. T cell activation was measured by intracellular interferon-gamma staining in CD8+ cells. T cell activation was suppressed in the presence of M2 macrophages, as indicated by a decrease in the percentage of interferon-gamma-positive cells. Addition of ST101 to cultures of matched donor human T cells containing M2 macrophages restored T cell activity. Treatment of cultures of matched donor human T cells containing M1 macrophages with ST101 enhanced activity compared with untreated cultures of matched donor human T cells containing M1 macrophages. The results are shown in Figure 6.

[0089] Example 3. C / EBPβ antagonists and checkpoint inhibitors exhibit antitumor activity Using a mouse model of triple-negative breast cancer (TNBC), we demonstrate that inhibition of C / EBPβ and PD-1 enhances antitumor activity compared with either treatment alone. Briefly, 4T1-luc TNBC tumor cells were orthotopically implanted into the mammary fat pads of immune-competent Balb / c mice. When treatment was initiated, tumors grew to approximately 100 mm 10 days after implantation. 3Animals were administered 10 mg / kg of ST101 subcutaneously on days 10, 12, 14, 16, 18, and 22 after tumor inoculation and / or 12.5 mg / kg of anti-PD-1 antibody (BioXCell, Lebanon, NH) intraperitoneally on days 10, 14, and 22 after tumor inoculation. The results are shown in Figure 7.

[0090] References Angelastro JM, et al. Selective destruction of glioblastoma cells by interference with the activity or expression of ATF5. Oncogene 2006;25: 907-916. Bernal F, et al. Reactivation of the p53 Tumor Suppressor Pathway by a Stapled p53 Peptide. J. Am. Chem. Soc. 2007;129:2456-2457. Bezy O, et al. Delta-interacting protein A, a new inhibitory partner of CCAAT / enhancer-binding protein beta, implicated in adipocyte differentiation. J Biol Chem 2005;280: 11432-11438. Bird GH, et al. Biophysical Determinants for Cellular Uptake of Hydrocarbon-Stapled Peptide Helices. Nat. Chem. Biol. 2017;12:845-852. Cates CC, et al. Regression / eradication of gliomas in mice by a systemically-deliverable ATF5 dominant-negative peptide. Oncotarget 2016;7: 12718-12730. Dougherty PG, et al. Understanding Cell Penetration of Cyclic Peptides. Chem. Rev. 2019;119(17):10241-10287. Eisenhauer EA, et al. New response evaluation criteria in solid tumours: Revised RECIST guideline (version 1.1). Eur J Cancer 2009;45(2):228-247. Ellingson B, et al. Modified Criteria for Radiographic Response Assessment in Glioblastoma Clinical Trials. Neurotherapeutics. 2017;14:307-320. Homma J, et al. Increased expression of CCAAT / enhancer binding protein beta correlates with prognosis in glioma patients. Oncol Rep 2006;15: 595-601. Huggins CJ, et al. C / EBPgamma suppresses senescence and inflammatory gene expression by heterodimerizing with C / EBPbeta. Mol Cell Biol 2013;33: 3242-3258. Karpel-Massler G, et al. A Synthetic Cell-Penetrating Dominant-Negative ATF5 Peptide Exerts Anticancer Activity against a Broad Spectrum of Treatment-Resistant Cancers. Clin Cancer Res 2016;22: 4698-4711. Marigo I, et al. Tumor-induced tolerance and immune suppression depend on the C / EBPβ transcription factor. Immunity. 2010;32:790-802. Ruffell D, et al. CREB-C / EBPβ cascade induces M2 macrophage-specific gene expression and promotes muscle injury repair. PNAS. 2009;106:17475-17480. Scher HI, et al. Trial Design and Objectives for Castration-Resistant Prostate Cancer: Updated Recommendations from the Prostate Cancer Clinical Trials Working Group 3. J Clin Oncology. 2016:34(12):1402-1418. Sheng Z, et al. An activating transcription factor 5-mediated survival pathway as a target for cancer therapy? Oncotarget 2010;1: 457-460. Zhang ZY, et al. Stabilization of ATF5 by TAK1-Nemo-like kinase critically regulates the interleukin-1beta-stimulated C / EBP signaling pathway. Mol Cell Biol 2015;35: 778-788. Zhao Y, et al. p300-dependent acetylation of activating transcription factor 5 enhances C / EBPbeta transactivation of C / EBPalpha during 3T3-L1 differentiation. Mol Cell Biol 2014;34: 315-324. ***

[0091] The present invention is further described by the following claims.

Claims

1. 1. A method for inhibiting the growth of a solid tumor in a subject, the method comprising combination therapy using (i) a pharmaceutical composition comprising an effective amount of a peptide antagonist of CCAAT-enhancer binding protein beta (C / EBPβ), and (ii) a pharmaceutical composition comprising an effective amount of an immunomodulatory agent, wherein the pharmaceutical composition comprising the C / EBPβ antagonist and the pharmaceutical composition comprising the immunomodulatory agent are administered to the subject simultaneously or separately.

2. 1. A method for reducing the volume of a solid tumor in a subject, the method comprising combination therapy using (i) a pharmaceutical composition comprising an effective amount of a peptide antagonist of CCAAT-enhancer binding protein beta (C / EBPβ), and (ii) a pharmaceutical composition comprising an effective amount of an immunomodulatory agent, wherein the pharmaceutical composition comprising the C / EBPβ antagonist and the pharmaceutical composition comprising the immunomodulatory agent are administered to the subject simultaneously or separately.

