Peptide vaccines and pembrolizumab for treating breast cancer

Abstract

To provide peptide vaccines and pembrolizumab for treating breast cancer.SOLUTION: The disclosure features, inter alia, combination therapies comprising pembrolizumab and one or more immunogenic XBP1-, CD138-, and CS1-derived peptides. The therapies herein can be used, e.g., for inducing an immune response in a subject having a cancer, and treating a cancer such as breast cancer, e.g., triple negative breast cancer. In embodiments, the immunogenic peptides bind to MHC class 1 molecules such as HLA-A molecules. Peptides from X-Box Protein 1 (XBP1)-, CD 138-, and CD2 Subset 1 (CS1) are immunogenic and are useful, e.g., to induce an immune response against various cancer cells.SELECTED DRAWING: None

Description

[Technical Field]

[0001] This application claims priority to U.S. Patent Application No. 62 / 576,404, filed October 24, 2017, the entire contents of which are incorporated herein by reference. [Background technology]

[0002] Several types of vaccines have been developed for the prevention of infectious diseases, including attenuated microorganisms, recombinant protein, and DNA vaccines. Recently, research has been conducted into the development of vaccine immunotherapy for treating cancer patients. Summary of the Invention [Means for solving the problem]

[0003] The present disclosure relates to a combination therapy, for example, one or more immunogenic peptides and pembrolizumab. In an embodiment, the immunogenic peptide binds to an MHC class 1 molecule, for example, an HLA-A molecule. Peptides derived from X-box protein 1 (XBP1), CD138, and CD2 subset 1 (CS1) are immunogenic and are useful, for example, for inducing immune responses against various cancer cells.

[0004] It will be apparent from the description below that the combination therapies herein can be used in a variety of applications, e.g., methods for inducing an immune response in a patient, methods for activating T cells (including, e.g., effector memory T cells and / or central memory T cells), and methods for treating cancer, e.g., breast cancer, e.g., triple-negative breast cancer.

[0005] In some aspects, the present disclosure provides a method of treating cancer, e.g., breast cancer, e.g., triple-negative breast cancer, comprising: (i) pembrolizumab; and (ii) one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. to a subject, wherein the subject has or is at risk of developing cancer, e.g., breast cancer, e.g., triple-negative breast cancer.

[0006] In some aspects, the present disclosure provides a method for treating cancer, e.g., breast cancer, e.g., triple-negative breast cancer, comprising: one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. and pembrolizumab in combination with

[0007] In some aspects, the present disclosure provides a method for treating cancer, e.g., breast cancer, e.g., triple-negative breast cancer, comprising: (a) an unspliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 1, e.g., an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a spliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 2, e.g., a spliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 3, e.g., a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3 and pembrolizumab in combination with

[0008] In some aspects, the present disclosure provides a method for treating cancer, e.g., breast cancer, e.g., triple-negative breast cancer, comprising: (a) an unspliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 1, e.g., an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a spliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 2, e.g., a spliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2; (c) a CD138 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 3, e.g., a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3; and (d) a CS-1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 4, e.g., a CS-1 peptide consisting of the amino acid sequence of SEQ ID NO: 4 and pembrolizumab in combination with

[0009] In some aspects, the present disclosure provides a method for treating cancer, e.g., breast cancer, e.g., triple-negative breast cancer, comprising: (a) an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a splice-form XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3 and pembrolizumab in combination with

[0010] In some aspects, the present disclosure provides a method for treating cancer, e.g., breast cancer, e.g., triple-negative breast cancer, comprising: (a) an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a splice-type XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2; (c) a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3, and (d) CS-1 peptide consisting of the amino acid sequence of SEQ ID NO: 4 and pembrolizumab in combination with

[0011] In some aspects, the present disclosure provides a method for inducing an immune response in a subject, comprising: (i) pembrolizumab; and (ii) one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. The method includes delivering to a subject

[0012] In some aspects, the present disclosure provides a method for inducing an immune response in a subject, e.g., a subject having breast cancer, e.g., triple-negative breast cancer, comprising: one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. and pembrolizumab in combination with

[0013] In some aspects, the present disclosure provides a method for inducing an immune response in a subject with breast cancer, e.g., triple-negative breast cancer, comprising: (a) an unspliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 1, e.g., an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a spliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 2, e.g., a spliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 3, e.g., a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3 and pembrolizumab in combination with

[0014] In some aspects, the present disclosure provides a method for inducing an immune response in a subject with breast cancer, e.g., triple-negative breast cancer, comprising: (a) an unspliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 1, e.g., an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a spliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 2, e.g., a spliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 3, e.g., a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3; and (d) a CS-1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 4, e.g., a CS-1 peptide consisting of the amino acid sequence of SEQ ID NO: 4 and pembrolizumab in combination with

[0015] In some aspects, the present disclosure provides a method for inducing an immune response in a subject with breast cancer, e.g., triple-negative breast cancer, comprising: (a) an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a splice-form XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3 and pembrolizumab in combination with

[0016] In some aspects, the present disclosure provides a method for inducing an immune response in a subject with breast cancer, e.g., triple-negative breast cancer, comprising: (a) an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a splice-type XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2; (c) a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3, and (d) CS-1 peptide consisting of the amino acid sequence of SEQ ID NO: 4 and pembrolizumab in combination with

[0017] In some aspects, the present disclosure provides a method of treating breast cancer, e.g., triple-negative breast cancer, comprising: (i) pembrolizumab; and (ii): (a) an unspliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 1, e.g., an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a spliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 2, e.g., a spliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 3, e.g., a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3 The method includes administering to a subject

[0018] In some aspects, the present disclosure provides a method of treating breast cancer, e.g., triple-negative breast cancer, comprising: (i) pembrolizumab; and (ii): (a) an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a splice-form XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3 The method includes administering to a subject

[0019] Any of the above aspects may also involve one or more of the following embodiments.

[0020] In some embodiments, the unspliced ​​XBP1 peptide is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 1. In some embodiments, the spliced ​​XBP1 peptide is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 2. In some embodiments, the CD138 peptide is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 3. In some embodiments, the CS-1 peptide is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 4.

[0021] In some embodiments, the unspliced ​​XBP1 peptide consists of the amino acid sequence of SEQ ID NO: 1. In some embodiments, the spliced ​​XBP1 peptide consists of the amino acid sequence of SEQ ID NO: 2. In some embodiments, the CD138 peptide consists of the amino acid sequence of SEQ ID NO: 3. In some embodiments, the CS-1 peptide consists of the amino acid sequence of SEQ ID NO: 4.

[0022] In some embodiments, the method comprises administering, or the composition for use comprises: (a) an unspliced ​​XBP1 peptide described herein, (b) a spliced ​​XBP1 peptide described herein, and (c) a CD138 peptide described herein. In some embodiments, the method comprises administering, or the composition for use comprises: (a) an unspliced ​​XBP1 peptide that is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 1, (b) a spliced ​​XBP1 peptide that is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide that is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 3. In some embodiments, the method comprises administering, or the composition for use comprises: (a) an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1, (b) a spliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3. In some embodiments, the method comprises administering, or the composition for use further comprises: (d) a CS-1 peptide described herein. In some embodiments, the method comprises administering, or the composition for use further comprises: (d) a CS-1 peptide that is 35 amino acids or less in length and that comprises the amino acid sequence of SEQ ID NO: 4. In some embodiments, the method comprises administering, or the composition for use further comprises: (d) a CS-1 peptide consisting of the amino acid sequence of SEQ ID NO: 4.

[0023] In some embodiments, the method includes administering one or more immunostimulatory agents, or the composition for use includes one or more immunostimulatory agents. In some embodiments, the immunostimulatory agent is an adjuvant including carboxymethylcellulose, polyinosinic acid-polycytidylic acid, and poly-L-lysine double-stranded RNA (e.g., poly IC-LC, e.g., hiltonol); an adjuvant including a water-and-oil emulsion; and adjuvants including proteins (e.g., cytokines, GCSF, or GM-CSF). In some embodiments, the method includes administering, or the composition for use further includes, an additional treatment, such as one or more chemotherapeutic agents, one or more forms of ionizing radiation, one or more immunotherapeutic agents (e.g., cancer vaccines, immune checkpoint inhibitors), one or more immune checkpoint inhibitors, e.g., antibodies that inhibit immune checkpoint molecules (e.g., anti-CTLA4 antibodies, e.g., ipilimumab or tremelimumab, PD-1 antibodies, or PDL-1 antibodies), or an adjuvant, e.g., a small molecule adjuvant (e.g., thalidomide or a thalidomide derivative, e.g., lenalidomide). In some embodiments, the method includes administering, or the composition for use further includes, lenalidomide. In some embodiments, the method comprises administering PolyIC-LC, or the composition for use further comprises PolyIC-LC.

[0024] In some embodiments, the cancer is breast cancer. In some embodiments, the breast cancer is triple-negative breast cancer. In some embodiments, the subject has, is at risk of developing, or is suspected of having breast cancer, for example, triple-negative breast cancer. In some embodiments, the subject has, or has been identified as having, one or more cancer cells that express XBP1, CD138, or CS1, or any combination thereof. In some embodiments, the method further comprises, after delivering the composition to the subject, determining whether an immune response has occurred in the subject. In some embodiments, the subject is human. In some embodiments, the subject is in remission from breast cancer, for example, triple-negative breast cancer.

[0025] In some embodiments, (i) pembrolizumab and (ii) peptide are administered separately or together. In some embodiments, (i) is administered before, together with, or after (ii). In some embodiments, (i) and (ii) are formulated for use separately or together. In some embodiments, (i) is formulated for administration before, together with, or after (ii).

[0026] In some embodiments, pembrolizumab is administered at a dose of 200 mg. In some embodiments, pembrolizumab is administered once every three weeks.

[0027] In some embodiments, the one or more peptides are administered at a dose of 0.8 mg total peptide, e.g., 0.2 mg unspliced ​​XBP1 peptide, 0.2 mg spliced ​​XBP1 peptide, 0.2 mg CD138 peptide, and 0.2 mg CS-1 peptide. In some embodiments, the one or more peptides are administered at a dose of 0.4-1.2 mg total peptide, e.g., 0.1-0.3 mg unspliced ​​XBP1 peptide, 0.1-0.3 mg spliced ​​XBP1 peptide, 0.1-0.3 mg CD138 peptide, and 0.1-0.3 mg CS-1 peptide. In some embodiments, the one or more peptides are administered at a dose of 0.8-1.2 mg total peptide, e.g., 0.2-0.3 mg unspliced ​​XBP1 peptide, 0.2-0.3 mg spliced ​​XBP1 peptide, 0.2-0.3 mg CD138 peptide, and 0.2-0.3 mg CS-1 peptide. In some embodiments, the one or more peptides are administered at a dose of 0.4-1.6 mg total peptide, e.g., 0.1-0.4 mg unspliced ​​XBP1 peptide, 0.1-0.4 mg spliced ​​XBP1 peptide, 0.1-0.4 mg CD138 peptide, and 0.1-0.4 mg CS-1 peptide. In some embodiments, the one or more peptides are administered at a dose of 0.8-1.6 mg total peptide, e.g., 0.2-0.4 mg unspliced ​​XBP1 peptide, 0.2-0.4 mg spliced ​​XBP1 peptide, 0.2-0.4 mg CD138 peptide, and 0.2-0.4 mg CS-1 peptide.

[0028] In some embodiments, the one or more peptides are administered at a dose of 0.6 mg total peptide, e.g., 0.2 mg unspliced ​​XBP1 peptide, 0.2 mg spliced ​​XBP1 peptide, and 0.2 mg CD138 peptide. In some embodiments, the one or more peptides are administered at a dose of 0.3-0.9 mg total peptide, e.g., 0.1-0.3 mg unspliced ​​XBP1 peptide, 0.1-0.3 mg spliced ​​XBP1 peptide, and 0.1-0.3 mg CD138 peptide. In some embodiments, the one or more peptides are administered at a dose of 0.6-0.9 mg total peptide, e.g., 0.2-0.3 mg unspliced ​​XBP1 peptide, 0.2-0.3 mg spliced ​​XBP1 peptide, and 0.2-0.3 mg CD138 peptide. In some embodiments, the one or more peptides are administered at a dose of 0.3-1.2 mg total peptide, e.g., 0.1-0.4 mg unspliced ​​XBP1 peptide, 0.1-0.4 mg spliced ​​XBP1 peptide, and 0.1-0.4 mg CD138 peptide. In some embodiments, the one or more peptides are administered at a dose of 0.6-1.2 mg total peptide, e.g., 0.2-0.4 mg unspliced ​​XBP1 peptide, 0.2-0.4 mg spliced ​​XBP1 peptide, and 0.2-0.4 mg CD138 peptide.

[0029] In embodiments, the peptides are administered as a first dose and one or more additional doses. In embodiments, the one or more peptides are administered every two weeks, e.g., for at least 1, 2, 3, 4, 5, 6, 8, 10, or 12 weeks. In embodiments, the one or more peptides are administered once weekly. In embodiments, the one or more peptides are administered every 1, 2, 3, or 4 weeks, e.g., for at least 1, 2, 3, 4, 5, 6, 8, 10, or 12 weeks. In embodiments, the one or more peptides are administered two or more times over the course of 12, 18, 24, 30, or 36 months, e.g., over the course of 18 to 24 months. In embodiments, the additional dose is administered about 9 to 12, 12 to 18, 18 to 24, 24 to 30, or 30 to 36 months after the first dose. In embodiments, the one or more peptides are administered to the subject at least 2, 3, 4, 5, 6, 7, 8, 9 or 10 times, for example, at least 4 or 6 times.

[0030] The present disclosure provides, in certain aspects, (i) pembrolizumab; and (ii) one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. Also provided is a composition comprising:

[0031] The present disclosure provides, in certain aspects, (i) pembrolizumab; and (ii) one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. Also provided is a kit comprising:

[0032] In embodiments, (i) and (ii) are mixed, and in embodiments, (i) and (ii) are separate, for example in separate containers.

[0033] In embodiments, the peptides herein comprise one or more of an XBP1 peptide, a CD138 peptide, and a CS-1 peptide that have affinity for multiple MHC molecules, e.g., HLA-A molecules, e.g., HLA-A2, increased stability within the peptide-binding cleft of multiple MHC molecules, e.g., HLA-A2, and the ability to induce activation and proliferation of T cells, including, for example, effector memory T cells and / or central memory T cells, when expressed on the surface of a cell (e.g., a cancer cell) in association with an MHC molecule, e.g., HLA-A2.

[0034] It will be apparent from the description herein that the combination therapy herein can be used in a variety of applications, such as a method for inducing an immune response in a patient with cancer, a method for activating T cells (e.g., effector memory T cells and / or central memory T cells) in a patient with cancer, a method for producing antibodies in a patient with cancer, and a method for treating cancer, such as breast cancer, for example, triple-negative breast cancer. In some embodiments, inducing an immune response comprises inducing a subject to produce antibodies against one or more of these peptides.

[0035] In some embodiments, the combination therapy comprises an isolated peptide comprising an amino acid sequence that is at least 66 (e.g., at least 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99)% identical to any one of SEQ ID NOs: 1-4. The peptide is capable of binding to a major histocompatibility complex (MHC) molecule, e.g., an MHC class I or class II molecule. In one embodiment, the peptide is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or up to 25, 30, or 35 amino acids in length and comprises the amino acid sequence of any one of SEQ ID NOs: 1-4, or an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of any one of SEQ ID NOs: 1-4. In one embodiment, the peptide is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or up to 25, 30, or 35 amino acids in length and comprises the amino acid sequence of any one of SEQ ID NOs: 1-4. In one embodiment, the peptide is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or up to 25, 30, or 35 amino acids in length, and the peptide comprises an amino acid sequence having three, two, or one substitution of the amino acid sequence of any one of SEQ ID NOs: 1-4. The substitutions can be conservative or non-conservative.

