Cancer treatment utilizing pre-existing microbial immunity

Abstract

To provide methods, compositions and kits for mobilizing a pre-existing immune response in an individual to reduce or stabilize a cancer in the individual.SOLUTION: A pharmaceutical composition for treating a solid tumor in an individual contains a drug for enhancing the immune response and at least one MHC II-restricted peptide from a cytomegalovirus (CMV) protein, where the CMV protein is selected from the group consisting of pp65, gB, IE-1, gH and gL, where the peptide mobilizes a pre-existing natural immune response to a tumor site, thereby treating the tumor, and where the pharmaceutical composition is for injection into solid tumors.SELECTED DRAWING: Figure 1-1

Description

[Technical field]

[0001] Cross-references to related applications This application claims the benefit of U.S. Provisional Patent Application No. 62 / 582,097, filed Nov. 6, 2017, which is incorporated by reference in its entirety.

[0002] Technical Field The present invention relates to immunology and cancer therapy, including methods, compositions and kits for directing a patient's existing immune response against cancer. [Background technology]

[0003] background Persistent asymptomatic viral infections are usually controlled by cellular and / or humoral immunity in healthy individuals, but can reactivate in immunocompromised individuals. Cellular immunity to some chronic viral infections increases with age, leading to the induction of many fully functional virus-specific T cells. Cytomegalovirus (CMV) is a β-herpesvirus that is widespread worldwide (50–90% of the human population is infected) and largely asymptomatic in healthy individuals. CMV establishes a lifelong persistent infection and requires persistent cellular immunity to prevent disease progression. Hence, CMV reactivation is a threat in immunosuppression, e.g., in hematopoietic stem cell transplantation. In immunocompetent individuals, CD4 and CD8 T cell responses to CMV are broad-spectrum to multiple CMV antigens. It exhibits high reactivity and intensity, has a high prevalence in the general human population, and increases with age (M. Bajwa et al., J Infect Dis 215, 1212-20(2017)). Inflation of memory T cells is a hallmark of persistent CMV infection and has been extensively studied in humans. CMV-specific CD8+ T cell responses are either inflationary or They are divided into two types, depending on whether they are quiescent (non-inflationary) or quiescent upon resolution of the primary infection (GA O'Hara, Trends Immunol 33:84-90 (2012)). The nature of the antigen and the pattern of antigen expression during persistent CMV infection result in CD8+ T cells with a memory phenotype (non-inflationary) or an effector phenotype. Murine CMV infection also results in CD8+ T cells that are resistant to human CMV. They establish lifelong persistent infection by inducing an immune response that mimics that of the host (Id).

[0004] Induction of antitumor T cell responses is of paramount importance in the development of effective immunotherapy against cancer. Only a portion of cancer patients respond to current immunotherapies. The generation of T cell immunity against cancer antigens often requires a highly personalized approach or relies on pre-existing anticancer T cells. It is also difficult to generate strong de novo T cell immunity in cancer patients, especially in the elderly. Personalized approaches rely on vaccines against tumor-associated antigens, neoantigens (i.e., mutated self-antigens) or viral oncoproteins. Other approaches are based on adoptive transfer of chimeric antigen receptor-transduced T cells or injection of monoclonal antibodies, which require difficult identification of tumor-specific antigens and are applicable only to a portion of cancer types or subtypes. And adoptive transfer of ex vivo expanded tumor-specific lymphocytes is a method that aims to take advantage of the natural antitumor response. All these approaches are highly personalized and require the identification of tumor epitopes and / or the expansion of the patient's autologous cells ex vivo.

[0005] In parallel, in situ tumor immunotherapies based on cytokines or TLR ligands have been used, many of which target innate immune recognition mechanisms to alter the tumor immune microenvironment, induce immunogenic cancer cell death, and promote epitope spreading.

[0006] Therefore, simple, broadly applicable, antigen-agnostic, immunotherapeutic methodologies remain needed to harness the effects of the immune system in early and long-term cancer control through direct cell death and enhanced epitope spreading, respectively. Summary of the Invention

[0007] overview The inventors recognized that the complex adaptive cell-mediated immunity that evolves over many years to potently control chronic viral infections in the elderly is a type of cell-mediated immunity that is effective in controlling tumor growth. To harness this type of antiviral immunity for cancer treatment, the inventors aimed to either directly target highly functional pre-existing antiviral T cells to the tumor environment using tumor-tropic papillomavirus pseudovirions or to induce minimal viral CD8 and CD4 T cell proliferation. We developed a novel approach to in situ immunotherapy by in situ injection of T cell cytomegalovirus (CMV) epitopes. Presentation of viral epitopes in the tumor environment leads to recruitment and activation of viral antigen-specific T cells in situ, which consequently differentially kill virus-negative tumor cells and alter the tumor microenvironment. This approach addresses an unmet need as it fulfills all the criteria for successful immunotherapy by promoting and establishing both early and long-term cancer cell killing and epitope spreading.

[0008] Thus, the present disclosure provides a method of treating cancer in an individual by recruiting a pre-existing immune response to the site of the cancer, thereby treating the cancer. The pre-existing immune response may be an immune memory response that is present in the individual before the individual is diagnosed with cancer. The pre-existing immune response may be a natural pre-existing immune response.

[0009] In these methods, recruiting a pre-existing immune response to the cancer cells may include introducing an antigen to the cancer that is not expressed by the cancer cells prior to the initiation of treatment, where the antigen is recognized by one or more components of the pre-existing immune response.

[0010] These methods may include confirming that the individual has a pre-existing immune response to the antigen before introducing the antigen into the tumor.These methods may also include evaluating the individual's pre-existing immune response to the antigen.In these methods, confirming the presence of a pre-existing immune response may include identifying the T cell response to the antigen in a sample from the individual.

[0011] In these methods, introducing the antigen may include injecting the antigen into the cancer. Additionally or alternatively, introducing the antigen may be accomplished by introducing a nucleic acid molecule encoding the antigen into the cancer. In these methods, the nucleic acid molecule may be DNA or RNA. When using RNA, the RNA may be modified to be more resistant to degradation. The nucleic acid molecule may be introduced into the cancer cells by injection. Additionally or alternatively, the nucleic acid molecule may be introduced into the cancer using a viral vector or pseudovirion such as a papillomavirus pseudovirion.

[0012] In these methods, the antigen can be a viral antigen. For example, the antigen can be a polypeptide that includes at least one epitope from a cytomegalovirus (CMV) protein, which peptide is recognized by one or more components of a pre-existing immune response. In these methods, the CMV protein can be selected from the group consisting of pp50, pp65, pp150, IE-1, IE-2, gB, US2, US6, UL16 and UL18. The polypeptide can be a 9-15 mer MHC I restricted peptide. Or Alternatively or additionally, the polypeptide is an at least 15-mer MHC II restricted peptide. Alternatively or additionally, the antigen may be a sequence selected from the sequences of SEQ ID NOs: 1-67. In these methods, one or more components of the immune response may be a T cell.

[0013] These methods may alter the cancer microenvironment by recruiting pre-existing immune responses.

[0014] In these methods, the antigen can be administered in combination with an agent that enhances the immune response. Exemplary agents include TLR agonists; IL-1R8 cytokine antagonists; intravenous immunoglobulin (IVIG); peptidoglycan isolated from gram-positive bacteria; lipoteichoic acid isolated from gram-positive bacteria; lipoproteins isolated from gram-positive bacteria; lipoarabinomannan isolated from mycobacteria; zymosan isolated from yeast cell walls; polyadenylic-polyuridylic acid; poly(IC); lipopolysaccharide; monophosphoryl lipid A; flagellin; Gardiquimod; imiquimod; R848; Included are agents selected from oligonucleosides containing CpG motifs, CD40 agonists, and 23S ribosomal RNA. In an exemplary method, the antigen may be administered in combination with Poly IC.

[0015] Another aspect provides kits for testing a patient and mobilizing a pre-existing immune response to a cancer site in the patient. These kits may include at least one CMV peptide antigen or a nucleic acid encoding the peptide, a pharma- ceutically acceptable carrier, a container, and a package insert or label indicating administration of the CMV peptide to reduce at least one symptom of cancer in a patient.

[0016] This summary is not intended to be, and should not be construed as, representative of the entire scope and scope of the invention. Moreover, references herein to "the disclosure" or aspects thereof should be understood to mean particular embodiments of the invention and should not necessarily be construed as limiting all embodiments to the particular description. The disclosure is described in this summary at various levels of detail, as well as in the accompanying figures and description of the embodiments, and no limitation as to the scope of the disclosure is intended by the inclusion or non-inclusion of elements, components, etc. in this summary. Further aspects of the invention may become readily apparent from the detailed description, particularly when taken in conjunction with the figures. [Brief description of the drawings]