3. 1. A method for treating a solid tumor in a subject, the method comprising combination therapy using (i) a pharmaceutical composition comprising an effective amount of a peptide antagonist of CCAAT-enhancer binding protein beta (C / EBPβ), and (ii) a pharmaceutical composition comprising an effective amount of an immunomodulatory agent, wherein the pharmaceutical composition comprising the C / EBPβ antagonist and the pharmaceutical composition comprising the immunomodulatory agent are administered to the subject simultaneously or separately.

4. 10. The method of any preceding claim, wherein the solid tumor is a melanoma, carcinoma, or sarcoma.

5. 10. The method of any preceding claim, wherein the subject has been diagnosed with locally advanced or metastatic breast cancer (LA / MBC), melanoma, glioblastoma (GBM), or castration-resistant prostate cancer (CRPC).

6. The method of any one of claims 1 to 3, wherein the peptide antagonist comprises the D-amino acid sequence VAEAREELERLEARLGQARGEL (SEQ ID NO: 1).

7. The method of any one of claims 1 to 3, wherein the peptide antagonist comprises the amino acid sequence LEGRAQGLRAELRELEERAEAV (SEQ ID NO: 3).

8. 10. The method of any preceding claim, wherein the peptide antagonist is a cell-penetrating peptide.

9. The method of any one of claims 1 to 3, wherein the peptide antagonist is ST101.

10. 10. The method of claim 9, wherein the peptide antagonist is administered to the subject at a dose of about 0.5 to 16 mg / kg.

11. 10. The method of claim 9, wherein the peptide antagonist is administered to the subject at a dose of about 500 mg.

12. The method of any one of claims 1 to 3, wherein the immunomodulatory agent is selected from the group consisting of a checkpoint inhibitor, a cytokine, and an immunoadjuvant.

13. The method of any one of claims 1 to 3, wherein the immunomodulatory agent is an antibody or an antibody-drug conjugate.

14. 14. The method of claim 13, wherein the antibody or antibody-drug conjugate is selected from the group consisting of amivantamab, belantamab mafodotin-blmf, bevacizumab, cetuximab, denosumab, dinutuximab, enfortumab vedotin-ejfv, margetuximab, naxitamab-gqgk, necitumumab, panitumumab, pertuzumab, ramucirumab, sacituzumab govitecan-hziy, tebentafsp-tebn, tisotumab vedotin, trastuzumab, trastuzumab deruxtecan, and trastuzumab emtansine.

15. The method of claim 13, wherein the antibody is an anti-PD-1 antibody.

16. 13. The method of claim 12, wherein the checkpoint inhibitor targets at least one of PD-1, PD-L1, CTLA-4, or LAG-3.

17. 13. The method of claim 12, wherein the checkpoint inhibitor is selected from the group consisting of atezolizumab, avelumab, cemiplimab, dostarlimab, durvalumab, ipilimumab, nivolumab, pembrolizumab, leratimab, and combinations thereof.

18. 13. The method of claim 12, wherein the cytokine targets at least one of the IL-2 pathway, the IL-2R pathway, the IFNAR1 pathway, or the IFNAR2 pathway.

19. 13. The method of claim 12, wherein the cytokine is selected from the group consisting of aldesleukin, granulocyte-macrophage colony-stimulating factor, interferon alpha-2a, interferon alpha-2b, and pegylated interferon alpha-2b.

20. 13. The method of claim 12, wherein the immune adjuvant targets the Toll-like receptor 7 pathway or the Toll-like receptor 3 pathway.

21. 13. The method of claim 12, wherein the immune adjuvant is selected from the group consisting of imiquimod and poly-ICLC.

22. 10. The method of any of the preceding claims, wherein the pharmaceutical composition comprising the C / EBPβ antagonist and / or the pharmaceutical composition comprising the immunomodulatory agent is administered intravenously.

23. 10. The method of any preceding claim, wherein the pharmaceutical composition comprising the C / EBPβ antagonist and / or the pharmaceutical composition comprising the immunomodulatory agent are administered to the subject on different days.

24. The method of any one of claims 1 to 23, wherein the pharmaceutical composition comprising the C / EBPβ antagonist is administered once a week for at least three weeks.

25. The method of any one of claims 1 to 23, wherein the pharmaceutical composition comprising the C / EBPβ antagonist is administered once every two weeks for at least four weeks.

26. 26. The method of any one of claims 1 to 25, wherein the pharmaceutical composition comprising the immunomodulatory agent is administered once a week for at least three weeks.

27. 26. The method of any one of claims 1 to 25, wherein the pharmaceutical composition comprising the immunomodulatory agent is administered once every two weeks for at least four weeks.

28. 26. The method of any one of claims 1 to 25, wherein the pharmaceutical composition comprising the immunomodulatory agent is administered once every three weeks for at least six weeks.

29. A pharmaceutical composition comprising an effective amount of a peptide antagonist of CCAAT enhancer-binding protein beta (C / EBPβ) and an effective amount of an immunomodulatory agent for use in a method of combination therapy to inhibit the growth of a solid tumor in a subject.

30. A pharmaceutical composition comprising an effective amount of a peptide antagonist of CCAAT enhancer-binding protein beta (C / EBPβ) and an effective amount of an immunomodulatory agent for use in a method of combination therapy to reduce the volume of a solid tumor in a subject.

31. A pharmaceutical composition comprising an effective amount of a peptide antagonist of CCAAT enhancer-binding protein beta (C / EBPβ) and an effective amount of an immunomodulatory agent for use in combination therapy to treat solid tumors in a human subject.

32. The pharmaceutical composition of any one of claims 29 to 31, wherein the C / EBPβ antagonist is ST101 and the immunomodulator is a PD-1 inhibitor.

Images