[0036] In one embodiment, the peptide consists of an amino acid sequence that is at least 66 (e.g., at least 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98 or 99)% identical to any one of SEQ ID NOs: 1-4. In one embodiment, the peptide consists of any one of the amino acid sequences of SEQ ID NOs: 1-4 with 3, 2 or 1 substitution. In one embodiment, the peptide consists of any one of the amino acid sequences of SEQ ID NOs: 1-4.

[0037] In some embodiments, any of the isolated peptides described herein can be associated with a major histocompatibility complex (MHC) molecule and recognized by an antigen-specific T cell receptor on a T cell.

[0038] In some embodiments, the combination therapy comprises a fusion protein comprising a first amino acid sequence consisting of a peptide described herein, e.g., an unspliced ​​XBP1 peptide described herein, a spliced ​​XBP1 peptide described herein, a CD138 peptide described herein, and / or a CS-1 peptide described herein; and a second amino acid sequence heterologous to the first amino acid sequence.

[0039] In some embodiments, the second amino acid sequence may comprise or be a targeting polypeptide, an immunostimulatory molecule, an immunoglobulin or an antigen-binding fragment thereof, an Fc receptor binding region of an immunoglobulin molecule, or a carrier polypeptide. The targeting polypeptide may be, for example, a polypeptide that directs the isolated peptide to an antigen-presenting cell (e.g., a dendritic cell, a macrophage, a monocyte, or a B cell). The immunostimulatory molecule may be, for example, a cytokine or a T-helper epitope. The immunoglobulin may be, for example, a single-chain Fv immunoglobulin fragment or an entire immunoglobulin molecule. The carrier polypeptide may comprise or be a KLH (keyhole limpet hemocyanin) polypeptide or an albumin polypeptide.

[0040] In some embodiments, any of the isolated peptides described herein may include a linker sequence. The linker sequence may directly or indirectly connect a first amino acid sequence to a second amino acid sequence. The linker sequence may include or consist of one or more amino acids, for example, at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids. In one embodiment, the linker may include or consist of at least one (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more) protease cleavage site.

[0041] In some embodiments, the second amino acid sequence can be amino-terminal or carboxy-terminal to the first amino acid sequence.

[0042] In some embodiments, any of the isolated peptides or fusion proteins described herein may be detectably labeled. The detectable label may be selected from the group consisting of a luminescent label, a fluorescent label, a radioactive label, and an enzymatic label.

[0043] In some embodiments, the combination therapy includes at least two peptides, e.g., two, three, four or more peptides described herein (e.g., a pharmaceutical composition herein includes, or a method of treatment herein includes, administering):

[0044] In one embodiment, the combination therapy comprises at least two peptides.For example, the combination therapy comprises unspliced ​​XBP1 peptide and spliced ​​XBP1 peptide; the combination therapy comprises unspliced ​​XBP1 peptide and CD138 peptide; the combination therapy comprises unspliced ​​XBP1 peptide and CS-1 peptide; the combination therapy comprises spliced ​​XBP1 peptide and CD138 peptide; the combination therapy comprises spliced ​​XBP1 peptide and CS-1 peptide; the combination therapy comprises CD138 peptide and CS-1 peptide.

[0045] In one embodiment, the combination therapy comprises at least three peptides.For example, the combination therapy comprises unspliced ​​XBP1 peptide, spliced ​​XBP1 peptide and CD138 peptide; the combination therapy comprises unspliced ​​XBP1 peptide, spliced ​​XBP1 peptide and CS-1 peptide; the combination therapy comprises unspliced ​​XBP1 peptide, CD138 peptide and CS-1 peptide; the combination therapy comprises spliced ​​XBP1 peptide, CD138 peptide and CS-1 peptide.In one embodiment, the combination therapy comprises at least three peptides, for example, unspliced ​​XBP1 peptide, spliced ​​XBP1 peptide and CD138 peptide.

[0046] In one embodiment, the combination therapy comprises four peptides, for example, the combination therapy comprises an unspliced ​​XBP1 peptide, a spliced ​​XBP1 peptide, a CD138 peptide, and a CS-1 peptide.

[0047] In one embodiment, the combination therapy comprises an unspliced ​​XBP1 peptide that is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or up to 25, 30, or 35 amino acids in length and comprises the amino acid sequence of SEQ ID NO: 1. In one embodiment, the combination therapy comprises a spliced ​​XBP1 peptide that is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or up to 25, 30, or 35 amino acids in length and comprises the amino acid sequence of SEQ ID NO: 2. In one embodiment, the combination therapy comprises a CD138 peptide that is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or up to 25, 30, or 35 amino acids in length and comprises the amino acid sequence of SEQ ID NO: 3. In one embodiment, the combination therapy comprises a CS-1 peptide that is 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, up to 25, 30, or 35 amino acids in length and comprises the amino acid sequence of SEQ ID NO:4.

[0048] In one embodiment, the combination therapy comprises four peptides, the four peptides being: a peptide comprising (e.g., consisting of) the amino acid sequence of SEQ ID NO: 1, a peptide comprising (e.g., consisting of) the amino acid sequence of SEQ ID NO: 2, a peptide comprising (e.g., consisting of) the amino acid sequence of SEQ ID NO: 3, and a peptide comprising (e.g., consisting of) the amino acid sequence of SEQ ID NO: 4.

[0049] The combination therapy may also include, for example, one or more additional agents, such as one or more therapeutic, diagnostic, or prophylactic agents, or immunostimulatory or immunomodulatory agents.Immunostimulatory agents include, but are not limited to, T-helper epitopes, modified peptide ligands, adjuvants, or any other immunostimulatory agents described herein.The T-helper epitope may be, for example, the PADRE sequence or the universal tetanus toxoid T-helper (TT Th) epitope. The adjuvant may be selected from the group consisting of Freund's complete adjuvant, Freund's incomplete adjuvant, alum, ligands for Toll receptors, saponins (e.g., QS21), RIBI, cholera toxin (CT), E. coli heat-labile toxin (LT), mutant CT (MCT), mutant E. coli heat-labile toxin (MLT), carboxymethylcellulose, adjuvants containing polyinosinic acid-polycytidylic acid and poly-L-lysine double-stranded RNA (e.g., poly IC-LC, e.g., hiltonol), adjuvants containing water-oil emulsions (e.g., montanide), and adjuvants containing proteins (e.g., cytokines, complement, GCSF, or GM-CSF). In one embodiment, the immunostimulatory agent is an adjuvant containing carboxymethylcellulose, polyinosinic acid-polycytidylic acid, and poly-L-lysine double-stranded RNA (e.g., poly IC-LC, e.g., hiltonol). In one embodiment, the adjuvant is a water-in-oil emulsion, e.g., montanide. In one embodiment, the adjuvant is a protein, e.g., a cytokine, complement, GCSF, or GM-CSF. In one embodiment, the additional agent can be a protein, e.g., an antibody. In one embodiment, the additional agent is an immune checkpoint inhibitor. For example, an antibody that inhibits an immune checkpoint molecule can be an anti-CTLA4 antibody, e.g., ipilimumab or tremelimumab, or an anti-PD-1 antibody, or an anti-PDL-1 antibody. In one embodiment, the additional agent can be a small molecule adjuvant, e.g., thalidomide or a thalidomide derivative, e.g., lenalidomide.

[0050] The combination therapy may also include immunogenic peptides other than those disclosed above, such as immunogenic peptides derived from WT1 or derivatives thereof. Exemplary WT1 peptides are described in U.S. Patent No. 7,598,221, the contents of which are incorporated herein by reference. In one embodiment, the combination therapy includes one or more immunogenic peptides derived from WT1 or derivatives thereof, for example, selected from one or more of the following: a WT1 class 1 epitope; a peptide comprising (or consisting of) RMFPNAPYL (WT1 126-134); a peptide comprising (or consisting of) YMFPNAPYL; a peptide comprising (or consisting of) RSDELVRHHNMHQRNMTKL (WT1 427-445); a peptide comprising (or consisting of) PGCNKRYFKLSHLQMHSRKHTG (WT1 331-352); a peptide comprising (or consisting of) SGQARMFPNAPYLPSCLES (WT1 122-140); and a peptide comprising (or consisting of) SGQAYMFPNAPYLPSCLES. Other immunogenic peptides include, but are not limited to, immunogenic peptides derived from MUC1, immunogenic peptides derived from gp100, immunogenic peptides derived from TRP-2, immunogenic peptides derived from MAG1, immunogenic peptides derived from NY-ESO1, immunogenic peptides derived from HER-2; and immunogenic peptides derived from AIM2.

[0051] In one embodiment, the compositions described herein are used to treat a subject having or at risk of having cancer, e.g., a breast cancer described herein, e.g., triple-negative breast cancer.

[0052] In some embodiments, the kits described herein include instructions for administering the peptide to a subject with cancer, e.g., breast cancer, e.g., triple-negative breast cancer.

[0053] In some embodiments, the kit may also include, for example, one or more pharmaceutically acceptable carriers, one or more immunostimulatory or immunomodulatory agents, or one or more therapeutic, diagnostic, or prophylactic agents. In one embodiment, the immunostimulatory agent is an immunostimulatory agent described herein. The one or more immunostimulatory agents may be selected from the group consisting of T-helper epitopes, modified peptide ligands, and adjuvants. In one embodiment, the immunostimulatory agent is an adjuvant comprising carboxymethylcellulose, polyinosinic acid-polycytidylic acid, and poly-L-lysine double-stranded RNA (e.g., poly IC-LC, e.g., hiltonol); an adjuvant comprising a water-oil emulsion (e.g., montanide); or an adjuvant comprising a protein (e.g., a cytokine, complement, GCSF, or GM-CSF). In one embodiment, the additional agent may be a protein, e.g., an antibody. In one embodiment, the additional agent is an immune checkpoint inhibitor. For example, the antibody that inhibits an immune checkpoint molecule can be an anti-CTLA4 antibody, e.g., ipilimumab or tremelimumab, or an anti-PD-1 antibody, or an anti-PDL-1 antibody. In one embodiment, the additional agent can be a small molecule adjuvant, e.g., thalidomide or a thalidomide derivative, e.g., lenalidomide. In one embodiment, the kit further comprises instructions for administering an immunostimulatory and / or immunomodulatory agent in combination with the peptide(s) described herein or the composition described herein.

[0054] In one embodiment, the kit further comprises an additional immunogenic peptide, for example, a WT1-derived immunogenic peptide or a derivative thereof, for example, an immunogenic WT1 peptide described herein. Other immunogenic peptides include, but are not limited to, MUC1-derived immunogenic peptides, gp100-derived immunogenic peptides, TRP-2-derived immunogenic peptides, MAG1-derived immunogenic peptides, NY-ESO1-derived immunogenic peptides, HER-2-derived immunogenic peptides, and AIM2-derived immunogenic peptides. In one embodiment, the kit further comprises instructions for administering the additional immunogenic peptide, for example, a WT1 peptide, in combination with the peptide(s) or composition described herein.

[0055] The methods described herein for inducing an immune response in a subject can include delivering, e.g., administering, to the subject one or more of any of the isolated peptides described herein and / or the compositions described herein. In one embodiment, the subject is administered at least two, e.g., two, three, or four, of the peptides set forth in SEQ ID NOs: 1-4. For example, the subject can be administered two or more of an unspliced ​​XBP1 peptide, a spliced ​​XBP1 peptide, a CD138 peptide, a CS-1 peptide, and combinations thereof.

[0056] This method can also include a step of determining whether an immune response has occurred in the subject after delivering one or more peptides or compositions to the subject.One or more peptides can be delivered to the subject as a pharmaceutical composition, for example, a pharmaceutical composition described herein.The subject can be, for example, a mammal (for example, a human) or any other subject described herein.The subject can have, be suspected of having, be at risk of developing, or be in remission from cancer, for example, breast cancer, for example, triple-negative breast cancer.

[0057] In some embodiments, the method may include determining whether the cancer cell (or cells) express one or more of XBP1, CD138, or CS-1.

[0058] In some embodiments, the method may further include administering to the subject one or more additional treatments, such as a chemotherapeutic agent, ionizing radiation, surgery, or one or more additional immunotherapeutic agents. The one or more forms of ionizing radiation may be, for example, gamma-irradiation, X-ray-irradiation, or beta-irradiation. The one or more chemotherapeutic agents may be a chemotherapeutic agent described herein, for example, a chemotherapeutic agent selected from the group consisting of platinum-based agents, taxanes, topoisomerase inhibitors, antimetabolites, alkylating agents, protease inhibitors, HDAC inhibitors, and vinca alkaloids. Exemplary chemotherapeutic agents include, but are not limited to, cisplatin, carboplatin, procarbazine, mechlorethamine, cyclophosphamide, camptothecin, adriamycin, ifosfamide, melphalan, chlorambucil, bisulfan, nitrosurea, dactinomycin, daunorubicin, doxorubicin, bleomycin, plicamycin, mitomycin, etoposide, verampil, podophyllotoxin, taxol, transplatinum, 5-fluorouracil, vincristine, vinblastin, methotrexate, and analogs of any of the foregoing. The method may also include administering to the subject one or more immunostimulatory agents, such as one or more immunostimulatory agents described herein.

[0059] In one embodiment, the method further comprises administering an additional immunogenic peptide, e.g., a WT1-derived immunogenic peptide or a derivative thereof, e.g., an immunogenic WT1 peptide described herein, in combination with one or more peptides described herein. Other immunogenic peptides include, but are not limited to, MUC1-derived immunogenic peptides, gp100-derived immunogenic peptides, TRP-2-derived immunogenic peptides, MAG1-derived immunogenic peptides, NY-ESO1-derived immunogenic peptides, HER-2-derived immunogenic peptides, and AIM2-derived immunogenic peptides.

[0060] In some embodiments, the delivering step comprises administering to the subject one or more peptides described herein (e.g., one or more peptides comprising any of SEQ ID NOS: 1-4). In some embodiments, the delivering step comprises administering to the subject one or more nucleic acids, each of which comprises a nucleotide sequence encoding one or more peptides, the nucleotide sequence being operably linked to an expression control sequence. The nucleic acid may be in a recombinant cell that has been transfected with the nucleic acid and expresses the one or more peptides. The recombinant cell may be a transfected cell, or a progeny of a transfected cell, produced by transfecting a cell obtained from the subject. The recombinant cell may be, for example, an antigen-presenting cell, such as, but not limited to, a dendritic cell, a macrophage, a monocyte, or a B cell.

[0061] In some embodiments of any of the above methods, the delivering step includes contacting one or more peptides with cells; and delivering the cells to a subject after contacting one or more peptides with the cells. The cells can be, for example, antigen-presenting cells, such as any of the antigen-presenting cells described herein. The cells can be, for example, cells obtained from a subject, or progeny of cells. In some embodiments, the cells can be cells obtained from another subject of the same species as the subject, or progeny of cells. The other subject can express at least one MHC molecule in common with the subject. The at least one MHC molecule can be, for example, an MHC class I molecule, such as an HLA-A2 molecule.

[0062] In one embodiment, the method further comprises administering an additional agent to the subject, e.g., a chemotherapeutic agent and / or an immunostimulatory and / or immunomodulatory agent. In one embodiment, the additional agent is an immunostimulatory agent, e.g., an immunostimulatory agent described herein. In one embodiment, the immunostimulatory agent is an adjuvant comprising carboxymethylcellulose, polyinosinic acid-polycytidylic acid, and poly-L-lysine double-stranded RNA (e.g., poly IC-LC, e.g., hiltonol); an adjuvant comprising a water-oil emulsion (e.g., montanide); or an adjuvant comprising a protein (e.g., a cytokine, complement, GCSF, or GM-CSF). In one embodiment, the additional agent can be a protein, e.g., an antibody. In one embodiment, the additional agent is an immune checkpoint inhibitor. For example, the antibody that inhibits an immune checkpoint molecule can be an anti-CTLA4 antibody, e.g., ipilimumab or tremelimumab, or an anti-PD-1 antibody, or an anti-PDL-1 antibody. In one embodiment, the additional agent may be a small molecule adjuvant, such as thalidomide or a thalidomide derivative, such as lenalidomide. In one embodiment, the method includes administering an additional immunogenic peptide, such as a WT1-derived immunogenic peptide or its derivative, such as a WT1 peptide described herein, in combination with one or more of these peptides. Other immunogenic peptides include, but are not limited to, MUC1-derived immunogenic peptides, gp100-derived immunogenic peptides, TRP-2-derived immunogenic peptides, MAG1-derived immunogenic peptides, NY-ESO1-derived immunogenic peptides, HER-2-derived immunogenic peptides, and AIM2-derived immunogenic peptides.