[0017] [Figure 1-1] Figure 1A shows that murine cytomegalovirus (mCMV) infection induces a large cytokine response to a mCMV peptide pool. Figure 1B shows IFN-γ production by splenic CD4+ and CD8+ T cells following peptide restimulation with the indicated MHC-I and MHC-II restricted mCMV peptides. [Figure 1-2] Figure 1A shows that murine cytomegalovirus (mCMV) infection induces a large cytokine response to a mCMV peptide pool. Figure 1B shows IFN-γ production by splenic CD4+ and CD8+ T cells following peptide restimulation with the indicated MHC-I and MHC-II restricted mCMV peptides. [Diagram 2] Figure 2A shows the injection protocol for intratumoral induction of solid tumors with HPV Psv expressing mCMV antigens. Figures 2B and 2C show the tumor volume following intratumoral injection of HPV16 Psv expressing m122 and m45, or HPV Psv expressing red fluorescent protein (RFP), respectively. [Figure 3-1]Figure 3A shows an injection protocol for intratumoral transfer of solid tumors with HPV Psv expressing mCMV antigens combined with poly(I:C) (PIC). Figures 3B-3E show that this intratumoral transfer protocol slows tumor growth. Figures 3F and 3G show infiltration of tumors with E7-, m45-, and m122-specific CD8+ T cells analyzed by MHC-I tetramer staining and FACS. [Figure 3-2] Figure 3A shows an injection protocol for intratumoral transfer of solid tumors with HPV Psv expressing mCMV antigens combined with poly(I:C) (PIC). Figures 3B-3E show that this intratumoral transfer protocol slows tumor growth. Figures 3F and 3G show infiltration of tumors with E7-, m45-, and m122-specific CD8+ T cells analyzed by MHC-I tetramer staining and FACS. [Figure 4] FIG. 4A shows the effect on survival and FIG. 4B shows the effect on tumor growth following intratumoral injection of MCMV MHC-I restricted peptides in C57Bl / 6 mice infected with murine cytomegalovirus (mCMV). [Diagram 5] FIG. 5 shows the effect of intratumoral injection of different doses of murine cytomegalovirus (mCMV) MHC-I restricted peptide on tumor growth in C57Bl / 6 mice infected with mCMV. [Figure 6-1] Figures 6A and 6B show the effect of intratumoral injection of a combination of mCMV MHC-I and MHC-II restricted peptides on tumor growth in mCMV-infected C57Bl / 6 mice, and Figure 6C shows blood E7-, m45-, and m122-specific CD8+ T cell responses analyzed by FACS with MHC-I tetramers to each peptide, demonstrating that sequential intratumoral inoculation with mCMV CD4 and then CD8 epitopes preferentially induces antitumor immunity. [Figure 6-2]Figures 6A and 6B show the effect of intratumoral injection of a combination of mCMV MHC-I and MHC-II restricted peptides on tumor growth in mCMV-infected C57Bl / 6 mice, and Figure 6C shows blood E7-, m45-, and m122-specific CD8+ T cell responses analyzed by FACS with MHC-I tetramers to each peptide, demonstrating that sequential intratumoral inoculation with mCMV CD4 and then CD8 epitopes preferentially induces antitumor immunity. [Figure 6-3] Figures 6A and 6B show the effect of intratumoral injection of a combination of mCMV MHC-I and MHC-II restricted peptides on tumor growth in mCMV-infected C57Bl / 6 mice, and Figure 6C shows blood E7-, m45-, and m122-specific CD8+ T cell responses analyzed by FACS with MHC-I tetramers to each peptide, demonstrating that sequential intratumoral inoculation with mCMV CD4 and then CD8 epitopes preferentially induces antitumor immunity. [Figure 7] FIG. 7 shows the effect of complete clearance of the primary tumor on long-term protection against secondary tumor challenge. [Figure 8] FIG. 8 shows that mCMV infection induces inflated CD8+ T cell responses in C57BL / 6 mice.

[0018] [Figure 9-1] Figure 9A shows IFN-γ production by inflated and non-inflated CD8+ T cells and IFN-γ production by CD4+ T cells, and Figure 9B shows cytokine production by mCMV CD8+ T cells against an MHC-I restricted peptide pool. [Figure 9-2] Figure 9A shows IFN-γ production by inflated and non-inflated CD8+ T cells and IFN-γ production by CD4+ T cells, and Figure 9B shows cytokine production by mCMV CD8+ T cells against an MHC-I restricted peptide pool. [Figure 10-1]Figure 10A shows the experimental protocol timing of the mouse TC1 tumor model for intratumoral administration of mCMV peptides. Figures 10B and 10C show the distribution of mCMV-specific CD8+ T cells in tumor-bearing mice. Inflated (IE3; Figure 10B) and non-inflated (m45; Figure 10C) specific CD8+ T cells were detected by FACS using MHC-I tetramer staining. [Figure 10-2] Figure 10A shows the experimental protocol timing of the mouse TC1 tumor model for intratumoral administration of mCMV peptides. Figures 10B and 10C show the distribution of mCMV-specific CD8+ T cells in tumor-bearing mice. Inflated (IE3; Figure 10B) and non-inflated (m45; Figure 10C) specific CD8+ T cells were detected by FACS using MHC-I tetramer staining. [Figure 10-3] Figure 10A shows the experimental protocol timing of the mouse TC1 tumor model for intratumoral administration of mCMV peptides. Figures 10B and 10C show the distribution of mCMV-specific CD8+ T cells in tumor-bearing mice. Inflated (IE3; Figure 10B) and non-inflated (m45; Figure 10C) specific CD8+ T cells were detected by FACS using MHC-I tetramer staining. [Figure 11-1] Figure 11A shows the experimental protocol timing of the mouse TC1 tumor model used for gene expression analysis of the tumor microenvironment. Figures 11B-11F show tumor infiltration by CD45+ cells (Figure 11B), Th1 cells (Figure 11C), cytotoxic CD8 T cells (Figure 11D), NK cells (Figure 11E) or dendritic cells (Figure 11F) after intratumoral administration. [Figure 11-2] Figure 11A shows the experimental protocol timing of the mouse TC1 tumor model used for gene expression analysis of the tumor microenvironment. Figures 11B-11F show tumor infiltration by CD45+ cells (Figure 11B), Th1 cells (Figure 11C), cytotoxic CD8 T cells (Figure 11D), NK cells (Figure 11E) or dendritic cells (Figure 11F) after intratumoral administration. [Figure 11-3]Figure 11A shows the experimental protocol timing of the mouse TC1 tumor model used for gene expression analysis of the tumor microenvironment. Figures 11B-11F show tumor infiltration by CD45+ cells (Figure 11B), Th1 cells (Figure 11C), cytotoxic CD8 T cells (Figure 11D), NK cells (Figure 11E) or dendritic cells (Figure 11F) after intratumoral administration. [Figure 12] Figures 12A and 12B show that intratumoral administration of mCMV CD8 epitopes slows tumor growth and poly(I:C) co-injection improves tumor control. Figure 12A shows the effect of intratumoral injection of MHC-I restricted mCMV peptide alone + / - poly(I:C). Figure 12B shows the effect of intratumoral injection of titrations of MHC-I restricted mCMV peptide. [Figure 13] Figures 13A and 13B show protection from TC1 tumor challenge by intratumoral injection of mCMV MHC-I and / or MHC-II peptides bearing poly(I:C). Sequential intratumoral inoculation with CD4 and then CD8 MCMV epitopes suppresses tumor growth (Figure 13A) and promotes long-term survival (Figure 13B). [Figure 14] FIG. 14 shows E7 tetramer positive CD8+ T cell responses in blood after six treatments with MHC-I restricted selected m38, m45 and m122 peptides, and / or MHC-II restricted m139 selected peptide, with or without poly(I:C) (30 ug), as well as saline or poly(I:C) alone as controls. [Figure 15] FIG. 15 shows that complete clearance of the primary tumor confers long-term protection against secondary tumor challenge. [Figure 16] FIG. 16 shows protection from MC38 tumor challenge by intratumoral injection of mCMV MHC-I and MHC-II peptides with poly(I:C). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0019] Detailed Description The present invention relates to novel methods of treating cancer. Specifically, the present invention relates to methods of treating cancer in an individual that utilize the individual's own immune system to attack cancer cells. The methods of the present invention take advantage of the fact that an individual has a pre-existing immune response that was not induced in response to cancer, but was induced by microorganisms in the environment. Because cancer cells do not normally express the microbial antigens that induced the pre-existing immune response, one would not expect such an immune response to attack the cancer. However, the inventors have discovered that such a pre-existing immune response can be mobilized to attack the cancer. One way this can be accomplished is by introducing one or more antigens recognized by the pre-existing immune response into the cancer, resulting in the cells of the immune response attacking the cancer cells that present the antigen. Thus, these methods are not directed at cancer cells that express the antigen prior to treatment of the cancer patient. For example, many glioblastoma cancer cells have been found to express CMV antigens, and the methods of the present disclosure may be directed against CMV. The existing immunity of the individual is not used to treat such glioblastoma. Furthermore, the destruction of cancer cells may result in the components of the existing immune response being exposed to cancer cell antigens. This may lead to the induction of an immune response against cancer cell antigens. Thus, the general method of the present invention may be carried out by mobilizing the existing immune response in an individual to the cancer site so that the existing immune response attacks the cancer. Mobilization may be achieved, for example, by introducing at least one antigen that is recognized by the components of the existing immune response (e.g., T cells) of the individual to the cancer.

[0020] The present invention is not limited to the specific embodiments described herein, as such embodiments may vary, and the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.

[0021] As used herein and in the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise. For example, a nucleic acid molecule means one or more nucleic acid molecules. Thus, the terms "a," "an," "one or more," and "at least one" can be used interchangeably. Similarly, the terms "comprising," "including," and "having" can be used interchangeably. It is further noted that the claims may be drafted to exclude any element. As such, the specification is intended to serve as antecedent to the use of exclusive terms such as "solely," "only," and the like in connection with the recitation of claim elements or the use of a "negative" limitation.

[0022] Certain features of the invention that are described in the context of separate embodiments may also be provided in combination in a single embodiment. Conversely, various features of the invention that are briefly described in the context of a single embodiment may also be provided separately or in any suitable subcombination. All combinations of embodiments are specifically embraced by the invention and are described herein as if each combination were individually and expressly disclosed. Moreover, all subcombinations are also specifically embraced by the invention and are described herein as if each such subcombination were individually and expressly disclosed.

[0023] The documents discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein should be construed as an admission that the present invention is not entitled to antedate such documents by virtue of prior invention. Further, the publication dates provided may be different from the actual publication dates, which may need to be independently confirmed. All documents discussed herein are incorporated by reference herein to disclose and describe the methods and / or materials in connection with which the documents are cited.

[0024] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention, the preferred methods and materials are described herein.

[0025] One aspect is a method of treating cancer in an individual, comprising recruiting a pre-existing immune response to the cancer, thereby treating the cancer.

[0026] As used herein, cancer refers to a disease in which abnormal cells divide without proper control of cell division and / or cellular senescence. The term cancer is meant to encompass not only solid tumors, but also blood borne cancers. Generally, tumors are masses of abnormal tissue that usually do not contain cysts or fluid areas. Solid tumors include benign (non-life threatening), Solid tumors can be noncancerous (non-cancerous) or malignant (life-threatening). Different types of solid tumors are named according to the type of cell that forms them. Examples of solid tumors include sarcomas, carcinomas, and lymphomas. Hematologic tumors (also called cancers of the blood) are cancers that begin in blood-forming tissues, such as the bone marrow, or in the cells of the immune system. Examples of cancers of the blood include leukemia, lymphoma, and multiple myeloma.

[0027] In some cancers, cells may invade tissues other than the tissue in which the original cancer cells originated. In some cancers, cancer cells may metastasize to other parts of the body via the blood and lymphatic system. Thus, cancers are usually named according to the organ or type of cell from which they originate. For example, cancer originating from the colon is called colon cancer, cancer originating from melanocytes in the skin is called melanoma, and so on. As used herein, cancer includes carcinoma, sarcoma, adenocarcinoma, lymphoma, leukemia including solid tumors and cancers of the lymphatic system, stomach cancer, kidney cancer, breast cancer, lung cancer (including non-small cell lung cancer and small cell lung cancer), bladder cancer, colon cancer, ovarian cancer, prostate cancer, pancreatic cancer, stomach cancer, brain cancer, head and neck cancer, skin cancer, uterine cancer, testicular cancer, esophageal cancer, liver cancer (including hepatocellular carcinoma), non-homologous cancer, and ovarian cancer. It may refer to lymphomas, including Hodgkin's lymphoma (e.g., Burkitt's lymphoma, small cell lymphoma, and large cell lymphoma) and Hodgkin's lymphoma, as well as multiple myeloma. In an exemplary embodiment, the cancer is lung cancer or adenocarcinoma.

[0028] The terms individual, subject, patient, etc., as used herein, are meant to encompass any mammal that may develop cancer, with the preferred mammal being a human. The terms individual, subject, and patient do not themselves denote a particular age, sex, race, etc. Thus, individuals of any age, whether male or female, are intended to be covered by the present disclosure. Similarly, the methods of the present invention can be applied to humans of any race, including, for example, Caucasian (white), African American (black), Native American, Native Hawaiian, Hispanic, Latino, Asian, and European. Such characteristics may be important. In such cases, significant characteristics (e.g., age, sex, race, etc.) may be indicated. These terms also encompass both humans and non-human animals. Non-human animals suitable for cancer testing or treatment include, but are not limited to, companion animals (i.e., pets), food animals, working animals, or zoo animals.

[0029] As used herein, immune response or immunological response refers to the presence in an individual of a humoral and / or cellular response to one or more antigens. For the purposes of this disclosure, a "humoral response" refers to an immune response mediated by B cells and antibody molecules, including secretory (IgA) or IgG molecules, while a "cellular response" refers to a response mediated by T lymphocytes and / or other white blood cells. One important aspect of cellular immunity involves antigen-specific responses by cytolytic T cells (CTLs). CTLs have specificity for peptide antigens presented in association with proteins encoded by major histocompatibility complexes (MHC) on the cell surface. CTLs help induce and promote the destruction of intracellular microbes, or the lysis of cells infected with such microbes. Another aspect of cellular immunity involves antigen-specific responses by helper T cells. Helper T cells act to help stimulate the function and focus the activity of nonspecific effector cells against cells presenting peptide antigens in association with MHC molecules on their surface. A cellular immune response also refers to the production of cytokines, chemokines and other such molecules produced by activated T cells and / or other white blood cells, including those derived from CD4+ and CD8+ T cells.

[0030] Thus, the immunological response may be one that stimulates CTLs and / or the production or activation of helper T cells. Production of chemokines and / or cytokines may also be stimulated. The immune response may also include an antibody-mediated immune response. Thus, the immune response may include one or more of the following effects: the production of antibodies (e.g., IgA or IgG) by B cells; and / or activation of suppressor, cytotoxic or helper T cells, and / or activation of T cells specifically directed against the antigen. Such responses can be determined using standard immunoassays and neutralization assays known in the art.

[0031] As used herein, a pre-existing immune response is an immune response that exists in an individual before the start of cancer treatment. Thus, an individual with a pre-existing immune response has an immune response to an antigen prior to the start of treatment with the antigen to treat cancer. A pre-existing immune response may be a natural immune response or an induced immune response. As used herein, a natural pre-existing immune response is an immune response of an individual that is induced in response to an antigen, such as a bacterial or viral antigen, that the individual unintentionally comes into contact with. That is, an individual with a pre-existing immune response is not exposed to an antigen with the intention of generating an immune response to the antigen. An induced pre-existing immune response is an immune response that occurs as a result of intentional exposure to an antigen, such as when receiving a vaccine. A pre-existing immune response may be a natural immune response or an induced immune response.

[0032] As used herein, the phrase "mobilizing an immune response" refers to the process by which an antigen is administered to an individual such that components of a pre-existing immune response migrate through the body to the site where the antigen was administered, resulting in an attack by components of the immune system against cells presenting the antigen. As used herein, "components of the immune response" refers to cells that can bind to an antigen and initiate an immune response against the antigen. Antigens useful in the practice of the present invention are any molecules that can be recognized by cells of the pre-existing immune system, particularly T cells. One example of such a compound is a protein, such as a bacterial or viral protein.

[0033] As used herein, the phrase "treating cancer" refers to various outcomes related to cancer. Treating cancer includes reducing the rate of increase in the number of cancer cells in the treated individual. Such a reduction in the rate of increase may be due to a decrease in the rate of replication of cancer cells. Alternatively, the rate of replication of cancer cells is not affected, and the increasing number of cancer cells may be killed by a pre-existing immune response. In certain aspects, treating cancer refers to a situation in which the increase in the number of cancer cells stops, but remains at a certain level. Such a situation occurs either due to inhibition of the replication of cancer cells by the recruitment of a pre-existing immune response, or due to a balance between the rate of killing of cancer cells by the recruited pre-existing immune response and the rate of production of new cancer cells. Treating cancer refers to stabilizing the cancer so that the cancer growth is reduced or stopped, or to reducing the number of cancer cells in the treated individual, and / or to preventing the individual from developing cancer (i.e., having no detectable cancer cells).

[0034] In an embodiment, the step of mobilizing a pre-existing immune response comprises introducing an antigen to the cancer that is recognized by one or more components of the pre-existing immune response. In a preferred embodiment, the antigen is not present in the cancer prior to treatment. Thus, one embodiment is a method of treating cancer in an individual, comprising mobilizing a pre-existing immune response to the cancer by introducing an antigen to the cancer that is recognized by one or more components of the pre-existing immune response, wherein the antigen is not present in the cancer prior to treating the cancer. Thus, as noted above, the pre-existing immune response may be a natural or induced immune response. Introduction of an antigen to the cancer can be accomplished using methods known in the art and may vary depending on the type of cancer being treated. For example, one type of cancer is a solid tumor. In such cancers, cancer cells replicate and remain adjacent to parent cancer cells, resulting in the formation of a mass of tissue formed from adjacent cancer cells. Because such cancers are a mass of cells, the antigen can be delivered directly to or within the mass. In one embodiment, a method of treating cancer in an individual, comprising mobilizing a pre-existing immune response to the solid tumor, wherein if the cancer is a solid tumor, the method comprises mobilizing a pre-existing immune response to the solid tumor, wherein .... The present invention includes recruiting a pre-existing immune response to a solid tumor by introducing an antigen to the solid tumor that is recognized by one or more components of the pre-existing immune response, where the antigen is not present in the solid tumor prior to treatment. In one embodiment, the pre-existing immune response is a natural immune response. In one embodiment, the pre-existing immune response is an induced immune response. In one embodiment, the antigen is delivered to the cancer (e.g., solid tumor) by injection of the antigen into the cancer (e.g., solid tumor). In such an embodiment, the antigen is displayed on MHC I molecules of cells, either by the antigen directly binding to such molecules or by uptake and processing of the antigen by the cancer cells. In these methods, the antigen can be combined with other molecules or compounds that enhance uptake and / or presentation of the antigen to the immune system.

[0035] As described above, in these methods, the antigen may be a protein. These protein antigens may be directly injected into the cancer (e.g., tumor) as described above. Thus, one aspect is a method of treating cancer in an individual, where the cancer is a solid tumor, comprising mobilizing a pre-existing immune response to the solid tumor by injecting an antigenic protein into the solid tumor, where the antigenic protein is recognized by one or more components of the pre-existing immune response, where the antigenic protein is not present in the solid tumor prior to treatment. Alternatively, the protein antigen can be introduced into the cancer by introducing a nucleic acid molecule encoding the protein into the cancer. Thus, one aspect is a method of treating cancer in an individual, where the cancer is a solid tumor, comprising mobilizing a pre-existing immune response to the solid tumor by introducing a nucleic acid molecule encoding an antigenic protein into the solid tumor, where the antigenic protein is recognized by one or more components of the pre-existing immune response, where the antigenic protein is not present in the solid tumor prior to treatment. The introduction of the nucleic acid molecule encoding the antigen into the cancer can be performed using any suitable method known in the art. In one aspect, a method of treating cancer in an individual, where the cancer is a solid tumor, comprises recruiting a pre-existing immune response to the solid tumor by injecting into the solid tumor a nucleic acid molecule encoding an antigenic protein, where the antigenic protein is recognized by one or more components of the pre-existing immune response, where the antigenic protein is not present in the solid tumor prior to treatment. In these methods, the nucleic acid molecule encoding the antigen is a naked nucleic acid molecule (i.e., a nucleic acid molecule that is capable of being vaccinated). The nucleic acid molecule encoding the antigen may be injected as is (uncomplexed with other molecules intended to enhance delivery of the nucleic acid molecule), or the injected nucleic acid molecule may be complexed with one or more compounds intended to enhance delivery, stability or longevity of the nucleic acid molecule. Examples of such compounds include lipids, proteins, carbohydrates and polymers, including synthetic polymers.