[0063] In one embodiment, the method further comprises administering one or more additional doses of the peptide or composition described herein.In one embodiment, the subject is administered one or more additional doses about 14 days after the previous dose, for example, the subject is administered 2, 3, 4, 5, 6, 7, 8, 9 or 10 doses of the peptide or composition described herein every other week.In one embodiment, the subject is administered one or more additional doses about 7 days after the previous dose, for example, the subject is administered 2, 3, 4, 5, 6, 7, 8, 9 or 10 doses of the peptide or composition described herein every week.

[0064] In some embodiments, the method herein comprises selecting a treatment for a mammal in need thereof. This method may comprise determining whether one or more cancer cells in the mammal express XBP1; and if one or more cancer cells express XBP1, selecting one or more of the peptides described herein, a fusion protein comprising such peptides, or a composition described herein as a therapeutic agent for the mammal. This method may also comprise, after determining that one or more cancer cells express XBP1, delivering one or more of the peptides described herein, a fusion protein comprising such peptides, or a composition described herein to the subject.

[0065] In some embodiments of any of the above methods, the subject or mammal may have undergone treatment for cancer, e.g., breast cancer, e.g., triple-negative breast cancer, and been non-responsive to that treatment, e.g., a combination therapy described herein may be a second-line, third-line, or fourth-line treatment.

[0066] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this invention belongs. In case of conflict, the present specification, including definitions, will control. Various suitable methods and materials are described below, but methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention. All publications, patent applications, patents and other references mentioned herein are incorporated herein by reference in their entirety. The materials, methods and examples disclosed herein are illustrative only and are not intended to be limiting.

[0067] Other features of the invention, such as methods for inducing an immune response in a subject, and Further advantages will become apparent from the following description and claims. DETAILED DESCRIPTION OF THE INVENTION

[0068] Detailed Description peptide The combination therapy herein comprises one or more immunogenic XBP1-derived peptides, CD138-derived peptides, and CS-1-derived peptides (and pharmaceutical compositions thereof). The treatment herein can be used, for example, to induce an immune response (e.g., stimulate a CTL response) or stimulate antibody production in a subject with cancer.

[0069] A detailed description of the peptides and exemplary methods for making and using the peptides is provided below.

[0070] The present disclosure features combination therapies comprising one or more isolated peptides comprising an amino acid sequence having sufficient identity to or identical to any one of SEQ ID NOs: 1-4 shown in Table 1. [Table 1]

[0071] In some embodiments, the isolated peptide is at least 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, or 35 amino acids in length (e.g., between 9 and 35 amino acids in length, e.g., 9-30, 9-25, 9-20, 9-15 amino acids in length) and comprises an amino acid sequence that has at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% identity to or is identical to the amino acid sequence of SEQ ID NOs: 1-4. Other suitable peptides can be at least 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, or 35 amino acids in length (e.g., between 9 and 35 amino acids in length, e.g., 9-30, 9-25, 9-20, 9-15 amino acids in length) and include the amino acid sequences of SEQ ID NOs: 1-4, or amino acid sequences having one, two, three, or four substitutions of the amino acid sequences of SEQ ID NOs: 1-4. The substitutions can be conservative or non-conservative substitutions.

[0072] "Unspliced ​​XBP1" peptides include the peptide of SEQ ID NO: 1, which has 261 amino acids and the following sequence: [ka] The term "XBP1" refers to peptides having an amino acid sequence of at least 5 (e.g., 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, 31, 32, 33, 34, or 35) contiguous amino acids derived from the unspliced ​​form of the human XBP1 protein having the sequence of SEQ ID NO: 5, and peptides having no more than 1, 2, 3, 4, 5 substitutions (e.g., conservative substitutions) of amino acids from the amino acid sequence of SEQ ID NO: 5. The amino acid positions of unspliced ​​XBP1 referred to in Table 1 are based on SEQ ID NO: 5.

[0073] The "splice-form XBP1" peptide includes the peptide of SEQ ID NO:2, which has 376 amino acids and the following sequence: [ka] The term "XBP1 splice form" refers to a peptide having an amino acid sequence of at least 5 (e.g., 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, 31, 32, 33, 34, or 35) consecutive amino acids from a spliced ​​form of the human XBP1 protein (XBP1 splice form) having the following structure:

[0074] The "CD138" peptide includes the peptide of SEQ ID NO: 3, which has 310 amino acids and the following sequence: [ka] The term "CD138" refers to a peptide having an amino acid sequence of at least 5 (e.g., 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, 31, 32, 33, 34, or 35) consecutive amino acids derived from the human CD138 protein having the following structure:

[0075] The "CS-1" peptide comprises the peptide of SEQ ID NO: 4, which has 335 amino acids and the following sequence: [ka] 8. The amino acid positions of CS-1 referred to in Table 1 are based on SEQ ID NO:8.

[0076] Peptides in general Since related sequences exist in the wild-type, full-length, mature human proteins having SEQ ID NOS: 5-8, the peptides described herein are identified by the residue numbers of the N- and C-terminal amino acids of the peptides (e.g., XBP1 118-126). These peptides frequently have sequences identical to the corresponding segments of the wild-type full-length mature protein having SEQ ID NOS: 5-8. However, the terms "unspliced ​​XBP1 peptide" (e.g., unspliced ​​XBP1 peptides having the following amino acid positions: 118-136, 185-193, 186-194, 190-198, 193-200, or 111-119), "spliced ​​XBP1 peptide" (e.g., spliced ​​XBP1 peptides having the following amino acid positions: 197-205, 194-202, 224-232, 368-376), "CD138 peptide" (e.g., CD138 ... It is understood that "CD138 peptides having amino acid positions: 256-264, 265-273, 260-268, 5-13, or 7-15," and "CS1 peptides" (e.g., CS-1 peptides having amino acid positions 236-245, 240-248, 239-247, 232-240, or 9-17) can be peptide fragments of XBP1 unspliced ​​peptides, XBP1 spliced ​​peptides, CD138, or CS-1 polypeptides (respectively) of species other than humans. As will be understood by those skilled in the art, the numbering of amino acids at the N- and C-termini of such peptide fragments of non-human polypeptides need not be the same as the numbering in the corresponding peptide fragments of human polypeptides. Furthermore, the length and / or amino acid sequence of peptide fragments of non-human polypeptides need not be the same as the length and / or amino acid sequence of the corresponding peptide fragments of human polypeptides. Those skilled in the art will know how to establish the N- and C-terminal amino acids, length, and amino acid sequence of peptides derived from non-human unspliced ​​XBP1, spliced ​​XBP1, CD138, and CS-1 polypeptides. One useful method for doing this is sequence alignment, particularly maximum homology sequence alignment.

[0077] The percent identity between two peptide sequences (e.g., peptides of SEQ ID NOS: 1-4) and another amino acid sequence that may be at least 66% identical to the peptide can be calculated using a number of algorithms, including Clustal W (The European Bioinformatics Institute (EMBL-EBI)), BLAST-Protein (National Center for Biotechnology Information (NCBI), United States National Institutes of Health), and PSAlign (University of Texas A&M; Sze et al. (2006) Journal of Computational Biology 13:309-319). The amount of the ATP that can be determined may be determined using a variety of algorithms and computer programs, including but not limited to:

[0078] Some of the peptides described herein are heteroclitic. As used herein, "heteroclitic" (e.g., heteroclitic peptide) refers to a form of peptide in which one or more amino acids are modified from the wild-type or original sequence to produce a peptide that is more immunogenic than the corresponding wild-type peptide. For example, in the exemplary heteroclitic peptides of SEQ ID NO: 1 and SEQ ID NO: 2, the bolded amino acids indicate the amino acids that are modified from the wild-type sequence of XBP1.

[0079] The variants of the above-mentioned human and non-human peptides are also disclosed herein.The variants of the human and non-human peptides described herein can include: (i) four or less (for example, three, two or one) amino acid substitutions (for example, conservative or non-conservative substitutions); (ii) terminal or internal deletions; or (iii) terminal or internal additions, and all of these are described in detail below.

[0080] The present disclosure also features combination therapies that include peptides comprising, consisting of, or consisting essentially of the amino acid sequence of any of SEQ ID NOs: 1-4, but with no more than four (e.g., no more than three, no more than two, or no more than one) substitutions. The substitutions can be, for example, conservative or non-conservative (as described above).

[0081] Conservative substitutions include substitutions within the following groups: valine, alanine, and glycine; leucine, valine, and isoleucine; aspartic acid and glutamic acid; asparagine and glutamine; serine, cysteine, and threonine; lysine and arginine; and phenylalanine and tyrosine. Nonpolar hydrophobic amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan, and methionine. Polar neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine. Positively charged (basic) amino acids include arginine, lysine, and histidine. Negatively charged (acidic) amino acids include aspartic acid and glutamic acid. Any substitution of one member of the above polar, basic, or acidic groups with another member of the same group can be considered a conservative substitution. In contrast, a non-conservative substitution is the substitution of one amino acid for another amino acid with dissimilar characteristics.

[0082] In some embodiments, one or more (e.g., 1, 2, 3, 4, or all 5) of positions 3, 4, 5, 6, 7, and 8 of any of the peptides are not substituted. In some embodiments, one or more of positions 3, 4, 5, 6, 7, and 8 of any of the peptides are identical to an amino acid of a peptide in Table 1.

[0083] Also featured are fusion proteins comprising a first amino acid sequence of a peptide described herein (e.g., an unspliced ​​XBP1 peptide described herein, a spliced ​​XBP1 peptide described herein, a CD138 peptide described herein, and / or a CS-1 peptide described herein); and a second amino acid sequence heterologous to the first amino acid sequence.

[0084] The second heterologous amino acid sequence(s) of the peptide generally does not (and is / are selected so as not to) adversely affect the production in cells of an immunogenic peptide of any of SEQ ID NOS: 1-4. The cellular machinery is expected to remove any additional sequence in the peptide to yield an immunogenic peptide of any of SEQ ID NOS: 1-4, which is presented by class I or class II MHC molecules to stimulate an immune response against XBP1-, CD138-, or CS1-expressing cancer cells.

[0085] An amino acid sequence "heterologous" to a first amino acid sequence, or the term "heterologous amino acid sequence," is any amino acid sequence other than the amino acid sequence(s) that flank the first amino acid sequence when it occurs in nature. For example, two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 or more) and / or fewer than 20 (e.g., 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1) carboxy- and / or amino-terminal amino acids immediately adjacent to GLVGLIFAV (SEQ ID NO:3) in human CD138 are not considered heterologous to SEQ ID NO:3. It is understood that a fusion protein comprising a first amino acid sequence that is less than 100% identical to the amino acid sequence of any of SEQ ID NOs: 1 to 4, or that contains one to four conservative substitutions in such an amino acid sequence, may not exist in nature at all.

[0086] In some embodiments, the second amino acid sequence can be a single amino acid. An amino acid "heterologous" to a first amino acid sequence, or the term "heterologous amino acid," is understood to be any amino acid other than the amino acid(s) adjacent to the first amino acid sequence when it occurs in nature. For example, the two amino acids immediately adjacent to GLVGLIFAV (SEQ ID NO: 3) in human CD138 are not considered heterologous to SEQ ID NO: 3.

[0087] The heterologous sequence can be, for example, a sequence used for purification of a recombinant protein (e.g., FLAG, polyhistidine (e.g., hexahistidine), hemagglutinin (HA), glutathione S-transferase (GST), or maltose-binding protein (MBP)). The heterologous sequence can also be a protein useful as a diagnostic or detectable marker, such as luciferase, green fluorescent protein (GFP), or chloramphenicol acetyltransferase (CAT). In some embodiments, the fusion protein can include a signal sequence from another protein, e.g., the KDEL (SEQ ID NO: 11) sequence or any other sequence described herein. In some embodiments, the fusion protein can include all or a portion of an immunoglobulin molecule (e.g., all or a portion of an immunoglobulin heavy chain constant region; see below). In some embodiments, the fusion protein may comprise, for example, a therapeutic or immunostimulatory polypeptide useful in eliciting an immune response (e.g., for antibody generation) (e.g., a T-helper epitope (e.g., the PADRE epitope or the tetanus toxoid universal T-helper cell epitope) or all or a portion of a cytokine or chemokine) and / or a carrier (e.g., KLH). In some embodiments, the fusion protein may comprise one or more linkers, e.g., a linker comprising a peptide sequence (see below). The fusion protein may also comprise a targeting polypeptide. The heterologous sequence may be of various lengths and, in some cases, may be longer than the first amino acid sequence to which the heterologous amino acid sequence is attached. It is understood that a fusion protein comprising a first amino acid sequence and a second amino acid sequence heterologous to the first amino acid sequence does not correspond in sequence to the naturally occurring protein.

[0088] Targeting polypeptides, as used herein, are polypeptides that direct the moiety(s) (e.g., first amino acid sequence) to which they are attached to a specific tissue (e.g., to a lymph node) or cell (e.g., to an antigen-presenting cell or other immune cell), or, in vitro, to a specific isolated molecule or molecular complex. Targeting polypeptides can be, for example, an antibody (immunoglobulin) or an antigen-binding fragment thereof, or a ligand for a cell surface receptor. The antibody (or antigen-binding fragment thereof) can be, for example, a monoclonal antibody, a polyclonal antibody, a humanized antibody, a fully human antibody, a single-chain antibody, a chimeric antibody, or a Fab, F(ab')2, Fab', Fv, or scFv fragment of an antibody. Antibody fragments that include or are the Fc region (with or without the antigen-binding region) can also be used to target reagents to Fc receptor-expressing cells (e.g., antigen-presenting cells, such as interdigitating dendritic cells, macrophages, monocytes, or B cells). The ligand for a cell surface receptor can be, for example, a chemokine, a cytokine (e.g., interleukin 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16) or a death receptor ligand (e.g., FasL or TNFα).

[0089] In some embodiments, the heterologous sequence may be, for example, a "trafficking sequence" that aids in delivery of the peptide into a cell or to a specific compartment of the cell (e.g., the endoplasmic reticulum or Golgi apparatus). Trafficking sequences may include, for example, membrane translocation sequences, transportan sequences, antennapedia sequences, cyclic integrin-binding peptides, and Tat-mediating peptides, or modified versions thereof.

[0090] A linker, e.g., a linker peptide, can directly or indirectly connect a first amino acid sequence to one or more heterologous amino acid sequences. For example, a linker can connect a first amino acid sequence to a second amino acid sequence. A linker peptide can be or include, for example, a stretch of amino acids in which at least four to six amino acids are glycine (see, e.g., Mancebo et al. (1990) Mol. Cell. Biol. 10:2492-2502). A linker peptide can also be or include six or more (e.g., 7, 8, 9, 10, 11, or 12 or more) histidine residues. A linker peptide can be or include at least one (e.g., 1, 2, 3, 4, 5, 6, 7, or 8 or more) protease cleavage site. The protease site can be, for example, a trypsin, chymotrypsin, or factor Xa cleavage site. Such protease sites can be useful, for example, to separate a first amino acid sequence from a heterologous sequence. For example, after expression and purification of a fusion protein comprising a first amino acid sequence linked by a trypsin protease cleavage site to a polyhistidine sequence (in this case used for purification), the polyhistidine sequence can be removed from the first amino acid sequence by contacting the fusion protein with trypsin.