[0036] Nucleic acid molecules encoding two or more antigens can also be introduced into cancer using a delivery vehicle such as a recombinant virus or pseudovirion. Examples of viruses useful for carrying out the method of the present invention include, but are not limited to, adenovirus, adeno-associated virus, herpes virus and papilloma virus. The use of such viruses to deliver nucleic acid molecules is known to those skilled in the art and is also disclosed in U.S. Patent No. 8,394,411, which is incorporated herein by reference. Examples of pseudoviruses useful for carrying out the method of the present invention include, but are not limited to, hepatitis pseudovirus, influenza pseudovirus, and papilloma pseudovirus. As used herein, pseudovirus refers to a particle consisting of viral capsid proteins assembled into a virus-like particle (VLP) that can bind and enter cancer cells. Such pseudovirion particles can, but preferably do not, package subgenomic amounts of viral nucleic acid molecules. Methods for making and using pseudovirions are known in the art and are also described in U.S. Pat. Nos. 6,599,739; 7,205,126; and 6,416,945, all of which are incorporated herein by reference in their entireties. Thus, the present disclosure provides a method of treating cancer in an individual, where the cancer is a solid tumor, comprising recruiting a pre-existing immune response to the solid tumor by introducing into the tumor a recombinant virus or pseudovirus comprised of a nucleic acid molecule encoding an antigenic protein, the antigenic protein being recognized by one or more components of the pre-existing immune response, the antigenic protein not being present in the solid tumor prior to treatment. Entry of the pseudovirus carrying the nucleic acid molecule of the present disclosure into a cell results in expression of the encoded antigenic protein by the cell and subsequent presentation of the antigen to the immune system. In these methods, the pseudovirus is a papilloma pseudovirus.

[0037] The introduction of the virus or pseudovirus comprising the nucleic acid molecule encoding the antigen into the cancer can be achieved by any suitable method known in the art. For example, the recombinant virus or pseudovirus comprising the nucleic acid molecule encoding the antigen can be injected near the cancer or directly into the cancer. Alternatively, the recombinant virus or pseudovirus comprising the nucleic acid molecule encoding the antigen can be administered to an individual by a route that results in the recombinant virus or pseudovirus being delivered to the cancer. Examples of such routes include, but are not limited to, intravenous (IV) injection, intramuscular (IM) injection, intraperitoneal (IP) injection, subcutaneous (SC) injection and oral delivery. Thus, one embodiment is a method of treating cancer in an individual, comprising administering to the individual a recombinant virus or pseudovirus comprising the nucleic acid molecule encoding an antigenic protein, wherein the cancer is a solid tumor, the antigenic protein is recognized by one or more components of the existing immune response, and the antigenic protein is not present in the solid tumor prior to treatment. In these methods, the recombinant virus or pseudovirus may be injected directly into the solid tumor, or the recombinant virus or pseudovirus may be delivered using a method selected from IV injection, IM injection, IP injection, SC injection, and oral delivery.

[0038] The disclosed method may be used to treat blood cancers. Blood cancers, blood cancers, blood tumors, etc., begin in blood-forming tissues, such as bone marrow, or cells of the immune system. Examples of blood cancers include leukemia, lymphoma, multiple myeloma, etc. Such cancers begin when cells of blood-forming tissues, or cells of the immune system, lose control of cell replication and begin replicating in an uncontrolled manner. Once formed, blood cancer cells find their way into the blood or lymphatic system, causing a significant increase in the number of cancer cells in the blood and / or lymphatic system. For example, leukemia is a cancer found in the blood and bone marrow. Leukemia results from uncontrolled replication of white blood cells, resulting in a large increase in the number of abnormal white blood cells in the blood and lymphatic tissues. These abnormal white blood cells do not function normally, and thus individuals with leukemia are unable to fight infections. Thus, the present disclosure provides a method of treating a hematological tumor in an individual, comprising recruiting a pre-existing immune response to the cancer cells of the hematological tumor in the individual by introducing an antigen recognized by one or more components of the pre-existing immune response to the cancer cells of the hematological tumor, wherein the antigen is not present in or present on the cancer cells of the hematological tumor prior to treatment. In these methods, the pre-existing immune response may be a natural immune response or an induced immune response. The introduction of an antigen to the hematological tumor cells can be performed using any suitable method. In these methods, the antigen can be introduced to the cancer cells of the hematological tumor by administering the antigen to the individual in a manner that results in delivery of the antigen to the cancer cells of the hematological tumor. For example, the antigen can be administered to the individual using a method selected from IV injection, IM injection, IP injection, SC injection, and oral administration. In these methods, the antigen can be targeted to the cancer cells of the hematological tumor, for example, by binding the antigen to a protein that binds a molecule on the cancer cells of the hematological tumor.

[0039] Antigens can also be introduced into cancer cells of a hematological tumor of an individual by introducing a nucleic acid molecule encoding an antigenic protein into the cancer cells of the hematological tumor. The present invention provides a method for treating a hematological tumor in an individual, comprising mobilizing an existing immune response to cancer cells of the hematological tumor by administering to the individual a nucleic acid molecule encoding an antigenic protein, wherein the antigenic protein is recognized by one or more components of the existing immune response, and the antigenic protein is not present in or on the cancer cells of the hematological tumor prior to treatment. The administration of the nucleic acid molecule encoding the antigen to the individual can be performed using any suitable method known in the art. For example, the nucleic acid molecule encoding the antigen can be injected as a naked nucleic acid molecule. Alternatively or additionally, the nucleic acid molecule encoding the antigen can be complexed with one or more compounds intended to improve the delivery, stability or longevity of the nucleic acid molecule. Examples of such compounds include lipids, proteins, carbohydrates and polymers, including synthetic polymers.

[0040] The nucleic acid molecule encoding two or more antigens can also be introduced into blood cancer cells using a delivery vehicle such as a recombinant virus or pseudovirus. Examples of such delivery vehicles are as described herein above. Examples of viruses useful for carrying out the method of the present invention include, but are not limited to, adenovirus, adeno-associated virus, herpes virus and papilloma virus. Examples of pseudoviruses useful for carrying out the method of the present invention include, but are not limited to, hepatitis pseudovirus, influenza pseudovirus and papilloma pseudovirus. Thus, the present disclosure provides a method of treating blood cancer in an individual, comprising mobilizing a pre-existing immune response to a solid tumor by introducing a recombinant virus or pseudovirus comprising a nucleic acid molecule encoding an antigenic protein into the tumor, wherein the antigenic protein is recognized by one or more components of the pre-existing immune response, and the antigenic protein is not present in or on the blood cancer cells prior to treatment.

[0041] Introduction of a virus or pseudovirus comprising a nucleic acid molecule encoding an antigen into a cancer can be accomplished using any suitable method known in the art. For example, a recombinant virus or pseudovirus comprising a nucleic acid molecule encoding an antigen can be administered to an individual by a route that results in the recombinant virus or pseudovirus being delivered to the cancer. Examples of such routes include, but are not limited to, intravenous (IV) injection, intramuscular (IM) injection, intraperitoneal (IP) injection, subcutaneous (SC) injection, and oral administration. Thus, the present disclosure provides a method of treating a blood cancer in an individual, comprising administering to the individual a recombinant virus or pseudovirus comprising a nucleic acid molecule encoding an antigenic protein, wherein the antigenic protein is recognized by one or more components of a pre-existing immune response, and the antigenic protein is not present in or on blood cancer cells prior to treatment. The recombinant virus or pseudovirus can be delivered using a method selected from the group consisting of IV injection, IM injection, IP injection, SC injection, and oral administration.

[0042] The methods described herein use one or more antigens to recruit an existing immune response to cancer. Any antigen can be used as long as it is recognized by one or more components of the existing immune response and the antigen is not present in or on the cancer cell prior to treatment. Examples of useful antigens include, but are not limited to, viral and bacterial antigens. One example of a viral antigen useful for carrying out the methods of the present invention is an antigen that contains at least one epitope from a cytomegalovirus protein. As used herein, an epitope is a cluster of amino acid residues that is recognized by the immune system, thereby inducing an immune response. Such an epitope may be composed of contiguous amino acid residues (i.e., amino acid residues that are adjacent to each other in the protein) or may be composed of non-contiguous amino acid residues (i.e., amino acid residues that are not adjacent to each other in the protein), but are in close special proximity in the final folded protein. In order for an epitope to be recognized by the immune system, it must contain at least six amino acid residues. It is generally understood by those skilled in the art that a cytomegalovirus residue is required. Thus, the methods of the present invention may include the use of an antigen that includes at least one epitope from a cytomegalovirus protein. Any suitable CMV protein may be used to generate an antigen useful for carrying out the methods of the present invention, so long as the antigen mobilizes a pre-existing immune response to cancer. Examples of CMV proteins suitable for use in the methods described herein include CMV pp50, CMV pp65, CMV pp150, CMV IE-1, CMV pp20, CMV pp10, CMV pp11, CMV pp21, CMV pp12, CMV pp13, CMV pp14, CMV pp15, CMV pp22, CMV pp23, CMV pp24, CMV pp25, CMV pp16, CMV pp17, CMV pp18, CMV pp19, CMV pp26, CMV pp27, CMV pp28, CMV pp29 ... Examples of such proteins and useful fragments thereof include, but are not limited to, IE-2, CMV gB, CMV US2, CMV UL16 and CMV UL18. Examples of such proteins and useful fragments thereof are disclosed in U.S. Patent Publication Nos. 2005 / 0019344 and 2010 / 0183647, both of which are incorporated herein by reference in their entirety. Useful fragments may also include any one or combination of peptides comprising the amino acid sequence of SEQ ID NOs: 1-67.

[0043] The disclosed methods can also be practiced with one or more antigens, each of which independently comprises an amino acid sequence that is a variant of at least 8 consecutive amino acids from a CMV protein. As used herein, variant refers to a protein or nucleic acid molecule having a sequence similar, but not identical, to a reference sequence, such that the activity (e.g., immunogenicity) of the variant protein (or protein encoded by the variant nucleic acid molecule) is not significantly altered. These sequence variations may be natural mutations or may be engineered using genetic engineering techniques known to those of skill in the art. Examples of such techniques are found in Sambrook J, Fritsch EF, Maniatis T et al., in Molecular Cloning-A Laboratory Manual, 2nd Edition, Cold Spring Harbor Laboratory Press, 1989, pp. 9.31-9.57, and in J. Immunol. 1999, 10, 111-115, 1999, 10, 112-114, 1999, 10, 113-115, 1999, 10, 114-115, 1999, 10, 115-116, 1999, 10, 116-117, 1999, 10, 117-118, 1999, 10, 118-119, 1999, 10, 119-120 ... can be found in Current Protocols in Molecular Biology, John Wiley & Sons, NY (1989), 6.3.1-6.3.6.

[0044] With respect to variants, any type of change in amino acid sequence is acceptable, so long as the resulting variant protein retains the ability to induce immune response. Examples of such variants include, but are not limited to, deletion, insertion, substitution, and combinations thereof. For example, it is well understood by those skilled in the art that in a protein, one or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acids can often be removed from the amino and / or carboxy termini of the protein without significantly affecting the activity of the protein. Similarly, one or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acids can often be inserted into a protein without significantly affecting the activity of the protein.

[0045] As described, mutant proteins may contain amino acid substitutions relative to control proteins (e.g., wild-type proteins). Any amino acid substitution is acceptable as long as the activity of the protein is not significantly affected. In this regard, it is understood in the art that amino acids can be classified based on their physical properties. Examples of such groups include, but are not limited to, charged amino acids, uncharged amino acids, polar uncharged amino acids, and hydrophobic amino acids. Preferred mutants that contain substitutions are those in which an amino acid is replaced with an amino acid from the same group. Such substitutions are called conservative substitutions.

[0046] Natural residues can be divided into classes based on common side chain properties: 1) hydrophobic: Met, Ala, Val, Leu, Ile; 2) neutral hydrophilic: Cys, Ser, Thr; 3) acidic: Asp, Glu; 4) basic: Asn, Gln, His, Lys, Arg; 5) residues that influence chain orientation: Gly, Pro; and 6) aromatic: Trp, Tyr, Phe.

[0047] For example, non-conservative substitutions substitute a member of one of these classes for a member of another class. This may include exchanging

[0048] When making amino acid mutations, the hydrophobicity index of the amino acid can be taken into consideration. Each amino acid is assigned a hydrophilicity index based on its hydrophobic and charge characteristics. The hydrophilicity indices are: isoleucine (+4.5); valine (+4.2); leucine (+3.8); phenylalanine (+2.8); cysteine / cystine (+2.5); methionine (+1.9); alanine (+1.8); glycine (-0.4); threonine (-0.7); serine (-0.7); serine (-0.8); tryptophan (-0.9); tyrosine (-1.3); proline (-1.6); histidine (-3.2); glutamic acid (-3.5); glutamine (-3.5); asparagine (-3.5); lysine (-3.9); and arginine (-4.5). The importance of the hydrophobic amino acid index in conferring interactive biological function on a protein is generally understood in the art (Kyte et al., 1982, J. Mol. Biol. 157:105-31). Certain amino acids have similar hydrophobicity indices or scores. It is known that amino acids having a hydrophilicity index of 1 to 5 are preferably substituted with other amino acids having a hydrophilicity index of 1 to 5 and still retain similar biological activity. When making modifications based on hydrophilicity index, substitution of amino acids having a hydrophilicity index of 1 to 5 is preferred, those having a hydrophilicity index of 1 to 5 are particularly preferred, and those having a hydrophilicity index of 0.5 to 5 are even more preferred.

[0049] It is understood in the art that substitutions of like amino acids can be made effectively on the basis of hydrophilicity, particularly when the biologically functional equivalent proteins or peptides produced thereby are intended for use in immunological inventions, as is the case herein. The greatest local average hydrophilicity of a protein, as governed by the hydrophilicity of its adjacent amino acids, correlates with its immunogenicity and antigenicity, i.e., with a biological property of the protein. The following hydrophilicity values ​​have been assigned to these amino acid residues: arginine (+3.0); lysine (+3.0); aspartic acid (+3.0±1); glutamic acid (+3.0±1); serine (+0.3); asparagine (+0.2); glutamine (+0.2); glycine (0); threonine (-0.4); proline (-0.5±1); alanine (-0.5); histidine (-0.5); cysteine ​​(-1.0); methionine (-1.3); valine (-1.5); leucine (-1.8); isoleucine (-1.8); tyrosine (-2.3); phenylalanine (-2.5); and tryptophan (-3.4). When making changes based on similar hydrophilicity values, substitutions of amino acids within ±2 of the hydrophilicity value are preferred, with those within ±1 being particularly preferred, and those within ±0.5 being even more preferred. Epitopes may also be identified from primary amino acid sequences on the basis of hydrophilicity.

[0050] Desired amino acid substitutions (whether conservative or non-conservative) can be determined by those skilled in the art at the time such substitutions are desired.For example, amino acid substitutions can be used to identify important residues of a protein, or to increase or decrease the immunogenicity, solubility or stability of a protein.Exemplary amino acid substitutions are shown in the following table.

[0051] [Table 1]

[0052] As used herein, the phrase "significantly affecting the activity of a protein" means a decrease in the activity of the protein by at least 10%, at least 20%, at least 30%, at least 40%, or at least 50%. In the context of the present invention, such activity can be measured, for example, as the ability of the protein to induce neutralizing antibodies or to induce a T cell response. Methods for determining such activity are known to those skilled in the art.

[0053] The disclosed methods may use one or more antigens, each independently consisting of at least 6 contiguous amino acids, at least 10 contiguous amino acids, at least 20 contiguous amino acids, at least 30 contiguous amino acids, at least 50 contiguous amino acids, at least 75 contiguous amino acids, at least 75 contiguous amino acids, or at least 100 contiguous amino acids from a CMV protein. The disclosed methods may use one or more antigens, each independently having an amino acid sequence at least 85% identical, at least 95% identical, at least 97% identical, or at least 99% identical to at least 10 contiguous amino acids, at least 20 contiguous amino acids, at least 30 contiguous amino acids, at least 50 contiguous amino acids, at least 75 contiguous amino acids, or at least 100 contiguous amino acids from a CMV protein. The disclosed methods may use one or more antigens, each independently comprising at least 6 contiguous amino acids, at least 10 contiguous amino acids, at least 20 contiguous amino acids, at least 30 contiguous amino acids, at least 50 contiguous amino acids, at least 75 contiguous amino acids, or at least 100 contiguous amino acids from a CMV protein. The disclosed method further comprises: for 9 to 15 consecutive amino acid residues from the CMV protein that is the I-restricting antigen, One or more antigens may be used, each of which independently comprises an amino acid sequence at least 95% identical, at least 97% identical, or at least 99% identical. The disclosed methods include those in which the antigen comprises 9 to 15 contiguous amino acid residues from a CMV protein that is an MHC I-restricted antigen. The disclosed method may use one or more antigens each independently comprising a group. At least 15 consecutive amino acid residues from a CMV protein that is a II restriction antigen The disclosed method can be used in which the antigen comprises one or more antigens that contain an amino acid sequence that is at least 95% identical, at least 97% identical, or at least 99% identical to a CMV protein that is an MHC II restricted antigen. The disclosed method may use one or more antigens comprising a group. The disclosed method may use one or more antigens comprising an amino acid sequence at least 95% identical, at least 97% identical, or at least 99% identical to a peptide comprising a sequence selected from the group consisting of peptides comprising the amino acid sequence of SEQ ID NO: 1-67, or any combination thereof. The disclosed method may use one or more antigens comprising an amino acid sequence at least 95% identical, at least 97% identical, or at least 99% identical to ...

[0054] [Table 2] [Table 3]

[0055] The methods of the present invention include treating cancer in an individual by mobilizing a pre-existing immune response against cancer. In these methods, the individual may be known to have a pre-existing immune response against an antigen before starting cancer treatment. The individual may be tested to confirm the presence of a pre-existing immune response before starting cancer treatment. Thus, these methods include treating cancer in an individual by confirming that the individual has a pre-existing immune response against an antigen, where the antigen is not present in or on the cancer. The antigen is then administered to the individual confirmed to have pre-existing immunity so that the antigen is introduced into the cancer, thereby treating the cancer.

[0056] Such methods may be used to treat any of the cancers previously described herein, including any solid tumor and / or hematological cancer.

[0057] Any method that identifies that the individual to be treated has a pre-existing immune response to an antigen can be used to practice the methods of the present disclosure. Examples of such methods include identifying, in a sample from the individual, B cells that recognize a particular antigen, antibodies that recognize a particular antigen, T cells that recognize a particular antigen, or T cell activity initiated in response to a particular antigen. Any suitable sample from the individual can be used to identify a pre-existing immune response. Examples of suitable samples include, but are not limited to, whole blood, serum, plasma, and tissue samples. As used herein, recognition of a specific antigen by a B cell, T cell, or antibody refers to the ability of such a B cell, T cell, or antibody to specifically bind an antigen. Specific binding of an antigen by a B cell, T cell, or antibody refers to the binding of a B cell, T cell, or antibody to a specific antigen with a higher affinity than the binding affinity of the same B cell, T cell, or antibody to a molecule unrelated to the antigen. For example, B cells that recognize or are specific for an antigen of the CMV pp50 protein. In one embodiment, the T cells or antibodies are directed against a protein unrelated to the CMV pp50 protein, e.g., human It binds the CMV pp50 antigen with an affinity that is significantly greater than the binding affinity of the same B cell, T cell, or antibody to albumin. Specific binding between the two entities is determined by their dissociation constants It can be scientifically expressed as approximately 10 -6 Less than, about 10 -7 Less than or about 10 -8 The specific binding between molecules, between cells and between molecules, and the methods for measuring such binding are well known to those skilled in the art, including, but not limited to, enzyme immunoassays (e.g., ELISA), immunoprecipitation, immunoblot assays and other immunoassays, for example, as described in Sambrook et al., supra, and Harlow et al., Antibodies, a Laboratory Manual (Cold Spring Harbor Labs Press, 1988). Such methods are also described in U.S. Patent No. 7,172,873, which is incorporated herein by reference. Methods for measuring T cell activation in samples from individuals are also known to those skilled in the art. Examples of such methods are disclosed in U.S. Patent Publication No. 2003 / 003485 and U.S. Patent No. 5,750,356, both of which are incorporated herein by reference.