[0091] A first amino acid sequence and a second amino acid sequence can be related to each other in various ways. As used herein, in the context of interactions between two or more atomic or molecular units, "related to" includes any covalent or non-covalent bond or physical intermixing of two or more atomic or molecular units (e.g., a first amino acid sequence and a second amino acid sequence). The chemical nature of covalent bonds (two atoms sharing one or more pairs of valence electrons) is known in the art and includes, for example, disulfide bonds or peptide bonds. A non-covalent bond is a chemical bond between atoms or molecules that does not involve the sharing of a pair of valence electrons. For example, non-covalent interactions include, for example, hydrophobic interactions, hydrogen bonding interactions, ionic bonds, van der Waals bonds, or dipole-dipole interactions. Examples of such non-covalent interactions include antibody-antigen complexation or binding pair interactions (interactions between a first and second member of a binding pair, e.g., the interaction between streptavidin and biotin). Thus, the term "related to" (eg, in the context of a first amino acid sequence and a second amino acid sequence) is understood to be coextensive with the term "comprising."

[0092] In some embodiments, the first amino acid sequence and the second amino acid sequence can be encoded by a single nucleic acid sequence (and can be expressed as a fusion protein from a single nucleic acid sequence). In some cases, the first amino acid sequence and the second amino acid sequence can be encoded by two or more (e.g., three, four, five, or six or more) different nucleic acid sequences. For example, the first amino acid sequence can be encoded by a first nucleic acid sequence, and the second amino acid sequence can be encoded by a second nucleic acid sequence.

[0093] When expressed or produced separately, the first and second amino acid sequences can be crosslinked to each other using any of several known chemical crosslinkers. Examples of such chemical crosslinkers are those that link two amino acid residues via a linkage containing a "hindered" disulfide bond. In these linkages, the disulfide bond within the crosslinking unit is protected (by hindering groups on either side of the disulfide bond) from reduction by, for example, reduced glutathione or the action of the enzyme disulfide reductase. One suitable chemical crosslinker, 4-succinimidyloxycarbonyl-α-methyl-α(2-pyridyldithio)toluene (SMPT), forms such a linkage between two amino acid sequences by utilizing a terminal lysine in one amino acid sequence and a terminal cysteine ​​in the other. Heterobifunctional reagents crosslink via different coupling moieties in each amino acid sequence. In this manner, the resulting "dimer" will be a heterodimer (a peptide comprising a first amino acid sequence and a second amino acid sequence), rather than either a homodimer (e.g., two first amino acid sequences or two second amino acid sequences) or a mixture of homodimers and heterodimers. Thus, the coupling moiety in the first amino acid sequence can be a cysteine ​​residue, and the coupling moiety in the other amino acid sequence can be a lysine residue. Other useful cross-linking agents include, without limitation, chemicals that link two amino groups (e.g., N-5-azido-2-nitrobenzoyloxysuccinimide), two sulfhydryl groups (e.g., 1,4-bis-maleimidobutane), one amino group and one sulfhydryl group (e.g., m-maleimidobenzoyl-N-hydroxysuccinimide ester), one amino group and one carboxyl group (e.g., 4-[p-azidosalicylamido]butylamine), and one amino group and one guanadium group present in the side chain of arginine (e.g., p-azidosalicylamido]butylamine). Didophenylglyoxal monohydrate).

[0094] In some embodiments, the coupling moieties are at the termini (C or N) of each amino acid sequence. As shown above, these can be cysteine ​​residues in each amino acid sequence, or a cysteine ​​in one amino acid sequence and a lysine in the other. If these are two cysteine ​​residues, cross-linking can be achieved, for example, by exposing the amino acid sequences to oxidizing conditions.

[0095] A fusion protein can include a first amino acid sequence and a second amino acid sequence, or the fusion protein can include more than one (e.g., 2, 3, 4, 5, 6, 7, or 8 or more) additional heterologous amino acid sequences. The additional heterologous amino acid sequences can be adjacent to or linked to the amino and / or carboxy termini of the first amino acid sequence.

[0096] When more than two amino acid sequences are linked, at least one of the amino acid sequences may have more than one bridging moiety. For example, a first amino acid sequence may have bridging moieties at the amino and carboxy termini. Such multimers may be constructed "sequentially." Thus, each amino acid sequence is linked to the next amino acid sequence such that only the terminal amino acid sequence in the chain has one residue involved in inter-domain (or inter-drug) binding, while the "internal" amino acid sequence(s) each have two residues involved in inter-domain binding. Alternatively, one amino acid sequence (e.g., a first amino acid sequence) may be linked to multiple (e.g., 2, 3, 4, or 5) other amino acid sequences.

[0097] The combination therapies described herein can include a first component and a second component, where the first component is a peptide described herein. The second component can be, for example, a heterologous amino acid sequence (see above), any other antigenic peptide (e.g., a peptide other than those described herein), a detectable label (see below), a therapeutic or prophylactic agent (see below). For example, a peptide composition can include an amino acid sequence consisting of or consisting essentially of any of SEQ ID NOS: 1-4, and a detectable label, such as a radionuclide.

[0098] In some embodiments, the peptides described herein may have up to 200 (e.g., 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, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, It is understood that the amino acid sequence may have heterologous amino acids (33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190 or 200).

[0099] The peptides described herein can bind to a major histocompatibility complex (MHC) molecule (e.g., an MHC class I molecule or an MHC class II molecule). A "major histocompatibility complex" or "MHC" is a cluster of genes that plays a role in controlling the cellular interactions responsible for the physiological immune response. In humans, the MHC is known as the HLA complex (see, e.g., Paul et al., FUNDAMENTAL IMMUNOLOGY, 3 rd Edition, Raven Press, New York, (1993) and Stites, et al., IMMUNOLOGY, 8 th Edition, Lange Publishing, Los Altos, Calif. (1994).

[0100] "HLA supertype or family" as used herein In this context, the term refers to a set of HLA molecules grouped based on shared peptide binding specificity. HLA class I molecules that share somewhat similar binding affinities for peptides with a particular amino acid motif are grouped into HLA supertypes. The terms HLA superfamily, HLA supertype family, HLA family, and HLA xx-like molecule (where xx indicates a particular HLA type) are synonymous. Types of HLA class I molecules include, for example, HLA-A1, HLA-A2, HLA-A3, HLA-A24, HLA-B7, HLA-B27, HLA-B44, HLA-B58, or HLA-B62. Such HLA molecules are described in detail in U.S. Pat. No. 7,026,443, the entire disclosure of which is incorporated herein by reference in its entirety.

[0101] A peptide can bind to an MHC molecule with high or moderate affinity. As used herein, "high affinity" binding of a peptide to an HLA class I molecule is defined as binding with a dissociation constant (K) of less than 50 nM (e.g., less than 45, 40, 35, 30, 25, 20, 15, 10, 5, 1, 0.5, 0.1 nM, or 0.05 nM). D "Moderate affinity" is defined as binding with a K between about 50 nM and about 500 nM (e.g., 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 115, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, or 500 nM). D"High affinity" binding of a peptide to an HLA class II molecule is a K of less than 100 nM (e.g., less than 95, 90, 85, 80, 75, 70, 65, 60, 55, 50, 45, 40, 35, 30, 25, 20, 15, 10, 5, 1, 0.5, 0.1 nM, or 0.05 nM). D A "moderate affinity" of a peptide for an HLA class II molecule is defined as binding between about 100 nM and about 1000 nM (e.g., 100, 110, 115, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 510, 520, 530, 540, 550, 560, 570, 580, 590, 600, 610, 620, 630, 640, 650, 660, 670, 680, 690, 700, 710, 720, 730, 740, 750, 760, 770, 780, 790, 800, 810, 820, 830, 840, 850, 860, 870, 880, 890, 900, 910, 920, 930, 940, 950, 960, 970, 980, 990, 1000, 1010, K of 0, 500, 510, 520, 530, 540, 550, 560, 570, 580, 590, 600, 610, 620, 630, 640, 650, 660, 670, 680, 690, 700, 710, 720, 730, 740, 750, 760, 770, 780, 790, 800, 810, 820, 830, 840, 850, 860, 870, 880, 890, 900, 910, 920, 930, 940, 950, 960, 970, 980, 990, or 1000 nM) D Methods for determining the binding affinity of a peptide to an MHC molecule are described, for example, in US Pat. No. 7,026,443.

[0102] The peptides described herein can also be associated with MHC molecules and recognized by antigen-specific T cell receptors on T cells. A variety of suitable methods can be used to determine whether a peptide is associated with an MHC molecule and recognized by a T cell receptor on a T cell. For example, peripheral blood lymphocytes (PBLs) from a normal subject can be cultured in vitro with a test peptide in the presence of antigen-presenting cells for a period of several weeks. T cells specific for the peptide are activated during this time and can be detected by, for example, a proliferation assay (carboxyfluoroscein succinimidyl ester (CFSE) assay or 3 H-thymidine assay), limiting dilution assay, cytotoxicity assay (e.g., calcein release assay), or cytokine (e.g., IFNγ) release assay, lymphokine release assay, or 51 Cr release assays (see, e.g., Wentworth, PA et al., Mol. Immunol. 32:603, 1995; Celis, E. et al., Proc. Natl. Acad. Sci. USA 91:2105, 1994; Tsai, V. et al., J. Immunol. 158:1796, 1997; Kawashima, I. et al., Human Immunol. 59:1, 1998, the disclosures of each of which are incorporated herein by reference in their entireties). Suitable in vivo methods include immunizing HLA transgenic mice, where the peptide in adjuvant is administered subcutaneously to the HLA transgenic mice, and several weeks after immunization, splenocytes are removed and cultured in vitro in the presence of the test peptide for approximately one week, and peptide-specific T cells are generated, e.g., 51Cr release assay (see, e.g., Wentworth, PA et al., J. Immunol. 26:97, 1996; Wentworth, PA et al., Int. Immunol. 8:651, 1996; Alexander, J. et al., J. Immunol. 159:4753, 1997, the disclosures of each of which are incorporated herein by reference in their entireties).

[0103] Additionally, direct quantification of antigen-specific T cells can be achieved by incubating T cells with detectably labeled MHC complexes, such as the MHC molecule multimer compositions described in International Application WO2014 / 071402, or HLA-I tetramers (see, e.g., Altman, JD et al., Proc. Natl. Acad. Sci. 2014, 10:101-102, the disclosures of each of which are incorporated herein by reference in their entireties). USA 90:10330, 1993 and Altman, JD et al., Science 274:94, 1996 This can be done by staining with any of the methods described above.

[0104] In some embodiments, peptides may be modified (e.g., amino acids of the peptide may be substituted) to modulate (e.g., increase or decrease) one or more properties of the peptide. For example, one or more (e.g., two, three, or four) amino acids of one of the peptides shown in Table 1 may be substituted to increase the affinity of the peptide for an MHC molecule. In some embodiments, one amino acid (e.g., a T cell receptor contact amino acid residue of the peptide) of a peptide described herein may be modified to enhance the binding interaction between the T cell receptor and the peptide (in the context of an MHC molecule). Such modified peptides are often referred to as "altered peptide ligands" (e.g., see Kalergis et al. (2000) J Immunol. 165(1):280; Conlon et al. (2002) Science 104:107-111, the disclosures of each of which are incorporated herein by reference in their entireties). 1801; and International Publication No. WO02070003).

[0105] Suitable methods for modifying peptides and for determining the effect of the modifications are described, for example, in International Application WO2014 / 071402 and Collins et al. (Immunological Reviews (1998) 163:151-160, the disclosures of each of which are incorporated herein by reference in their entireties. (This is listed in the table below.)

[0106] Suitable methods for producing the peptides and nucleic acids encoding the peptides herein are described, for example, in the sections entitled "Nucleic Acids and Methods for Producing the Peptides" and "Additional Processing of the Peptides" in International Application WO2014 / 071402, which is incorporated herein by reference in its entirety.

[0107] The peptides (and fusion proteins) described herein can be, but need not be, isolated. The term "isolated," as applied to any of the peptides (or fusion proteins) described herein, refers to a peptide, fragment thereof (or, for compositions, macromolecular complex) that has been separated or purified from components that naturally accompany it (e.g., proteins or other naturally occurring biological or organic molecules). Recombinant molecules (e.g., recombinant peptides) are always understood to be "isolated." Typically, a peptide (or fragment or macromolecular complex) is isolated if it constitutes at least 60% by weight of the total molecules of its type in a preparation, e.g., 60% of the total molecules of its type in a sample. For example, a peptide described herein is considered isolated if it constitutes at least 60% by weight of the total protein in a preparation or sample. In some embodiments, the molecules in a preparation consist of at least 75%, at least 90%, or at least 99% by weight of the total molecules of its type in the preparation. A peptide can also be "isolated" if it is present in a mixture with other isolated peptides, for example, a mixture of 2, 3 or 4 different peptides of equal mass.

[0108] In some embodiments, an isolated peptide, fusion protein, peptide coding sequence, fusion protein coding sequence, or vector may be frozen, lyophilized, or immobilized and stored under appropriate conditions that allow the molecule to retain activity (e.g., the ability of a peptide to bind to an MHC molecule, such as an MHC class I molecule, or the ability of a vector to support expression of a peptide in a cell).

[0109] Further processing of peptides After expression or synthesis of any of the peptides (or fusion proteins) described herein, the peptide (or fusion protein) may be further processed. Further processing may include chemical or enzymatic modifications to the peptide (or fusion protein), or if the peptide (or fusion protein) is modified, the processing may include enzymatic or chemical alteration of existing modifications, or both. Further processing of the peptide may include, for example, the addition (covalent or non-covalent linkage) of a heterologous amino acid sequence, such as, but not limited to, any of the heterologous amino acid sequences described above. Enzymatic processing may include contacting the peptide with, for example, one or more proteases, phosphatases, or kinases under conditions that allow the peptide to be modified. Enzymatic processing may include contacting the peptide with one or more enzymes (e.g., oligosaccharyltransferases or mannosidases) capable of glycosylating the peptide or modifying the glycosylation of the peptide.

[0110] This treatment can include, for example, the addition of a detectable label to the peptide. For example, the peptide can be coupled to an enzyme (e.g., horseradish peroxidase, alkaline phosphatase, β-galactosidase, or acetylcholinesterase), a fluorescent material (e.g., umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine, fluorescein, dansyl chloride, allophycocyanin (APC), or phycoerythrin), a luminescent material (e.g., a lanthanide or a chelate thereof), a bioluminescent material (e.g., luciferase, luciferin, or aequorin), or a radionuclide (e.g., 3 H, 32 P, 33 P, 125 I or 35 S), can be detectably labeled.

[0111] This processing may also include coupling of the peptide (or fusion protein) to a polymer (e.g., a polyalkylene glycol moiety, e.g., a polyethylene glycol moiety). In some embodiments, the polymer is coupled to the peptide at a site that is N-terminal on the peptide. In some embodiments, the peptide may contain one or more internal amino acid insertions that provide internal polymer conjugation sites to which a polymer can be conjugated.

[0112] Pembrolizumab The combination therapy herein includes pembrolizumab, a monoclonal anti-PD-1 antibody molecule having the light and heavy chain sequences shown below.

[0113] International Nonproprietary Names for Pharmaceutical Substances The International Journal of Clinical Nucleic Acids (INN) (WHO Drug Information, Vol. 28, No. 3, 2014) (which is incorporated by reference in its entirety, including the section entitled "pembrolizumab" on page 407) provides the heavy and light chain sequences of pembrolizumab as follows: [ka]

[0114] Pembrolizumab can be formulated for parenteral administration, e.g., intravenous administration.

[0115] In embodiments, pembrolizumab is administered at 200 mg every 3 weeks. In embodiments, pembrolizumab is administered every 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 weeks. In embodiments, pembrolizumab is administered intravenously. In embodiments, pembrolizumab is administered intravenously over the course of 30 minutes. In embodiments, pembrolizumab is administered at 2 mg / kg body weight every 3 weeks.