[0058] Such methods generally include contacting a T cell-containing sample from an individual with an antigen and measuring the sample for T cell activation.Methods for measuring T cell activation are also well known in the art and are disclosed in Walker, S., et al., Transplant Infectious Disease, 2007:9:165-70; and Kotton, CN et al. (2013) Transplantation 96, 333.

[0059] Commercially available CMV tests (QuantiFERON (商標) -CMV, QIAGEN Sciences Inc. Germantown, MD) was used to stimulate cells in heparinized whole blood by binding to the human cytomegalovirus protein. It is available as an in vitro diagnostic test using a peptide cocktail that stimulates the antigen (CMV). Patients exposed to a disease / infection have specific T cell lymphocytes in their blood that have immunological memory for the antigen (immunologically reactive molecule) that primes the disease / infection. The addition of antigen to blood drawn from an induced individual results in rapid restimulation of antigen-specific effector T cells, resulting in the release of cytokines (e.g., IFN-γ). Effector T cells can respond rapidly when exposed to a priming antigen. Thus, the production of IFN-γ in response to antigen exposure is a specific marker of the cellular immune response to that antigen. This IFN-γ response can be used to quantify the immune response. Detection of interferon-γ (IFN-γ) by enzyme-linked immunosorbent assay (ELISA) is used to identify in vitro responses to peptide antigens associated with CMV infection. QuantiFERON (商標) - The intended use of CMV is in humans. The aim is to monitor the level of anti-CMV immunity.

[0060] Thus, in any of the disclosed methods for treating cancer in an individual, the individual may first be identified as having a pre-existing immune response to an antigen not present in or on the cancer. This pre-existing immune response may be identified by identifying in a sample from the individual: i) B cells that recognize a particular antigen; ii) antibodies that recognize specific antigens; iii) T cells that recognize a specific antigen; and iv) T cell activity initiated in response to a specific antigen. The specific antigen can then be administered to an individual identified as having a pre-existing immune response, such that the antigen is introduced into the cancer, thereby treating the cancer.

[0061] In any of the methods provided in this disclosure, other agents may be used (i.e., administered) in combination with the CMV antigen to enhance immune regulation or recruitment within the scope of the present invention. Such other agents include TLR agonists; intravenous immunoglobulin (IVIG); peptidoglycan isolated from gram-positive bacteria; lipotecholic acid isolated from gram-positive bacteria; lipoproteins isolated from gram-positive bacteria; lipoarabinomannan isolated from mycobacteria; zymosan isolated from yeast cell walls; polyadenylic-polyuridylic acid; poly(IC); lipopolysaccharide; monophosphoryl lipid A; flagellin; gardikimod; imiquimod; R848; oligonucleosides containing CpG motifs, CD40 agonists, and 23S ribosomal RNA. In a preferred aspect of these methods, the TLR agonist is poly-IC.

[0062] Another aspect of the present disclosure is a kit for testing an individual and mobilizing a pre-existing immune response against cancer in the individual. The kit may include at least one CMV peptide antigen or a nucleic acid encoding a peptide thereof, a pharma- ceutically acceptable carrier, a container, and a package insert or label indicating administration of the CMV peptide to reduce at least one symptom of cancer in a patient. These kits may further include a means for testing the antigenic response of the patient to the CMV antigen. For example, the kit may include sterile plasticware for obtaining and testing a whole blood sample, and an in vitro test of the response to the CMV peptide antigen, and / or detection of interferon-gamma (IFN-gamma) by enzyme-linked immunosorbent assay (ELISA) to identify the in vitro response to these peptide antigens. EXAMPLES

[0063] Working Example Chronic viral infections that are normally well controlled by the host, such as human cytomegalovirus (hCMV), often result in the induction of increasingly large numbers of fully functional virus-specific T cells with age. Using a murine mCMV model that mimics key aspects of the human immune response to hCMV, we have developed methods and reagents to attract these antiviral T cells to tumors, followed by tumor cell killing and inducing the spreading of potent epitopes to tumor neoantigens, resulting in an adaptive immune response that confers long-term control of tumor growth and protection from re-exposure to allogeneic tumor cells.

[0064] Example 1 Murine cytomegalovirus infection induces cytokine responses to mCMV peptide pools 1x10 to C57Bl / 6 mice 4 pfu of murine cytomegalovirus (mCMV ) were infected. Blood samples were taken 12 days after infection. Leukocytes were cultured using a selection of m38, m45, m57, m122, m139, m141 and m164 mCMV proteins. The CD8+ T cells were stimulated with the immunogenic peptides. IL-α and IL-2 cytokine production was assessed by intracellular cytokine staining and analyzed by fluorescence-activated cell sorting (FACS) (Figure 1A). Blood samples were collected 2 months after infection. Inflationary (m122) and non-inflationary (m45) specific CD8+ T cells were isolated by immunofluorescence staining with MHC-I tetramers. Memory CD8+ T cell responses were detected by FACS using IgG staining. Spleens were harvested 6 months after infection. MHC-I restriction and m139 560-574 IFN-γ production by CD8+ and CD4+ T cells after in vitro stimulation with MHC-II restricted mCMV peptides was measured using intracellular cytokines. The results were evaluated by immunofluorescence staining (Figure 1B).

[0065] Example 2 Intratumoral transduction of solid tumors with HPV Psv expressing mCMV antigens 1x10 to C57Bl / 6 mice 4 pfu of murine cytomegalovirus (mCMV Six months after infection, mice were infected with 2x10 5 TC-1 tumor cells were injected subcutaneously (sc) (injection protocol, Fig. 2A). Tumor growth was measured using electronic calipers. On days 13 and 15 after tumor injection, HPV16 Psv expressing m122 and m45 (Fig. 2B), or red fluorescent protein (RFP) were injected into the tumor. HPV Psv expressing radiofrequency adenosine phosphatase (RFP) (Figure 2C) were injected intratumorally (1 per Psv 0 8 infectious units).

[0066] Example 3 Intratumoral transduction of solid tumors using mCMV antigens in combination with poly(I:C) C57Bl / 6 mice, 1x10 4 pfu of murine cytomegalovirus (mCM Four months after infection, mice were infected with 2x10 5 TC-1 tumor cells (Figure 3A) were injected subcutaneously. Tumors were treated with m122, m38, and m12C on days 11 and 13 in the presence or absence of poly(I:C) (30 μg) (PIC). HPV16 expressing m122, m38 and m45 on days 16 and 18, HPV45 expressing m122, m38 and m45 on days 21 and 23, or control RFP (10 per PsV) 8 Infectious units (IU) were injected intratumorally. Tumor growth was measured using electronic calipers (Figures 3B-3E). These tumor volume / growth data are consistent with the intratumoral induction of solid tumors by HPV Psv expressing mCMV antigens. We demonstrate that co-administration of E7- (Fig. 3F), m45-, and m122- (Fig. 3G)-specific CD8+ T cells slows tumor growth, and that co-administration with poly(I:C) further slows tumor growth (compare Fig. 3B and 3D; and compare Fig. 3C and 3E). Infiltration of CD8+ T cells was analyzed by MHC-I tetramer staining and FACS. These data show that when these CMV antigens were administered in combination with poly(IC), tumor There is evidence that invasion was significantly enhanced.

[0067] Example 4 Intratumoral injection of mCMV MHC-I-restricted peptide results in improved survival C57Bl / 6 mice, 1x10 4 pfu of murine cytomegalovirus (mCM Four months after infection, mice were infected with 2x10 5 TC-1 tumor cells were injected subcutaneously (Figure 3A). Tumors were injected intratumorally with selected m38, m45 and m122 peptides (1 μg each) with or without poly(I:C) (30 ug), and with saline or poly(I:C) alone as controls, on days 11, 13, 16, 18, 21 and 23. Animal deaths were recorded (Figure 4A) and tumor growth was measured using electronic calipers (Figure 4B). These data indicate that intratumoral injection of mCMV MHC-I restricted peptides retards tumor growth and results in increased survival. This proves that.

[0068] Example 5 Intratumoral injection of mCMV MHC-I-restricted peptides delays tumor growth C57Bl / 6 mice, 1x10 4 pfu of murine cytomegalovirus (mCM Four months after infection, 2x10 5TC-1 tumor cells were subcutaneously injected into the tumors. Tumors were incubated with selected Decreasing doses (1 μg, 0.1 μg, and 0.01 μg) of the m38, m45, and m122 peptides were injected intratumorally, as well as saline or poly(I:C) alone as controls. Tumor growth was measured using electronic calipers (Figure 5). These data support the conclusion that mCMV M We show that intratumoral injection of HC-I restricted peptides retards tumor growth.