[0116] In embodiments, the combination therapy herein comprises administering pembrolizumab and a peptide composition described herein. In embodiments, the combination therapy herein comprises administering an antibody molecule related to pembrolizumab (e.g., in place of pembrolizumab in any of the methods herein). In embodiments, the antibody molecule comprises a heavy chain sequence having at least 70%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 9. In embodiments, the antibody molecule comprises a light chain sequence having at least 70%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 10. In embodiments, the antibody molecule comprises a VH region having at least 70%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identity to the VH region derived from SEQ ID NO: 9, or has a CH region derived from SEQ ID NO: 9. In embodiments, the antibody molecule comprises a VL region having at least 70%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identity to the VL region derived from SEQ ID NO: 10, or has a VL region derived from SEQ ID NO: 10. In embodiments, the antibody molecule comprises HC CDR1, HC CDR2 and HC CDR3 from SEQ ID NO: 9, wherein these CDRs are defined according to Kabat, Chothia, or a combination of Kabat and Chothia. In embodiments, the antibody molecule comprises LC CDR1, LC CDR2 and LC CDR3 from SEQ ID NO: 10, wherein these CDRs are defined according to Kabat, Chothia, or a combination of Kabat and Chothia.

[0117] As used herein, an "antibody molecule" refers to a protein that includes at least one immunoglobulin variable domain sequence, e.g., an immunoglobulin chain or fragment thereof. The term "antibody molecule" encompasses antibodies and antibody fragments. In certain embodiments, an antibody molecule is a multispecific antibody molecule, e.g., a bispecific antibody molecule.

[0118] The term "antibody," as used herein, refers to a protein or polypeptide sequence derived from an immunoglobulin molecule that specifically binds to an antigen. Antibodies can be polyclonal or monoclonal, multi-chain or single-chain, or intact immunoglobulins, and can be derived from natural or recombinant sources. An antibody can be a tetramer of immunoglobulin molecules.

[0119] The term "antibody fragment" refers to at least a portion of an antibody that retains the ability to specifically interact with an epitope of an antigen (e.g., by binding, steric hindrance, stabilization / destabilization, spatial distribution). Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab'), Fv fragments, scFv antibody fragments, disulfide-linked Fvs (sdFv), Fd fragments consisting of a VH domain and a CH1 domain, linear antibodies, single-domain antibodies such as sdAbs (either VL or VH), camelid VHH domains, multispecific antibodies formed from antibody fragments, e.g., bivalent fragments comprising two Fab fragments linked by a disulfide bridge in the hinge region, and isolated CDR or other epitope-binding fragments of antibodies. Antigen-binding fragments can also be incorporated into single domain antibodies, maxibodies, minibodies, nanobodies, intrabodies, diabodies, triabodies, tetrabodies, v-NARs, and bis-scFvs (see, e.g., Hollinger and Hudson, Nature Biotechnology 23:1126-1136, 2005).

[0120] The term "complementarity determining region" or "CDR" as used herein refers to the sequence of amino acids in an antibody variable region that confers antigen specificity and binding affinity. For example, typically, there are three CDRs (e.g., HCDR1, HCDR2, and HCDR3) in each heavy chain variable region, and three CDRs (LCDR1, LCDR2, and LCDR3) in each light chain variable region. The exact amino acid sequence boundaries of a given CDR can be found in Kabat et al. (1991), "Sequences of Proteins of Immunological Interest," 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD ("Kabat" numbering scheme), Al-Lazikani et al., (1997) JMB 273,927-948 ("Cho The CDR amino acid residues in the heavy chain variable domain (VH) are numbered 31-35 (HCDR1), 50-65 (HCDR2), and 95-102 (HCDR3), and the CDR amino acid residues in the light chain variable domain (VL) are numbered 24-34 (LCDR1), 50-56 (LCDR2), and 89-97 (LCDR3). The CDR amino acid residues in the VH are numbered 26-32 (HCDR1), 52-56 (HCDR2), and 95-102 (HCDR3), and the CDR amino acid residues in the VL are numbered 26-32 (HCDR1), 50-52 (HCDR2), and 91-96 (LCDR3), and the CDR amino acid residues in the VL are numbered 26-32 (LCDR1), 50-52 (LCDR2), and 91-96 (LCDR3). In a combined Kabat and Chothia numbering scheme, in some embodiments, the CDRs correspond to amino acid residues that are part of a Kabat CDR, a Chothia CDR, or both. For example, in some embodiments, the CDRs correspond to amino acid residues 26-35 (HCDR1), 50-65 (HCDR2), and 95-102 (HCDR3) in a VH, e.g., a mammalian VH, e.g., a human VH; and amino acid residues 24-34 (LCDR1), 50-56 (LCDR2), and 89-97 (LCDR3) in a VL, e.g., a mammalian VL, e.g., a human VL.

[0121] Pharmaceutical Composition Any of the peptides, fusion proteins, or other therapeutic agents described herein can be incorporated into pharmaceutical compositions. Such compositions can include one or more peptides (and / or nucleic acids encoding the peptides) and a pharmaceutically acceptable carrier. The composition can further include pembrolizumab. As used herein, the term "pharmaceutically acceptable carrier" includes solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, that are compatible with pharmaceutical administration. One or more peptides can be formulated into pharmaceutical compositions in the form of syrups, elixirs, suspensions, powders, granules, tablets, capsules, lozenges, troches, aqueous solutions, creams, ointments, lotions, gels, emulsions, and the like. Supplementary active compounds (e.g., one or more chemotherapeutic agents) can also be incorporated into the composition. In embodiments, the composition includes two or more (e.g., 2, 3, 4, 5, or 6) peptides described herein. The composition may also include immunogenic peptides other than the immunogenic peptides disclosed herein, for example, peptides derived from WT1 or its derivatives described herein. Other immunogenic peptides include, but are not limited to, immunogenic peptides derived from MUC1, gp100, TRP-2, MAG1, NY-ESO1, HER-2, and AIM2.

[0122] Pharmaceutical compositions are generally formulated to be compatible with their intended route of administration. Examples of routes of administration include oral, rectal, and parenteral, e.g., intravenous, intramuscular, intradermal, subcutaneous, inhalation, transdermal, or transmucosal. Solutions or suspensions used for parenteral application may contain the following components: a sterile diluent, e.g., water for injection, saline solution, fixed oils, polyethylene glycol, glycerine, propylene glycol, or other synthetic solvents. The composition may contain a solvent; an antibacterial agent, such as benzyl alcohol or methylparaben; an antioxidant, such as ascorbic acid or sodium bisulfite; a chelating agent, such as ethylenediaminetetraacetic acid; a buffer, such as acetate, citrate, or phosphate, and an agent for adjusting osmolality, such as sodium chloride or dextrose. The pH can be adjusted using an acid or base, such as hydrochloric acid or sodium hydroxide. The composition may be packaged in a glass or plastic ampule, disposable syringe, or multi-dose vial. Further formulations, for example, for peptide compositions, are described in International Application WO 2014 / 071402, which is incorporated herein by reference in its entirety, including the section entitled "Pharmaceutical Compositions," pages 77-81.

[0123] In embodiments, the one or more peptides are formulated for injection, e.g., subcutaneous injection. In embodiments, pembrolizumab is formulated for injection, e.g., intravenous injection.

[0124] Systemic administration can also be via transmucosal or transdermal means. For transmucosal or transdermal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives. Transmucosal administration can be achieved through the use of nasal sprays or suppositories. For transdermal administration, peptides (or fusion proteins or nucleic acids) can be formulated into ointments, salves, gels, or creams generally known in the art.

[0125] In one embodiment, the compositions (e.g., peptides and / or antibody molecules) herein are administered in a variety of dosage forms, such as sustained release formulations, including implants and microencapsulated delivery systems. The peptide or antibody molecule may be prepared using a carrier that protects it from rapid elimination from the body. Biodegradable, biocompatible polymers, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid, may be used. Methods for preparing such formulations are clear to those skilled in the art. Materials are also commercially available from Alza Corporation and Nova Pharmaceuticals, Inc. Liposomal suspensions (including liposomes directed to APCs, for example, using monoclonal antibodies against APC-specific antigens) may also be used as pharmaceutically acceptable carriers. These may be prepared according to methods known to those skilled in the art, for example, as described in U.S. Pat. No. 4,522,811.

[0126] Any of the pharmaceutical compositions described herein can be included in a container, pack, or dispenser together with instructions for administration, as described below.

[0127] Applicable The combination therapy, composition, pharmaceutical composition and kit herein can be used in various methods.For example, the combination therapy and composition described herein can be used for the following purposes: (i) to induce immune response in subjects with solid tumors, such as breast cancer; (ii) to activate T cells (e.g., central memory T cells and / or effector memory T cells) in culture; and / or (iii) to treat or even prevent solid tumors, such as breast cancer.Solid tumors include, for example, lung cancer, liver cancer, bile duct cancer, gastric cancer, cervical cancer, nasopharyngeal cancer, breast cancer, colon cancer, pancreatic cancer and prostate cancer.In some embodiments, the cancer, such as breast cancer, is recurrent or treatment-resistant.In some embodiments, the cancer is recurrent or treatment-resistant triple-negative breast cancer.

[0128] Although the utility of the combination therapies, compositions, pharmaceutical compositions and kits herein is in no way limited to any of the specific embodiments described herein, exemplary methods in which these reagents may be used are provided below.

[0129] Methods for inducing an immune response The present disclosure also features various methods for inducing an immune response in a subject with cancer, for example, breast cancer. The method for inducing an immune response in a subject with cancer can include administering to the subject one or more of any of the combinations described herein or any of the pharmaceutical compositions described herein. The immune response can be mediated by CD8 + T cells, CD4 + T cells, cytotoxic T lymphocytes (CTLs), T H 1 Response, T H There may be two responses, or a combination of both types of responses.

[0130] The combination therapies herein can be used in a variety of applications, e.g., methods for inducing an immune response in a subject, methods for producing antibodies in a subject, and methods for treating cancer, e.g., breast cancer, e.g., triple-negative breast cancer.

[0131] Any of the methods herein can also be, for example, a method for treating or preventing (preventing against) cancer in a subject (e.g., breast cancer, e.g., triple-negative breast cancer, or any other cancer that expresses XBP1, CD138, or CS1). The terms "prevent," "preventing," or "prevention" are used herein to refer to a particular As used herein with respect to a given treatment for a given condition, they mean that the treated subject never develops a clinically observable level of the condition (e.g., the subject does not exhibit one or more symptoms of the condition, or in embodiments, the subject does not develop a detectable level of cancer).

[0132] As used herein, "treating" a subject with a disorder, e.g., cancer, The terms "treatment," "treatment," or "treating" are used in reference to a given treatment for a given disorder, wherein at least one symptom of the disorder is cured, cured, or cured. Healed, alleviated, relieved, altered, remedied, ameliorated, or improved. Treatment includes administering an amount of a composition effective to alleviate, alleviate, alter, treat, ameliorate, improve, or affect the disorder or symptoms of the disorder. Treatment may inhibit the progression or worsening of symptoms of the disorder, or may cause the condition to progress more slowly and / or to a lesser extent (e.g., fewer symptoms or fewer cancer cells in the subject) than would occur in the absence of treatment. For example, treatment may improve the progression of a condition, e.g., an early stage of cancer that would be predicted to result in a given manifestation of the condition (advanced cancer), during the course of the condition. A condition is said to be "treated" if, given during diagnosis (e.g., detection of a small number of cancer cells in a sample from a subject), the subject experiences fewer and / or milder symptoms of the condition than would otherwise be predicted. A treatment can "treat" a cancer (e.g., breast cancer, e.g., triple-negative breast cancer) if the subject exhibits only mild overt symptoms of the cancer.

[0133] In some embodiments, the cancer is a cancer described herein. For example, the cancer may be bladder (including aggressive and metastatic bladder cancer), breast (e.g., estrogen receptor-positive breast cancer, estrogen receptor-negative breast cancer, HER-2-positive breast cancer, HER-2-negative breast cancer, triple-negative breast cancer, inflammatory breast cancer), colon (including colorectal cancer), kidney (e.g., renal cell carcinoma (e.g., papillary renal cell carcinoma, clear cell carcinoma, chromophobic carcinoma)), liver kidney, lung (including small cell lung cancer and non-small cell lung cancer (including adenocarcinoma, squamous cell carcinoma, bronchoalveolar carcinoma and large cell carcinoma)), urogenital organs, e.g., ovary (fallopian cancer) The cancer may be of the following: uterine, endometrial and peritoneal cancer), cervix, prostate and testes, lymphatic system, rectum, larynx, pancreas (including exocrine pancreatic cancer), stomach (e.g., gastroesophageal, upper stomach or lower stomach cancer), digestive cancer (e.g., anal or bile duct cancer), gallbladder, thyroid, leukemia (e.g., acute myeloid leukemia), neural and glial cell cancer (e.g., glioblastoma multiforme), and head and neck (e.g., nasopharyngeal carcinoma).

[0134] Administration can be by periodic injection of a bolus of the pharmaceutical composition, or can be by external (e.g., IV bag) or internal (e.g., bioerodable implant, bio It can be uninterrupted or continuous, by intravenous or intraperitoneal administration from a reservoir (artificial organ, or colony of implanted reagent-producing cells). See, e.g., U.S. Patent Nos. 4,407,957, 5,798,113, and 5,800,828, each of which is incorporated herein by reference in its entirety.

[0135] Generally, the dosage of peptide or antibody molecule required will depend on the choice of the route of administration; the nature of the formulation; the nature and severity of the subject's illness; the subject's immune status; the subject's size, weight, surface area, age, and sex; other medications being administered; and the judgment of the attending health-care professional.

[0136] Suitable dosages of peptides for inducing an immune response range from 0.000001 to 10 mg of reagent or antigenic / immunogenic composition per kg of subject body weight. Variations in the required dosage are expected given the different efficiencies of various reagents and various routes of administration. For example, nasal or rectal administration may require higher dosages than administration by intravenous injection. Variations in these dosage levels can be adjusted using standard empirical routines for optimization, as is well understood in the art. Administration can be single or multiple (e.g., 2, 3, 4, 6, 8, 10, 20, 50, 100, 150, or more) doses. For example, the peptide(s) can be administered as a primary immunization, followed by one or more subsequent booster immunizations.

[0137] To optimize therapeutic efficacy (e.g., efficacy of one or more peptides or nucleic acids encoding peptides to induce an immune response in a subject), compositions containing peptides can be initially administered in different dosage regimens. The dosage and regimen depend on factors including, for example, the species of mammal, its immune status, and the body weight of the mammal.

[0138] The frequency of dosing of pharmaceutical compositions (for example, pharmaceutical compositions described herein) is within the skill and clinical judgment of medical practitioners (for example, doctors or nurses).Typically, dosing regimen is established by clinical trials that can establish optimal dosing parameters.However, medical practitioners can change such dosing regimen according to the age, health, weight, sex and medical condition of the subject.

[0139] In some embodiments, pharmaceutical compositions can be administered to the subject at least twice (for example, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 15 or 20 times or more).For example, pharmaceutical compositions can be administered to the subject once a month for 3 months; once a week for 1 month; every other week, once a year for 3 years, once a year for 5 years; once every 5 years; once every 10 years; or once every 3 years for life.

[0140] In some embodiments, the reagent may be administered in conjunction with an immune modulator, such as a Toll receptor ligand or an adjuvant (see below).

[0141] As defined herein, a "therapeutically effective amount" of a peptide or a nucleic acid encoding a peptide is an amount of peptide or nucleic acid capable of generating an immune response in a treated subject. Therapeutically effective amounts (i.e., effective dosages) of peptide include milligram, microgram, nanogram, or picogram amounts of reagent per kilogram of subject or sample weight (e.g., about 1 nanogram / kilogram to about 500 micrograms / kilogram, about 1 microgram / kilogram to about 500 milligrams / kilogram, about 100 micrograms / kilogram to about 5 milligrams / kilogram, or about 1 microgram / kilogram to about 50 micrograms / kilogram). A therapeutically effective amount of a nucleic acid also includes microgram, nanogram, or picogram amounts of reagent per kilogram of subject or sample weight (e.g., about 1 nanogram / kilogram to about 500 micrograms / kilogram, about 1 microgram / kilogram to about 500 micrograms / kilogram, about 100 micrograms / kilogram to about 500 micrograms / kilogram, or about 1 microgram / kilogram to about 50 micrograms / kilogram).

[0142] As defined herein, a "prophylactically effective amount" of a peptide or a nucleic acid encoding a peptide is an amount of peptide or nucleic acid capable of generating an immune response against cancer cells (e.g., breast cancer cells) in a treated subject, which immune response can prevent the development of cancer in the subject or substantially reduce the subject's chance of developing or continuing to develop cancer (see above). Prophylactically effective amounts (i.e., effective dosages) of peptides include milligram, microgram, nanogram, or picogram amounts of reagent per kilogram of subject or sample weight (e.g., about 1 nanogram / kilogram to about 500 micrograms / kilogram, about 1 microgram / kilogram to about 500 milligrams / kilogram, about 100 micrograms / kilogram to about 5 milligrams / kilogram, or about 1 microgram / kilogram to about 50 micrograms / kilogram). Prophylactically effective amounts of nucleic acids also include microgram, nanogram, or picogram amounts of reagent per kilogram of subject or sample weight (e.g., about 1 nanogram / kilogram to about 500 micrograms / kilogram, about 1 microgram / kilogram to about 500 micrograms / kilogram, about 100 micrograms / kilogram to about 500 micrograms / kilogram, or about 1 microgram / kilogram to about 50 micrograms / kilogram).

[0143] The subject can be any animal capable of immune response to an antigen, for example, a mammal, for example, a human (e.g., a human patient) or a non-human primate (e.g., a chimpanzee, a baboon, or a monkey), a mouse, a rat, a rabbit, a guinea pig, a gerbil, a hamster, a horse, some livestock (e.g., a cow, a pig, a sheep, or a goat), a dog, a cat, or a whale. The subject can be a subject who has, is suspected of having, or is at risk of developing a solid tumor, for example, breast cancer, for example, triple-negative breast cancer, or any other type of solid tumor that expresses XBP1, CD138, or CS-1 (e.g., lung cancer, liver cancer, bile duct cancer, gastric cancer, cervical cancer, nasopharyngeal cancer, colon cancer, or pancreatic cancer). The subject can be a subject in remission from cancer, for example, breast cancer.

[0144] These methods may also include determining whether one or more cancer cells of the subject's cancer (e.g., breast cancer) express XBP1, CD138, or CS-1 before administering one or more peptides (or nucleic acids) to the subject.The expression of these proteins includes both mRNA and protein expression.Methods for detecting protein and mRNA expression in cells include, for example, enzyme-linked immunosorbent assay (ELISA), Western blotting and dot blotting techniques, or immunohistochemistry techniques for detecting protein, and reverse transcription-polymerase chain reaction (RT-PCR) or Northern blotting techniques for detecting mRNA.

[0145] The peptide or composition may be used in combination with other known therapies. As used herein, "administered in combination" means that two (or more) different treatments are delivered to a subject while the subject is suffering from a disorder, for example, two or more treatments are delivered after the subject is diagnosed with a disorder and before the disorder is cured or eliminated, or before the treatments are stopped for other reasons. In some embodiments, the delivery of one treatment is still occurring when the delivery of a second treatment begins, so that there is an overlap in the period of administration. This may be referred to herein as "simultaneous" or "concurrent delivery." In other embodiments, the delivery of one treatment ends before the delivery of the other treatment begins. In some embodiments in either case, the treatments are more effective due to the combined administration. For example, the second treatment is more effective, for example, an equivalent effect is seen by a smaller amount of the second treatment, or the second treatment reduces symptoms to a greater extent than that seen when the second treatment is administered in the absence of the first treatment, or a similar situation is seen with the first treatment.In some embodiments, the delivery is such that the reduction in symptoms or other parameters related to the disorder is greater than that observed by one treatment delivered in the absence of the other treatment.The effect of the two treatments can be partially additive, completely additive, or greater than additive.The delivery can be such that the effect of the first treatment delivered is still detectable at the time the second treatment is delivered.

[0146] In some embodiments, the delivery is such that the combination therapy produces a greater immune response than that observed in patients treated with the same dose of one or more peptides alone. In some embodiments, the delivery is such that the combination therapy produces a greater clinical response than that observed in patients treated with the same dose of one or more peptides alone. In some embodiments, the delivery is such that the combination therapy produces a greater clinical response than that observed in patients treated with the same dose of pembrolizumab alone. In some embodiments, the delivery is such that the combination therapy produces a similar clinical response between the combination therapy and pembrolizumab monotherapy, where pembrolizumab is administered less frequently or at a lower dose as part of the combination therapy than as monotherapy.

[0147] In some embodiments, the method of treatment comprises administering (a) one or more peptides described herein, (b) pembrolizumab, and (c) one or more additional treatments. In embodiments, the additional treatments comprise surgery, chemotherapy (e.g., adjuvant or neoadjuvant chemotherapy).

[0148] The additional treatment may be, for example, surgery, one or more chemotherapeutic agents, one or more forms of ionizing radiation, and / or one or more immunomodulatory agents.

[0149] The one or more forms of ionizing radiation can be gamma-irradiation, X-ray-irradiation or beta-irradiation.

[0150] Exemplary classes of chemotherapeutic agents include, for example, the following: Alkylating agents (including, without limitation, nitrogen mustards, ethylenimine derivatives, alkyl sulfonates, nitrosoureas, and triazenes): uracil mustard (Aminouracil Mustard®, Chlorethaminacil®, Demethyldopan®, Desmethyldopan®, Haemanthamine®, Nordopan®, Uracil nitrogen mustard, mustard®, Uracillost®, Uracilmostaza®, Uramustin®, Uramustine®), chlormethine (Mustargen®), cyclophosphamide (Cytoxan®, Neosar®, Clafen®, Endoxan®, Procytox®, Revimmune™), ifosfamide (Mitoxana®), melphalan (Alkeran®), chlorambucil (Leukeran®), pipobroman (A medel®, Vercyte®), triethylenemelamine (Hemel®, Hexalen®, Hexastat®), triethylenethiophosphoramine, temozolomide (Temodar®), thiotepa (Thioplex®), busulfan (Busilvex®, Myleran®), carmustine (BiCNU®), lomustine (CeeNU®), streptozocin (Zanosar®), and dacarbazine (DTIC-Dome®).

[0151] Anti-EGFR antibodies (e.g., cetuximab (Erbitux®) and panitumumab (Vectibix®)).

[0152] Anti-HER-2 antibodies (e.g., trastuzumab (Herceptin®)).

[0153] Antimetabolites (including, without limitation, folate antagonists (also referred to herein as antifolates), pyrimidine analogs, purine analogs, and adenosine deaminase inhibitors): methotrexate (Rheumatrex®, Trexall®), 5-fluorouracil (Adrucil®, Efudex®, Fluoroplex®), floxuridine (FUDF®), cytarabine (Cytosar-U®, Tarabine PFS), 6-mercaptopurine (Puri-Nethol®), 6-thioguanine (Thioguanine Tabloid®), fludarabine phosphate (Fludara®), pentostatin (Nipent®), pemetrexed (Alimta®), raltitrexed (Tomudex®), cladribine (Leustatin®), clofarabine (Clofarex®, Clolar®), mercaptopurine (Puri-Nethol®), capecitabine (Xeloda®), nelarabine (Arranon®), azacitidine (Vidaza®), and gemcitabine (Gemzar®). Suitable antimetabolites include, for example, 5-fluorouracil (Adrucil®, Efudex®, Fluoroplex®), floxuridine (FUDF®), capecitabine (Xeloda®), pemetrexed (Alimta®), raltitrexed (Tomudex®), and gemcitabine (Gemzar®).

[0154] Vinca alkaloids: vinblastine (Velban®, Velsar®), vincristine (Vincasar®, Oncovin®), vindesine (Eldisine®), vinorelbine (Navelbine®).

[0155] Platinum-based agents: carboplatin (Paraplat®, Paraplatin®), cisplatin (Platinol®), oxaliplatin (Eloxatin®).

[0156] Anthracyclines: daunorubicin (Cerubidine®, Rubidomycin®), doxorubicin (Adriamycin®), epirubicin (Ellence®), idarubicin (Idamycin®), mitoxantrone (Novantrone®), valrubicin (Valstar®).

[0157] Topoisomerase inhibitors: topotecan (Hycamtin®), irinotecan (Camptosar®), etoposide (Toposar®, VePesid®), teniposide (Vumon®), lamellarin D, SN-38, camptothecin.

[0158] Taxanes: paclitaxel (Taxol®), docetaxel (Taxotere®), larotaxel, cabazitaxel.

[0159] Epothilones: ixabepilone, epothilone B, epothilone D, BMS310705, dehydelone, ZK-epothilone (ZK-EPO).

[0160] Poly ADP-ribose polymerase (PARP) inhibitors: (e.g., BSI201, olaparib (AZD-2281), ABT-888, AG014699, CEP 9722, MK 4827, KU-0059436 (AZD2281), LT-673, 3-aminobenzamide).

[0161] Antibiotics: actinomycin (Cosmegen®), bleomycin (Blenoxane®), hydroxyurea (Droxia®, Hydrea®), mitomycin (Mitozytrex®, Mutamycin®).

[0162] Immunomodulators: lenalidomide (Revlimid®), thalidomide (Thalomid®).

[0163] Immune cell antibodies: alemtuzamab (Campath®), Mutuzumab (Myelotarg®), rituximab (Rituxan®), tositumomab (Bexxar®).

[0164] Interferons (e.g., IFN-alpha (Alferon®, Roferon-A®, Intron®-A) or IFN-gamma (Actimmune®)).

[0165] Interleukins: IL-1, IL-2 (Proleukin®), IL-24, IL-6 (Sigosix®), IL-12.

[0166] An HSP90 inhibitor (e.g., geldanamycin or any of its derivatives). In certain embodiments, the HSP90 inhibitor is selected from geldanamycin, 17-alkylamino-17-desmethoxygeldanamycin ("17-AAG"), or 17-(2-dimethylaminoethyl)amino-17-desmethoxygeldanamycin ("17-DMAG").

[0167] Angiogenesis inhibitors, including without limitation: A6 (Angstrom Pharmaceuticals), ABT-510 (Abbott Laboratories), ABT-627 (atrasentan) (Abbott Laboratories / Xinlay), ABT-869 (Abbott Laboratories), Actimid (CC4047, pomalidomide) (Celgene Corporation ), AdGVPEDF.11D (GenVec), ADH-1 (Exherin )) (Adherex Technologies), AEE788 (Novartis), AG-013736 (axitinib) (Pfizer), AG3340 (prinomastat) (Agouron Pharmaceuticals), AGX1053 (AngioGenex), AGX51 (AngioGenex), ALN-VSP (ALN-VSP O2) (Alnylam Pharmaceuticals), AMG 386 (Amgen), AMG706 (Amgen), apatinib (YN968D1) (Jiangsu Hengrui Medicine), AP23573 (ridaforolimus / MK8669) (Ariad Pharmaceuticals), AQ4N (Novavea), ARQ 197 (ArQule), ASA404 (Novartis / Antisoma), atiprimod (Callisto Pharmaceuticals), ATN-161 (Attenuon), AV-412 (Aveo Pharmaceuticals), AV-951 (Aveo Pharmaceuticals), Avastin (bevacizumab) (Genentech), AZD2171 (cediranib / resentin) (AstraZeneca), BAY 57-9352 (telatinib) (Bayer), BEZ235 (Novartis), BIBF1120 (Boehringer Ingelheim Pharmaceuticals), BIBW 2992 (Boehringer Ingelheim Pharmaceuticals), BMS-275291 (Bristol-Myers Squibb), BMS-582664 (brivanib) (Bristol-Myers Squibb), BMS-690514 (Bristol-Myers Squibb), calcitriol, CCI-779 (Torisel) (Wyeth), CDP-791 (ImClone Systems), Ceflatonin (homoharringtonine / HHT) (ChemGenex Therapeutics), Celebrex (celecoxib) (Pfizer), CEP-7055 (Cephalon / Sanofi), CHIR-265 (Chiron Corporation), NGR-TNF, COL-3 (Metastat) (Collagenex Pharmaceuticals), combretastatin (Oxigene), CP-751,871 (figitumumab) (Pfizer), CP-547,632 (Pfizer), CS-7017 (Daiichi Sankyo) Pharma), CT-322 (Angiocept) (Adnexus), curcumin, dalteparin (Fragmin) (Pfizer), disulfiram (Antabuse), E7820 (Eisai Limited), E7080 (Eisai Limited), EMD 121974 (cilengitide) (EMD Pharmaceuticals), ENMD-1198 (EntreMed), ENMD-2076 (EntreMed), Endostar (Simcere), Erbitux (ImClone / Bristol-Myers Squibb), EZN-2208 (Enzon Pharmaceuticals), EZN-2968 (Enzon Pharmaceuticals), GC1008 (Genzyme), genistein, GSK1363089 (Foretinib) (GlaxoSmithKline), G W786034 (pazopanib) (GlaxoSmithKline), GT-111 (Vascular Biogenics Ltd.), IMC-1121B (ramucirumab) (ImClone Systems), IMC-18F1 (ImClone Systems), IMC-3G3 (ImClone LLC), INCB007839 (Incyte Corporation), INGN 241 (Introgen Therapeutics), Iressa (ZD1839 / gefitinib), LBH589 (Faridak / panobinostat) (Nova rtis), Lucentis (ranibizumab) (Genentech / Novartis), LY317615 (enzastaurin) (Eli Lilly and Company), Macugen (pegaptanib) (Pfizer), MEDI522 (Abegrin) (MedImmune), MLN518 (tandutinib) (Millennium), Neovastat (AE941 / Benefin) (Aeterna Zentaris) , Nexavar (Bayer / Onyx), NM-3 (Genzyme Corporation), Noscapine (Cougar Biotechnology), NPI-2358 (Nereus Pharmaceuticals), OSI-930 (OSI), Palomid 529 (Paloma Pharmaceuticals, Inc.), Panzem Capsules (2ME2) ​​(EntreMed), Panzem NCD (2ME2) ​​(EntreMed), PF-02341066 (Pfizer), PF-04554878 (Pfizer), PI-88 (Progen Industries / Medigen Biotechnology), PKC412 (Novartis), Polyphenon E (green tea extract) (Polypheno E International, Inc), PPI-2458 (Praecis) Pharmaceuticals), PTC299(PTC Therapeutics, PTK787 (vatalanib) (Novartis), PXD101 (belinostat) (CuraGen Corporation), RAD001 (everolimus) (Novartis), RAF265 (Novartis), regorafenib (BAY73-4506) (Bayer), Revlimid (Celgene), Retaane (Alcon Research), SN38 (liposomal) (Neopharm), SNS-032 (BMS-387032) (Sunesis), SOM230 (pasireotide) (Novartis), squalamine (Genaera), suramin, Sutent (Pfizer), Tarceva (Genentech), TB-403 (Thrombogenics), Tempostatin (Collard Biopharmaceuticals), tetrathiomolybdate (Sigma-Aldrich), TG100801 (TargeGen), thalidomide (Celgene Corporation), tinzaparin sodium, TKI258 (Novartis), TRC093 (Tracon Pharmaceuticals Inc.), VEGF Trap (aflibercept) (Regeneron Pharmaceuticals), VEGF Trap-Eye (Regeneron Pharmaceuticals), Veglin (VasGene Therapeutics), bortezomib (Millennium), XL184 (Exelixis), XL647 (Exelixis), XL784 (Exelixis), XL820 (Exelixis), XL999 (Exelixis), ZD6474 (AstraZeneca), vorinostat (Merck), and ZSTK474.