[0069] Example 6 Combination of mCMV MHC-I and MHC-II restricted peptides delays tumor growth 2.5x10 5 Four months after infection, mice were infected with 2x10 5 TC-1 tumor cells were subcutaneously injected. Tumors were injected intratumorally six times from day 12 to day 28 with MHC-I restricted selected m38, m45 and m122 peptides, and / or MHC-II restricted m139 selected peptides or saline. All peptides were injected together with poly(I:C) (30 μg). Groups were injected sequentially with 6 times MHC-I peptides, or 6 times MHC-II peptides, or 6 times MHCCI and MHCII peptides together, or 3 times MHC-I peptides followed by 3 times MHC-II peptides, or 3 times MHC-II peptides followed by 3 times MHC-I peptides. Tumor growth was measured using electronic calipers (Figures 6A and 6B). These data support the conclusion that mCMV MHC-I and MHC-II restricted peptides to delay tumor growth. Circulating E7-, m45-, and m122-specific CD8+ T cell responses were also The results were analyzed by FACS using MHC-I tetramers of each peptide (Figure 6C). These data support the conclusion that sequential tumor inoculation with mCMV CD4 epitopes followed by CD8 epitopes is consistent with previous data. These results indicate that certain species preferentially induce antitumor immunity.

[0070] Example 7 Complete clearance of primary tumors confers long-term tumor protection Protected C57Bl / 6 mice surviving the primary tumor challenge described in Example 6 were injected with 2x10 5 TC-1 tumor cells were injected subcutaneously. As a control for tumor harvest, young (12 week old) and age-matched (10 month old) mice were exposed to TC-1 tumor cells. Tumor growth was measured using electronic calipers (Figure 7). These data indicate that complete clearance of the primary tumor provides long-term protection against secondary tumor challenge.

[0071] Example 8 Intratumoral injection of MCMV alters the tumor immune microenvironment Effects of mCMV MHC-I and MHC-I on the tumor immune microenvironment in the presence or absence of PolyIC The effect of intratumoral injection of II-restricted peptide was analyzed for immune gene expression in RNA samples using the Nanostring Cancer immunology gene set (nCounter) 2 days after the end of the last intratumoral treatment. Results were summarized as score change for each gene set analyzed. Global score of expression difference by gene set was scored relative to saline-treated group (n=4 per group). Microenvironment characteristics evaluated included: B cell function, interleukins, TNF superfamily, antigen processing, MHC, adaptive immunity, transporter function, adhesion, NK cell function, T cell function, CD4 molecules, leukocyte function, complement pathway, microglia function, humoral immunity, TLR, inflammation, dendritic cell function, interferon, innate immunity, macrophage function, chemokines and receptors, senescence, apoptosis, cytokines and receptors, cancer progression, basal cell function, cell cycle, and pathogen response.

[0072] Example 9 mCMV infection was observed in CD8+ Inducing T cell responses C57Bl / 6 mice, 5x10 3 pfu of murine cytomegalovirus (mCM V). Blood samples were taken 1 month or 5 months after infection. Inflation (IE3) and non-inflation (m45) specific CD8+ T cells were detected by FACS using MHC-I tetramer staining. As shown in Figure 8, mCMV infection induced distinct effector and memory CD8+ T cell responses.

[0073] Example 10 mCMV infection induces potent CD8+ and CD4+ T cell proliferation in C57BL / 6 mice. Inducing a response C57Bl / 6 mice, 5x10 3 pfu of murine cytomegalovirus (mCM V). Blood samples were taken 12 days after infection. Spleen cells were cultured with the indicated peptides and m38, m45, m57, m122, m139, m141 and m164. The CD4+ and CD8+ T cells were stimulated with blood cells containing a collection of immunogenic peptides selected from mCMV proteins. Kine production was assessed by intracellular cytokine staining and analyzed by FACS (FIGS. 9A, 9B). These results indicate that murine cytomegalovirus infection induces a massive cytokine response.

[0074] Example 11 Tissue distribution of mCMV-specific CD8+ T cells The distribution of mCMV-specific CD8+ T cells in tumor-bearing mice was examined. / 6 mice, 5x10 3mCMV. The experimental schedule is shown in Figure 10A. Four months after infection, mice were infected with 2x10 5 TC-1 tumor cells were injected subcutaneously. Lymph node, spleen, salivary gland and tumor tissues were harvested and inflated (IE3; Fig. 10B) and non-inflated (m45; Fig. 10C) specific CD8+ T cells were detected by FACS using MHC-I tetramer staining. Expression of resident memory T cell markers was assessed using CD69 and CD103 antibodies. These results indicate that TC1 tumors are infiltrated by mCMV-specific CD8+ T cells. It was shown that.

[0075] Example 12 Gene expression analysis of the tumor microenvironment The expression of genes in tumor cells in a mouse model was measured using 100 μg / mL IgG antibody with 100 μg / mL IgG antibody (100 μg / mL); saline; poly I:C (PIC) (50 μg); mCMV m139 peptide (MHC-II restricted / CD4) (CD4) ( 3μg); mCMV m38, m122, m45 peptide (MHC-I restricted / CD8) ( CD8) (1 μg each); mCMV m139 + poly I:C (PIC CD4) (3 μg each); mCMV m38, m122, m45 peptide (MHC-I restricted / CD8) + poly I: C(PIC CD8) (1 μg each) intratumorally (4 animals per group). TC1 tumor cells were placed subcutaneously and treated three times at weeks 11, 13, and 16. A timeline of the experimental protocol is shown in Figure 11A. Following treatment and tumor harvest, tumors were analyzed using the QIACube. Tumor RNA was extracted using the Tumor PanCancer Immune Profiling The analysis was performed using the Nanostring Cancer immunology gene set (NS_MM_CANCERIMM_C3400), which measures gene transcripts forming 770 genes in the panel. The collected data is represented as heat maps of gene set expression within specific biological processes (adaptive immunity, antigen processing, T cell function, dendritic cell function, NK cell function, interferon, TNF superfamily genes). Construct volcano plots of gene expression changes relative to saline treatment (plots represent the change (represented as fold increase or fold decrease) of treatment group relative to control treatment (saline) with statistical significance). Apply cell infiltration quantification algorithms (CD45, cytotoxic CD8, CD4 Th1, NK cell The results were based on MHC-I-restricted / CD8 and MHC-I-restricted / CD8+ The greatest change in global significance score was observed in I:C-treated animals.

[0076] Immune gene profiling of total tumor RNA following intratumoral treatment showed significant upregulation of immune genes in three groups: 1) mCMV m139 peptide: MHC-II restricted / CD4 - 3mg (230 genes genes were upregulated and four genes were downregulated). 2) mCMV m38, IE3, m45 peptide: MHC-I restricted / CD8 - 1mg ( 359 genes were upregulated and 43 genes were downregulated). 3) mCMV m38, IE3, m45 peptide:MHC-I restricted / CD8+ poly(I:C) (309 genes were upregulated and 49 genes were downregulated).

[0077] Infiltration of the tumor by leukocytes was also analyzed after intratumoral treatment. Figures 11B to 11F show tumor infiltration by different leukocytes. These data indicate that CD8 mCMV epitopes ( We showed that intratumoral injection of mCMV with or without poly(I:C) induces recruitment of T cells and non-T cells (NK cells) within the tumor; and that the CD4 mCMV epitope together with poly(I:C) have shown that intratumoral injection of poly(I:C) induces the recruitment of T cells and non-T cells (NK cells) within tumors; and that intratumoral injection of poly(I:C) together with CD8 or CD4 epitopes induces the recruitment of dendritic cells within tumors.

[0078] Example 13 Intratumoral injection of mCMV slows tumor growth of mCMV CD8 epitopes C57Bl / 6 mice, 5x10 3 pfu of murine cytomegalovirus (mCM Four months after infection, 2x10 5 TC-1 tumor cells were injected subcutaneously. Tumor growth was measured using electronic calipers. Tumors were cultured on days 11, 13, 16, 18, 21 and 23 with selected MHC-I restricted m38, m45 and m122 peptides (0.0 μg each) in the presence or absence of poly(I:C) (30 μg). 1 μg, 0.1 μg, or 1 μg of mCMV MHC-I restricted peptide was injected intratumorally, and saline or poly(I:C) alone was injected as a control. We show that intratumoral injection of ribozyme slows tumor growth and that poly(I:C) coinjection improves tumor control.

[0079] Example 14 Tumor targeting of mCMV MHC-I and / or MHC-II peptides using poly(I:C) Protection from TC1 and MC38 tumor challenge by intratumoral injection C57Bl / 6 mice, 5x10 3 Four months after infection, 2x10 mice expressing E6 and E7 oncoproteins 5TC-1 tumor cells were subcutaneously injected at 100x100x100 / mL. Tumor growth and survival were monitored. Tumors were injected intratumorally 6 times with selected MHC-I restricted m38, m45 and m122 peptides, and / or selected MHC-II restricted m139 peptide, with or without poly(I:C) (30 μg), and saline or poly(I:C) alone as controls, from day 12 to day 28. Groups were injected sequentially 6 times with MHC-I peptides, or 6 times with MHC-II peptides, or 6 times with both MHC-I and MHC-II peptides, or 3 times with MHC-I peptides followed by 3 times with MHC-II peptides, or 3 times with MHC-II peptides followed by 3 times with MHC-I peptides. Figure 13A shows the mCMV MHC-I and MHC Figure 13B shows that intratumoral injection of a combination of MHC-II-restricted peptides delays tumor growth, and Figure 13B shows that sequential intratumoral inoculation with a CD4 (MHC-II) mCMV epitope followed by a CD8 (MHC-I) mCMV epitope promotes long-term survival.

[0080] Example 15 Blood E7 tetramer positive CD8 after treatment + T cell response C57Bl / 6 mice, 5x10 3 Four months after infection, 2x10 mice expressing E6 and E7 oncoproteins 5 TC-1 tumor cells s.c. Tumors were injected intratumorally six times from day 12 to day 28 with selected MHC-I restricted m38, m45 and m122 peptides, and / or selected MHC-II restricted m139 peptide, with or without poly(I:C) (30ug), as well as saline or poly(I:C) alone as controls. All peptides were injected with poly(I:C) (30ug). Groups were injected sequentially six times with MHC-I peptides, or six times with MHC-II peptides, or six times with both MHC-I and MHC-II peptides, or three times with MHC-I peptides followed by three with MHC-II peptides, or three times with MHC-II peptides followed by three with MHC-I peptides. Blood E7-, m45-, m122-specific CD8+ T cell responses were measured. The mCMV IgG antibody was analyzed by FACS using the MHC-I tetramer pairs of each peptide. Sequential intratumoral inoculation with CD4 epitopes followed by CD8 epitopes resulted in preferential antitumor The results show that it induces tumor immunity.