[0168] Antiandrogens, including without limitation: nilutamide (Nilandron®) and bicalutamide (Caxodex®).

[0169] Antiestrogens, including without limitation: tamoxifen (Nolvadex®), toremifene (Fareston®), letrozole (Femara®), testolactone (Teslac®), anastrozole (Arimidex®), bicalutamide (Casodex®), exemestane (Aromasin®), flutamide (Eulexin®), fulvestrant (Faslodex®), raloxifene (Evista®, Keoxifene®), and raloxifene hydrochloride.

[0170] Antihypercalcemic agents, including without limitation: gallium(III) nitrate hydrate (Ganite®) and pamidronate disodium (Aredia®).

[0171] Apoptosis inducers, including, without limitation, ethanol, 2-[[3-(2,3-dichlorophenoxy)propyl]amino]-(9Cl), gambogic acid, embelin, and arsenic trioxide (Trisenox®).

[0172] Aurora kinase inhibitors, including without limitation: binuclein 2.

[0173] Bruton's tyrosine kinase inhibitors, including without limitation: terreic acid.

[0174] Calcineurin inhibitors, including without limitation: cypermethrin, deltamethrin, fenvalerate, and tyrphostin 8.

[0175] CaM kinase II inhibitors, including, without limitation, 5-isoquinolinesulfonic acid, 4-[{2S)-2-[(5-isoquinolinylsulfonyl)methylamino]-3-oxo-3-{4-phenyl-1-piperazinyl)propyl]phenyl ester, and benzenesulfonamide.

[0176] CD45 tyrosine phosphatase inhibitors, including without limitation: phosphonic acids.

[0177] CDC25 phosphatase inhibitors, including without limitation: 1,4-naphthalenedione, 2,3-bis[(2-hydroxyethyl)thio]-(9Cl).

[0178] CHK kinase inhibitors, including without limitation: debromohymenialdisine.

[0179] Cyclooxygenase inhibitors, including without limitation: 1H-indole-3-acetamide, 1-(4-chlorobenzoyl)-5-methoxy-2-methyl-N-(2-phenylethyl)-(9Cl), 5-alkyl substituted 2-arylaminophenylacetic acids and derivatives thereof (e.g., celecoxib (Celebrex®), rofecoxib (Vioxx®), etoricoxib (Arcoxia®), lumiracoxib (Prexige®), valdecoxib (Bextra®), or 5-alkyl-2-arylaminophenylacetic acids).

[0180] cRAF kinase inhibitors, including without limitation: 3-(3,5-dibromo-4-hydroxybenzylidene)-5-iodo-1,3-dihydroindol-2-one and benzamide, 3-(dimethylamino)-N-[3-[(4-hydroxybenzoyl)amino]-4-methylphenyl]-(9Cl).

[0181] Cyclin-dependent kinase inhibitors, including, without limitation, olomoucine and its derivatives, purvalanol B, roascovitine (Seliciclib®), indirubin, kempauron, purvalanol A, and indirubin-3′-monoxime.

[0182] Cysteine ​​protease inhibitors, including without limitation: 4-morpholinecarboxamide, N-[(1S)-3-fluoro-2-oxo-1-(2-phenylethyl)propyl]amino]-2-oxo-1-(phenylmethyl)ethyl]-(9Cl).

[0183] DNA intercalators, including without limitation: plicamycin (Mithracin®) and daptomycin (Cubicin®).

[0184] DNA strand breaking agents, including without limitation: bleomycin (Blenoxane®).

[0185] E3 ligase inhibitors, including without limitation: N-((3,3,3-trifluoro-2-trifluoromethyl)propionyl)sulfanilamide.

[0186] EGF pathway inhibitors, including without limitation: tyrphostin 46, EKB-569, erlotinib (Tarceva®), gefitinib (Iressa®), lapatinib (Tykerb®), and the compounds described in WO 97 / 02266, EP EP 0 564 409, WO99 / 03854, EP 0 520 722, EP 0 566 226, EP 0 787 722, EP 0 837 063, US 5,747,498, WO98 / 10767, WO97 / 30034, WO97 / 49688, WO97 / 38983 and WO96 / 33980.

[0187] Farnesyltransferase inhibitors, including without limitation: A-hydroxyfarnesylphosphonic acid, butanoic acid, 2-[(2S)-2-[[(2S,3S)-2-[[(2R)-2-amino-3-mercaptopropyl]amino]-3-methylpentyl]oxy]-1-oxo-3-phenylpropyl]amino]-4-(methylsulfonyl)-1-methylethyl ester (2S)-(9Cl), and manumycin A.

[0188] Flk-1 kinase inhibitors, including without limitation: 2-propenamide, 2-cyano-3-[4-hydroxy-3,5-bis(1-methylethyl)phenyl]-N-(3-phenylpropyl)-(2E)-(9Cl).

[0189] Glycogen synthase kinase-3 (GSK3) inhibitors, including without limitation: indirubin-3'-monoxime.

[0190] Heat shock protein 90 (Hsp90) chaperone modulators, including without limitation: AUY922, STA-9090, ATI13387, MCP-3100, IPI-504, IPI-493, SNX-5422, Debio0932, HSP990, DS-2248, PU-H71, 17-DMAG (Alvespimycin), and XL888.

[0191] Histone deacetylase (HDAC) inhibitors, including, without limitation, suberoylanilide hydroxamic acid (SAHA), [4-(2-amino-phenylcarbamoyl)-benzyl]-carbamic acid pyridin-3-ylmethyl ester and its derivatives, butyric acid, pyroxamide, trichostatin A, oxamflatin, apicidin, depsipeptide, depudecin, trapoxin, and WO02 The compound disclosed in / 22577.

[0192] I-kappa B-alpha kinase inhibitors (IKK), including without limitation: 2-propenenitrile, 3-[(4-methylphenyl)sulfonyl]-(2E)-(9Cl).

[0193] Imidazotetrazinones, including without limitation: temozolomide (Methazolastone®), Temodar® and its derivatives (e.g., as disclosed generically and specifically in US 5,260,291), and Mitozolomide.

[0194] Insulin-like growth factor pathway inhibitors, for example, IGF inhibitors or IGF receptor (IGFR1 or IGFR2) inhibitors, include, without limitation: small molecule inhibitors, for example, OSI-906; anti-IGF or anti-IGFR antibodies, for example, AVE-1642, MK-0646, IMC-A12 (cixutumab), R1507, CP-751 ,871 (figitumumab).

[0195] Insulin tyrosine kinase inhibitors, including without limitation: hydroxyl-2-naphthalenylmethylphosphonic acid.

[0196] c-Jun-N-terminal kinase (JNK) inhibitors, including without limitation: pyrazoleanthrone and epigallocatechin gallate.

[0197] Mitogen-activated protein kinase (MAP) inhibitors, including without limitation: benzenesulfonamide, N-[2-[[[3-(4-chlorophenyl)-2-propenyl]methyl]amino]methyl]phenyl]-N-(2-hydroxyethyl)-4-methoxy-(9Cl).

[0198] MDM2 inhibitors, including without limitation: trans-4-iodo, 4'-boranyl-chalcone.

[0199] MEK inhibitors, including without limitation: butanedinitrile, bis[amino[2-aminophenyl)thio]methylene]-(9Cl).

[0200] MMP inhibitors, including without limitation: actinonin, epigallocatechin gallate, collagen peptidomimetic and non-peptidomimetic inhibitors, tetracycline derivatives Marimastat®, prinomastat, incyclinide (Metastat®), shark cartilage extract AE-94 1 (Neovastat®), Tanomastat, TAA211, MMI270B or AAJ996.

[0201] mTor inhibitors, including without limitation: rapamycin (Rapamune®) and its analogs and derivatives, AP23573 (also known as ridaforolimus, deforolimus, or MK-8669), CCI-779 (also known as temsirolimus) (Torisel®), and SDZ-RAD.

[0202] NGFR tyrosine kinase inhibitors, including without limitation: Tyrphostin AG 879.

[0203] p38 MAP kinase inhibitors, including without limitation: phenol, 4-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-imidazol-2-yl]-(9Cl) and benzamide, 3-(dimethylamino)-N-[3-[(4-hydroxylbenzoyl)amino]-4-methylphenyl]-(9Cl).

[0204] p56 tyrosine kinase inhibitors, including without limitation: damnacanthal and tyrphostin 46.

[0205] PDGF pathway inhibitors, including without limitation: tyrphostin AG 1296, tyrphostin 9, 1,3-butadiene-1,1,3-tricarbonitrile, 2-amino-4-(1H-indol-5-yl)-(9Cl), imatinib (Gleevec®) and gefitinib (Iressa®) and compounds disclosed generically and specifically in European Patent No. 0 564 409 and PCT Publication No. WO 99 / 03854.

[0206] Phosphatidylinositol 3-kinase inhibitors, including without limitation: wortmannin and quercetin dihydrate.

[0207] Phosphatase inhibitors, including without limitation: cantharidic acid, cantharidin, and L-leucinamide.

[0208] PKC inhibitors, including without limitation: 1-H-pyrrolo-2,5-dione, 3-[1-[3-(dimethylamino)propyl]-1H-indol-3-yl]-4-(1H-indol-3-yl)-(9Cl), bisindolylmaleimide IX, sphinogosine, staurosporine, and hypericin.

[0209] PKC delta kinase inhibitors, including without limitation: rottlerin.

[0210] Polyamine synthesis inhibitors, including without limitation: DMFO.

[0211] Proteasome inhibitors, including without limitation: aclacinomycin A, gliotoxin, and bortezomib (Velcade®).

[0212] Protein phosphatase inhibitors, including without limitation: cantharidic acid, cantharidin, LP-bromotetramisole oxalate, 2(5H)-furanone, 4-hydroxy-5-(hydroxymethyl)-3-(1-oxohexadecyl)-(5R)-(9Cl), and benzylphosphonic acid.

[0213] Protein tyrosine kinase inhibitors, including without limitation: tyrphostin Ag 216, tyrphostin Ag 1288, tyrphostin Ag 1295, geldanamycin, genistein, and 7H-pyrrolo[2,3-d]pyrimidine derivatives;

[0214] PTP1B inhibitors, including without limitation: L-leucinamide.

[0215] SRC family tyrosine kinase inhibitors, including without limitation: PP1 and PP2.

[0216] Syk tyrosine kinase inhibitors, including without limitation: piceatannol.

[0217] Janus (JAK-2 and / or JAK-3) tyrosine kinase inhibitors, including without limitation: Tyrphostin AG 490 and 2-naphthyl vinyl ketone.

[0218] Retinoids, including without limitation: isotretinoin (Accutane®, Amnesteem®, Cistane®, Claravis®, Sotret®) and tretinoin (Aberel®, Aknoten®, Avita®, Renova®, Retin-A®, Retin-A MICRO®, Vesanoid®).

[0219] RNA polymerase II elongation inhibitors, including without limitation: 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole.

[0220] Serine / threonine kinase inhibitors, including without limitation: 2-aminopurine.

[0221] Sterol biosynthesis inhibitors, including without limitation: squalene epoxidase and CYP2D6.

[0222] VEGF pathway inhibitors, including, without limitation, anti-VEGF antibodies, such as bevacizumab, and small molecules, such as sunitinib (Sutent®), sorafinib (Nexavar®), ZD6474 (also known as vandetanib) known as cediranib) (Zactima™), SU6668, CP-547632, AV-951 (tivozanib) and AZD2171 (also known as cediranib) (Recentin™).

[0223] For example, the one or more chemotherapeutic agents may be selected from the group consisting of cisplatin, carboplatin, procarbazine, mechlorethamine, cyclophosphamide, camptothecin, adriamycin, ifosfamide, melphalan, chlorambucil, bisulfan, nitrosurea, dactinomycin, daunorubicin, doxorubicin, bleomycin, plicamycin, mitomycin, etoposide, verampil, podophyllotoxin, tamoxifen, taxol, thalidomide, lenalidomide, proteosome inhibitors (e.g., bortezomib), hsp90 inhibitors (e.g., tenespinmycin), transplatin, 5-fluorouracil, vincristine, vinblastine, methotrexate, or an analog of any of the foregoing. Immunomodulatory agents include, for example, various chemokines and cytokines, such as interleukin 2 (IL-2), granulocyte / macrophage colony-stimulating factor (GM-CSF), and interleukin 12 (IL-12).

[0224] In one embodiment, the additional therapy is one or more additional immunogenic peptides, for example, one or more immunogenic peptides derived from WT1 or derivatives thereof. Exemplary WT1 immunogenic peptides include, but are not limited to, the following WT1 class 1 epitopes: a peptide comprising (or consisting of) RMFPNAPYL (WT1 126-134); a peptide comprising (or consisting of) YMFPNAPYL; a peptide comprising (or consisting of) RSDELVRHHNMHQRNMTKL (WT1 427-445); a peptide comprising (or consisting of) PGCNKRYFKLSHLQMHSRKHTG (WT1 331-352); a peptide comprising (or consisting of) SGQARMFPNAPYLPSCLES (WT1 122-140); and a peptide comprising (or consisting of) SGQAYMFPNAPYLPSCLES. Other WT1 immunogenic peptides are described in U.S. Patent No. 7,598,221, the contents of which are incorporated herein by reference. Other immunogenic peptides include, but are not limited to, immunogenic peptides derived from MUC1, gp100, TRP-2, MAG1, NY-ESO1, HER-2, and AIM2.

[0225] The subject may have, be suspected of having, or be at risk of developing cancer, such as breast cancer, for example, triple-negative breast cancer, for example, metastatic triple-negative breast cancer. A subject "suspected of having cancer" is a subject who has one or more symptoms of cancer, or one or more laboratory test results, such as blood test results, that suggest cancer. Symptoms of cancer are well known to those skilled in the art and generally include, but are not limited to, pain, weight loss, weakness, excessive fatigue, difficulty eating, loss of appetite, chronic cough, worsening shortness of breath, hemoptysis, blood in the urine, blood in the stool, nausea, vomiting, abdominal distension, flatulence, fluid in the abdominal cavity, vaginal bleeding, constipation, abdominal distension, colon perforation, acute peritonitis (infection, fever, pain), pain, vomiting blood, excessive sweating, fever, high blood pressure, anemia, diarrhea, jaundice, dizziness, chills, muscle cramps, difficulty swallowing, etc.

[0226] As used herein, a subject "at risk of developing cancer" is one who has a predisposition to developing cancer, i.e., a genetic predisposition to developing cancer, such as a mutation in a tumor suppressor gene (e.g., a mutation in BRCA1, p53, RB, or APC), has been exposed to a condition that may cause cancer, or is currently affected by such a condition. Thus, a subject can also be "at risk of developing cancer" if the subject has been exposed to mutagenic or carcinogenic levels of a particular compound (e.g., carcinogenic compounds in cigarette smoke, such as acrolein, 4-aminobiphenyl, aromatic amines, benzene, benz{a}anthracene, benzo{a}pyrene, formaldehyde, hydrazine, polonium-210 (radon), urethane, or vinyl chloride). A subject may be "at risk for developing cancer" if the subject has been exposed to, for example, high doses of ultraviolet light or X-ray radiation, or if the subject has been exposed to (e.g., infected with) a tumor-causing / associated virus, such as papillomavirus, Epstein-Barr virus, hepatitis B virus, or human T-cell leukemia-lymphoma virus. Additionally, a subject may be "at risk for developing cancer" if the subject suffers from inflammation (e.g., chronic inflammation).

[0227] Triple-negative breast cancer refers to breast cancer that is negative for estrogen receptor (ER), progesterone receptor (PR) and HER2. The presence or absence of ER, PR and HER2 can be evaluated, for example, by immunohistochemistry or quantitative PCR on biopsy samples. This type of cancer is often treated with a combination of surgery and chemotherapy.

[0228] In some embodiments, the patient is human leukocyte antigen (HLA)-A2 positive.

[0229] In some embodiments, the patient has 0 or 1 Eastern Cooperative Eastern Cooperative Oncology Group (ECOG) performance status; platelet count ≥ 75 × 10 9 / L and absolute neutrophil count (ANC) ≥ 1.0 × 10 9 / L; and / or have adequate liver function as evidenced by bilirubin ≦2.0 mg / dL and alanine transaminase (ALT) and aspartate transaminase (AST) ≦2.5 × upper limit of normal (ULN).

[0230] In embodiments, (i) pembrolizumab and (ii) one or more peptides are administered in an amount sufficient to increase progression-free survival (PFS) compared to the predicted course of disease without treatment, or compared to the predicted course of disease upon treatment with (i) alone or (ii) alone, or compared to the predicted course of disease upon standard of care treatment. In embodiments, (i) and (ii) are administered in an amount sufficient to increase overall survival (OS) compared to the predicted course of disease without treatment, or compared to the predicted course of disease upon treatment with (i) alone or (ii) alone, or compared to the predicted course of disease upon standard of care treatment. In embodiments, (i) pembrolizumab and (ii) one or more peptides are administered in an amount sufficient to obtain a clinical response (e.g., as measured by PFS or OS) similar to that of monotherapy with pembrolizumab, e.g., a clinical response with fewer adverse effects. PFS and OS may be determined according to the Response Evaluation Criteria in Solid Tumors (RECIST) guidelines (version 1.1). In embodiments, (i) and (ii) are administered in amounts sufficient to induce an immune response to one or more of the administered peptides, e.g., a higher immune response than observed without treatment, or upon treatment with (i) alone or (ii) alone, or compared to the expected course of disease with standard of care treatment. The immune response can be measured, for example, by ELISPOT assay or as described in International Application WO2014 / 071402, which is incorporated herein by reference in its entirety.

[0231] In some embodiments, the method may also include a step of determining whether an immune response has occurred in the subject after administering a combination therapy described herein to the subject. Suitable methods for determining whether an immune response has occurred in the subject include, for example, the use of an immunoassay to detect the presence of antibodies specific to the peptide in a biological sample from the subject. For example, after administering a peptide or composition to the subject, a biological sample (e.g., a blood sample) can be obtained from the subject and tested for the presence of antibodies specific to the peptide(s). An immune response can also be detected by assaying for the presence or amount of activated T cells in the sample. Such assays include, for example, a proliferation assay, a limiting dilution assay, a cytotoxicity assay (e.g., a lymphokine release assay or 51 Cr release assay) or flow cytometry assay.

[0232] In some embodiments, these methods can also include determining whether the subject has cancer.The appropriate method for such determination depends on the type of cancer detected in the subject, but is known in the art.Such method can be qualitative or quantitative.Methods for diagnosing breast cancer include mammogram, ultrasound, MRI, biopsy and molecular testing.Methods for diagnosing triple-negative breast cancer include immunohistochemistry for PR, ER and HER2.

[0233] Methods for choosing treatment A method for selecting a treatment for a subject having cancer, e.g., breast cancer, or any cancer in which XBP1, CD138, or CS1 is expressed, optionally includes determining whether one or more cells of the subject's cancer (e.g., breast cancer cells) express XBP1; and if one or more cells express XBP1, selecting (i) pembrolizumab and (ii) a peptide or composition described herein, e.g., an XBP1 peptide described herein or a composition comprising an XBP1 peptide, as a treatment for the subject.

[0234] A method for selecting a treatment for a subject having cancer, e.g., breast cancer, may optionally include determining whether one or more cells of the subject's cancer (e.g., breast cancer cells) express CD138; and if one or more cells express CD138, selecting (i) pembrolizumab and (ii) a peptide or composition described herein, e.g., a CD138 peptide described herein or a composition comprising a CD138 peptide, as a treatment for the subject.

[0235] A method for selecting a treatment for a subject having cancer, e.g., breast cancer, may optionally include determining whether one or more cells of the subject's cancer (e.g., breast cancer cells) express CS-1; and if the one or more cells express CS-1, selecting (i) pembrolizumab and (ii) a peptide or composition described herein, e.g., a CS-1 peptide or a composition comprising a CS-1 peptide described herein, as a treatment for the subject.

[0236] It is understood that if one or more cells of a subject's cancer (e.g., breast cancer cells) express two or more of XBP1, CD138, and CS-1, a suitable peptide combination can be delivered to the subject in combination with pembrolizumab, for example, via a composition described herein. Methods for determining whether one or more cells express XBP1, CD138, or CS-1 are described, for example, in the section entitled "Methods for Selecting a Therapy," pages 104-107 of International Application WO2014 / 071402, which is incorporated herein by reference in its entirety.

[0237] Kits and Products The present disclosure also features various kits. The kits may include, for example, (i) pembrolizumab, (ii) one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more) of any of the peptides or compositions described herein (or expression vectors containing nucleic acid sequences encoding one or more peptides), and (iii) instructions for administering the peptides or compositions to a subject. In embodiments, (ii) includes one or more, e.g., all, of: (a) an unspliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 1; (b) a spliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 2; or (c) a CD138 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 3. In an embodiment, (ii) includes one or more, e.g., all, of: (a) an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a spliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2; or (c) a CD138 peptide consisting of the amino acid sequence of SEQ ID NO: 3. The kit may include one or more pharmaceutically acceptable carriers and / or one or more immunostimulatory agents and / or one or more immunomodulatory agents. The immunostimulatory agent may be, for example, a T-helper epitope, an altered peptide ligand, or an adjuvant. In one embodiment, the immunostimulatory agent may be carboxymethylcellulose, a combination of polyinosinic acid-polycytidylic acid and poly-L-lysine double-stranded RNA (e.g., poly IC-LC, e.g., hiltonol); a water-oil emulsion (e.g., montanide); or a protein (e.g., a cytokine, complement, GCSF, or GM-CSF). In one embodiment, the additional agent may be a protein, e.g., an antibody. In one embodiment, the additional agent is an immune checkpoint inhibitor. For example, the antibody that inhibits an immune checkpoint molecule can be an anti-CTLA4 antibody, such as ipilimumab or tremelimumab, or an anti-PD-1 antibody, or an anti-PDL-1 antibody.In one embodiment, the additional agent may be a small molecule adjuvant, such as thalidomide or a thalidomide derivative, such as lenalidomide.

[0238] The kit may also include one or more therapeutic, diagnostic, or prophylactic agents, including, but not limited to: (i) agents that modulate inflammatory responses (e.g., aspirin, indomethacin, ibuprofen, naproxen, steroids, cromolyn sodium, or theophylline); (ii) agents that affect renal and / or cardiovascular function (e.g., furosemide, thiazides, amiloride, spironolactone, captopril, enalapril, lisinopril, diltiazem, nifedipine, verapamil, digoxin, isordil, dobutamine, lidocaine, quinidine, adenosine, digitalis, mevastatin). , lovastatin, simvastatin or mevalonate; (iii) drugs affecting gastrointestinal function (e.g., omeprazole or sucralfate); (iv) antibiotics (e.g., tetracycline, clindamycin, amphotericin B, quinine, methicillin, vancomycin, penicillin G, amoxicillin, gentamicin, erythromycin, ciprofloxacin, doxycycline, streptomycin, gentamicin, tobramycin, chloramphenicol, isoniazid, fluconazole or amantadine); (v) anticancer drugs (e.g., cyclophosphamide, methotrexate, fluorouracil, cytarabine, mercaptopurine, vinblastine, vincristine, doxorubicin, bleomycin, mitomycin C, hydroxyurea, prednisone, tamoxifen, cisplatin or decarbazine); (vi) immunomodulatory agents (e.g., Interleukins, interferons (e.g., interferon gamma (IFN-γ), granulocyte-macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor alpha (TNFα), tumor necrosis factor beta (TNFβ), cyclosporine, FK506, azathioprine, steroids); (ix) drugs acting on the blood and / or blood-forming organs (e.g., interleukins, G-CSF, GM-CSF, erythropoietin, heparin, warfarin, or coumarin); or ( vii) Hormones (e.g., growth hormone (GH), prolactin, luteinizing hormone, TSH, ACTH, insulin, FSH, CG, somatostatin, estrogen, androgen, progesterone, gonadotropin-releasing hormone (GnRH), thyroxine, triiodothyronine); hormone antagonists; drugs affecting mineralization and bone turnover (e.g., calcium, phosphate, parathyroid hormone (PTH), vitamin D, bisphosphonates, calcitonin, fluoride).

[0239] Other embodiments While the present invention has been described in conjunction with its detailed description, the above description is intended to be illustrative, not limiting, of the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims. In certain embodiments, for example, the following items are provided: (Item 1) 1. A method of treating breast cancer, e.g., triple-negative breast cancer, comprising: (i) pembrolizumab; and (ii) one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. to a subject, wherein the subject has or is at risk of developing breast cancer, e.g., triple-negative breast cancer. (Item 2) For use in treating breast cancer, e.g., triple-negative breast cancer, Pembrolizumab in combination with one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. (Item 3) 1. A method for inducing an immune response in a subject, comprising: (i) pembrolizumab; and (ii) one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. The method comprises delivering to a subject. (Item 4) For use in inducing an immune response in a subject, one or more of the unspliced ​​XBP1 peptides described herein, the spliced ​​XBP1 peptides described herein, the CD138 peptides described herein, and the CS-1 peptides described herein. and pembrolizumab in combination. (Item 5) 5. The method or composition for use according to any one of items 1 to 4, wherein the unspliced ​​XBP1 peptide is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 1. (Item 6) 5. The method or composition for use according to any one of items 1 to 4, wherein the spliced ​​XBP1 peptide is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 2. (Item 7) 8. The method or composition for use according to any one of items 1 to 4, wherein the CD138 peptide is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 3. 5. The method or composition for use according to any one of items 1 to 4, wherein the CS-1 peptide is 35 amino acids or less in length and comprises the amino acid sequence of SEQ ID NO: 4. (Item 9) 5. The method or composition for use according to any one of items 1 to 4, wherein the unspliced ​​XBP1 peptide consists of the amino acid sequence of SEQ ID NO: 1. (Item 10) 5. The method or composition for use according to any one of items 1 to 4, wherein the spliced ​​XBP1 peptide consists of the amino acid sequence of SEQ ID NO: 2. (Item 11) 5. The method or composition for use according to any one of items 1 to 4, wherein the CD138 peptide consists of the amino acid sequence of SEQ ID NO: 3. (Item 12) 5. The method or composition for use according to any one of items 1 to 4, wherein the CS-1 peptide consists of the amino acid sequence of SEQ ID NO: 4. (Item 13) The method or composition for use according to any of the preceding items, wherein the method comprises administering, or the composition for use comprises: (a) an unspliced ​​XBP1 peptide described herein; (b) a splice form of an XBP1 peptide described herein, and (c) A CD138 peptide as described herein. (Item 14) The method or composition for use according to any of the preceding items, wherein the method comprises administering, or the composition for use comprises: (a) an unspliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 1; (b) a spliced ​​XBP1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO: 2; and (c) A CD138 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO:3. (Item 15) The method or composition for use according to any of the preceding items, wherein the method comprises administering, or the composition for use comprises: (a) an unspliced ​​XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 1; (b) a splice-form XBP1 peptide consisting of the amino acid sequence of SEQ ID NO: 2, and (c) CD138 peptide consisting of the amino acid sequence of SEQ ID NO:3. (Item 16) 16. The method or composition for use according to any of items 13 to 15, wherein the method comprises administering, or the composition for use further comprises: (d) A CS-1 peptide described herein. (Item 17) 16. The method or composition for use according to any of items 13 to 15, wherein the method comprises administering, or the composition for use further comprises: (d) A CS-1 peptide having a length of 35 amino acids or less and comprising the amino acid sequence of SEQ ID NO:4. (Item 18) 16. The method or composition for use according to any of items 13 to 15, wherein the method comprises administering, or the composition for use further comprises: (d) CS-1 peptide consisting of the amino acid sequence of SEQ ID NO: 4. (Item 19) 10. The method or composition for use according to any preceding item, wherein the method further comprises administering in combination with one or more immunostimulatory agents, or the composition for use further comprises administering in combination with one or more immunostimulatory agents. (Item 20) 20. The method or composition for use according to item 19, wherein the immunostimulatory agent is selected from an adjuvant comprising carboxymethylcellulose, polyinosinic acid-polycytidylic acid, and poly-L-lysine double-stranded RNA (e.g., poly IC-LC, e.g., hiltonol); an adjuvant comprising a water-oil emulsion (e.g., montanide); and an adjuvant comprising a protein (e.g., a cytokine, GCSF, or GM-CSF). (Item 21) The method or composition for use according to any preceding item, wherein the method comprises administering, or the composition for use further comprises: Further treatments, such as one or more chemotherapeutic agents, one or more forms of ionizing radiation, one or more immunotherapeutic agents (e.g., a cancer vaccine, an immune checkpoint inhibitor), one or more immune checkpoint inhibitors, e.g., an antibody that inhibits an immune checkpoint molecule (e.g., an anti-CTLA4 antibody, e.g., ipilimumab or tremelimumab, a PD-1 antibody, or a PDL-1 antibody), or an adjuvant, e.g., a small molecule adjuvant (e.g., thalidomide or a thalidomide derivative, e.g., lenalidomide). (Item 22) 2. The method or composition for use according to any preceding item, wherein the method comprises administering PolyIC-LC, or the composition for use further comprises PolyIC-LC. (Item 23) 3. The method or composition for use according to item 1 or 2, wherein the breast cancer is triple-negative breast cancer. (Item 24) 5. The method or composition for use according to item 3 or 4, wherein the subject has, is at risk of developing, or is suspected of having cancer, such as breast cancer, such as triple-negative breast cancer. (Item 25) The method or composition for use according to any preceding item, wherein the breast cancer is metastatic breast cancer, for example, metastatic triple-negative breast cancer. (Item 26) The method or composition for use according to any preceding item, wherein the subject has, or is identified as having, one or more cancer cells that express XBP1, CD138 or CS1, or any combination thereof. (Item 27) 10. The method of any preceding item, further comprising, after delivering the composition to the subject, determining whether an immune response has occurred in the subject. (Item 28) The method of any preceding item, wherein the subject is a human. (Item 29) The method of any preceding item, wherein the subject is in remission from breast cancer, e.g., triple-negative breast cancer. (Item 30) The method of any preceding item, wherein (i) and (ii) are administered separately or together. (Item 31) The method of any preceding item, wherein (i) is administered before, together with, or after (ii). (Item 32) A composition for use according to any of the preceding items, wherein (i) and (ii) are formulated for use, separately or together. (Item 33) The composition for use according to any of the preceding items, wherein (i) is formulated for administration before, together with, or after (ii). (Item 34) The method or composition for use according to any preceding item, wherein the pembrolizumab is administered at a dose of 200 mg. (Item 35) The method or composition for use according to any preceding item, wherein the pembrolizumab is administered every three weeks. (Item 36) The method or composition for use according to any preceding item, wherein the one or more peptides are administered at a dose of 0.8 mg of total peptide, for example, 0.2 mg of the unspliced ​​XBP1 peptide, 0.2 mg of the spliced ​​XBP1 peptide, 0.2 mg of the CD138 peptide, and 0.2 mg of the CS-1 peptide. (Item 37) The method or composition for use according to any of the preceding items, wherein the one or more peptides are administered at a dose of 0.6 mg of total peptide, for example, 0.2 mg of the unspliced ​​XBP1 peptide, 0.2 mg of the spliced ​​XBP1 peptide, and 0.2 mg of the CD138 peptide. (Item 38) The method or composition for use according to any preceding item, wherein the one or more peptides are administered once a week, for example for at least 1, 2, 3, 4, 5, or 6 weeks.

Claims

[Claim 1] A method, combination, or composition as described in the specification.

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