[0081] Example 16 Long-term protection against secondary tumor exposure Protected C57Bl / 6 mice that survived primary tumor challenge as described above were inoculated with 2x10 mice expressing E6 and E7 oncoproteins in the flank contralateral to the primary challenge. 5 TC-1 tumor cells were injected subcutaneously. Tumor growth was measured using electronic calipers. As a control for tumor ingestion, young (12 weeks old) and age-matched (10 months old) mice were challenged with TC-1 tumor cells. Figure 15 shows that complete clearance of the primary tumor provides long-term protection against secondary tumor challenge.

[0082] Example 17 Tumor mCMV MHC-I and MHC-II peptides with poly(I:C) Protection from MC38 tumor challenge by intravenous injection 5x10 C57Bl / 6 mice 3 Four months after infection, mice were infected with mCMV. In this study, 5x10 cells derived from a mouse colon adenocarcinoma exhibiting hypermutation and microsatellite instability were 5 MCM38 tumor cells were injected subcutaneously, and tumor growth was monitored. Tumors were injected intratumorally six times from day 12 to day 28 with selected MHC-I restricted m38, m45 and m122 peptides, or selected MHC-II restricted m139 peptide alone, with or without poly(I:C) (30 μg), and saline alone as a control. Figure 16 shows that complete clearance of the primary tumor provides long-term protection against secondary tumor challenge. Figure 16 shows that mCMV MHC-I and MHC-II 1 shows that intratumoral injection of a combination of I-restricting peptides slows tumor growth and results in tumor clearance.

[0083] The studies described in Examples 1 to 17 demonstrate that both non-inflating and inflating mCMV-specific T cells infiltrate tumors during latent mCMV infection, and that redirecting established antiviral T cells to solid tumors leads to profound changes in the tumor immune microenvironment and tumor regression. The data also demonstrate that the retargeting of established antiviral CD4+ T cells to solid tumors leads to profound changes in the tumor immune microenvironment and tumor regression. We show that redirecting T cells to solid tumors promotes epitope spreading to tumor-associated antigens and complete tumor clearance. Thus, these methods provide a broadly applicable "antigen agonistic" tumor treatment based on pre-existing antiviral T cells.

[0084] HPV L1 and L2 particles have strong tropism for many tumor cells. However, they do not bind to or infect intact epithelium. Thus, HPV PsV or VLPs can be used as a carrier to deliver antitumor agents genetically or directly to tumor cells.

[0085] While the present invention has been described with reference to specific embodiments, it will be understood by those skilled in the art that various modifications may be made and equivalents substituted without departing from the true spirit and scope of the invention. In addition, specific situations, materials, compositions of matter, processes, process steps (or Many modifications may be made to adapt the disclosed subject matter (including, but not limited to, the components, methods, and apparatus) to the objective, spirit and scope of the present invention, and all such modifications are intended to be within the scope of the appended claims.

[0086] The aspects of the present disclosure also include the following aspects. <1> A method of treating cancer in an individual, comprising recruiting an existing immune response to the cancer site, thereby treating the cancer. <2> the pre-existing immune response is a natural pre-existing immune response; <1> The method described above. <3> Recruiting a pre-existing immune response to the cancer cells includes introducing an antigen to the cancer that is not expressed by the cancer cells prior to the initiation of treatment, where the antigen is recognized by one or more components of the pre-existing immune response. <1> or <2> The method described above. <4> Prior to introducing the antigen into the tumor, it has been determined that the individual has a pre-existing immune response to the antigen. <1> from <3> 2. The method according to claim 1 , <5> the step of confirming the presence of a pre-existing immune response comprises identifying a T cell response to the antigen in a sample from the individual; <4> The method described above. <6> the step of introducing the antigen comprises injecting the antigen into the cancer; <1> from <5> 2. The method according to claim 1 , <7> the step of introducing an antigen comprises introducing a nucleic acid molecule encoding the antigen into the cancer; <1> from <6> 2. The method according to claim 1 , <8> the nucleic acid molecule is DNA; <7> The method described above. <9> the nucleic acid molecule is RNA; <7> The method described above. <10> RNA is modified to become more resistant to degradation; <9> The method described above. <11> The nucleic acid molecule is introduced into the cancer by injection; <7> The method described above. <12> The nucleic acid molecule is introduced into the cancer using a viral vector. <7> The method described above. <13> Viral vectors are introduced into cancer cells using pseudovirions. <12> The method described above. <14> the pseudovirion is a papillomavirus pseudovirion; <13> The method described above. <15> the antigen is a viral antigen; <3> from <14> 2. The method according to claim 1 , <16> the antigen is a polypeptide comprising at least one epitope derived from a cytomegalovirus (CMV) protein, wherein the at least one epitope is recognized by one or more components of a pre-existing immune response; <3> from <14> 2. The method according to claim 1 , <17> one or more of the components is a T cell; <16> The method described above. <18> The CMV protein is selected from the group consisting of pp50, pp65, pp150, IE-1, IE-2, gB, US2, US6, UL16 and UL18; <16> The method described above. <19> the polypeptide is an MHC I restricted peptide of 9-15 amino acids; <16> The method described above. <20> The polypeptide is an MHC II restricted peptide of at least 15 amino acids. <16> The method described above. <21> The antigen comprises a sequence at least 90% identical to a sequence selected from the group consisting of SEQ ID NOs: 1-67; <16> The method described above. <22> The antigen comprises a sequence selected from SEQ ID NOs: 1-67; <16> The method described above. <23> Recruitment of pre-existing immune responses alters the cancer microenvironment, selected from B cell function, interleukins, TNF superfamily, antigen processing, MHC, adaptive immunity, transporter function, adhesion, NK cell function, T cell function, CD molecules, leukocyte function, complement pathway, microglia function, humoral immunity, TLR, inflammation, dendritic cell function, interferon, innate immunity, macrophage function, chemokines and receptors, senescence, apoptosis, cytokines and receptors, cancer progression, basic cell functions, cell cycle, and pathogen response. <3> from <22> 2. The method according to claim 1 , <24> The antigen is administered in combination with an agent that enhances the immune response selected from TLR agonists; IL-1R8 cytokine antagonists; intravenous immunoglobulin (IVIG); peptidoglycan isolated from gram-positive bacteria; lipoteichoic acid isolated from gram-positive bacteria; lipoproteins isolated from gram-positive bacteria; lipoarabinomannan isolated from mycobacteria, zymosan isolated from yeast cell walls; polyadenylic-polyuridylic acid; poly(IC); lipopolysaccharide; monophosphoryl lipid A; flagellin; guardiquimod; imiquimod; R848; oligonucleosides containing CpG motifs, CD40 agonists and 23S ribosomal RNA, <3> from <23> 2. The method according to claim 1 , <25> The antigen is administered in combination with poly-IC; <3> from <23> 2. The method according to claim 1 , <26> the cancer is a solid tumor; <1> from <25> 2. The method according to claim 1 , <27> The cancer is a blood cancer, <1> from <26> 2. The method according to claim 1 , <28> A kit for mobilizing an existing immune response against cancer in an individual, comprising at least one CMV peptide antigen or a nucleic acid encoding the peptide, a pharma- ceutically acceptable carrier, a container, and a package insert or label describing administration of the CMV peptide to reduce cancer in a patient. <29> A kit for testing a patient and mobilizing an existing immune response in the patient against the site of cancer.

Claims

1. 1. A pharmaceutical composition for treating a solid tumor in an individual, comprising at least one MHC II-restricted peptide derived from a cytomegalovirus (CMV) protein and poly(I:C), wherein each MHC II-restricted peptide independently comprises any one of SEQ ID NOs: 1-33, each peptide recruiting a natural pre-existing immune response to the tumor site, thereby treating the tumor, wherein the natural pre-existing immune response is due to a previous CMV infection, and wherein the pharmaceutical composition is for injection into the solid tumor.

2. 10. The pharmaceutical composition of claim 1, comprising an MHC II restricted peptide comprising SEQ ID NOs: 1-33. thing.

3. 3. The pharmaceutical composition of claim 1 or claim 2, wherein the peptide is not expressed by tumor cells prior to the initiation of treatment and the peptide is recognized by one or more components of a pre-existing immune response.

4. The pharmaceutical composition of claim 3 , wherein the one or more components are T cells.

5. 5. The pharmaceutical composition of claim 1, wherein the individual is confirmed to have a pre-existing immune response to the at least one MHC II-restricted peptide as an antigen prior to administering the pharmaceutical composition to the individual.

6. The pharmaceutical composition of claim 5, wherein the presence of the pre-existing immune response is confirmed by identifying a T cell response to the antigen in a sample from the individual.

7. 7. The pharmaceutical composition of any one of claims 1 to 6, wherein the mobilization of a pre-existing immune response alters the cancer microenvironment selected from B cell function, interleukins, TNF superfamily, antigen processing, MHC, adaptive immunity, transporter function, adhesion, NK cell function, T cell function, CD molecules, leukocyte function, complement pathway, microglia function, humoral immunity, TLR, inflammation, dendritic cell function, interferon, innate immunity, macrophage function, chemokines and receptors, senescence, apoptosis, cytokines and receptors, cancer progression, basic cell functions, cell cycle, and pathogen response.

8. 8. The pharmaceutical composition of any one of claims 1 to 7, further comprising at least one MHC I-restricted peptide derived from a CMV protein, said CMV protein being selected from the group consisting of pp50, pp65, pp150, gB, IE-1, IE-2, US2, US6, UL16 and UL18.

9. 9. The pharmaceutical composition of claim 8, wherein each MHC I-restricted CMV peptide independently comprises the sequence of one of SEQ ID NOs: 34-67.

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