Cancer vaccine compositions and methods of use thereof

Abstract

To provide cancer vaccine compositions and methods of use thereof.SOLUTION: The disclosure provides a cancer vaccine composition comprising inactivated cancer cells and an adjuvant, wherein the inactivated cancer cells are incapable of performing replication. Also there is provided a method for producing a cancer vaccine composition, the method comprising contacting cancer cells with light (e.g., UV light) in the presence of a photosensitizer (e.g., riboflavin). Cancer immunotherapy involves the use of compositions and methods to elicit and enhance an individual's own immune system against cancerous cells, or infections that are factors predisposing to cancer.SELECTED DRAWING: None

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Patent Application No. 62 / 755,741, filed November 5, 2018; U.S. Provisional Patent Application No. 62 / 688,051, filed June 21, 2018; and U.S. Provisional Patent Application No. 62 / 645,975, filed March 21, 2018, each of which is incorporated by reference in its entirety for all purposes.

[0002] The present disclosure relates generally to compositions and methods for inhibiting tumor growth and promoting anti-tumor immune responses. More particularly, the present disclosure relates to cancer vaccine compositions and methods that activate the immune system's response against tumors. The present disclosure also relates to methods for making cancer cell vaccines. [Background technology]

[0003] Cancer immunotherapy involves the use of compositions and methods to induce and boost an individual's own immune system against cancer cells or infections that predispose to cancer. Cancer vaccines work by inducing the immune system to mount a response against an antigen (e.g., typically a protein, peptide, or carbohydrate) that is introduced into the body in a non-oncogenic form and induces the body to confer immunity or obtain a durable "memory" immune response. Once an immune system response is established, exposure of the immune system to this antigen (e.g., in the form of a cancerous tumor) results in a rapid and robust immune response. Summary of the Invention [Problem to be solved by the invention]

[0004] One of the challenges of cancer immunotherapy is that the clinical response between one patient and another often varies significantly.Some patients may have significant and durable response, while others do not derive obvious clinical benefit.Therefore, there is a need in the art for a composition that can reliably and effectively stimulate immune system as cancer immunotherapy. [Means for solving the problem]

[0005] Provided herein is a cancer vaccine composition comprising inactivated cancer cells that are incapable of replicating. The cancer cells may be isolated or derived from a patient suffering from one or more types of cancer.

[0006] Further provided is a method for treating cancer in a patient in need thereof, comprising administering to the patient a cancer vaccine of the present disclosure.

[0007] Further provided is a method for making a cancer vaccine composition, comprising treating cancer cells with light (e.g., UV light) in the presence of a photosensitizer (e.g., riboflavin).

[0008] Further provided is a cancer vaccine composition for use in a method of treating cancer.

[0009] Further provided is a cancer vaccine composition for use as a medicament for treating cancer, and the use of the cancer vaccine composition in the manufacture of a medicament for treating cancer. These and other aspects are described in further detail below. [Brief description of the drawings]

[0010] [Figure 1] The proliferation of CAMA cells following treatment with riboflavin and UV light is illustrated on the day of treatment (day 0) and 2, 4, 6, and 8 days after treatment. Cells were treated with a Mirasol® PRT lighting device at illumination intensities of 10%, 20%, 30%, 40%, 50%, or 100%. Cells treated with UV light (live) were included as a control. [Diagram 2] 1 shows the viability of CAMA cells after treatment with riboflavin and UV light. [Diagram 3] FIG. 1 shows the expression of the surface marker EpCAM in CAMA cells following treatment with riboflavin and UV light. [Figure 4]Fluorescence microscopy images are presented comparing surface marker expression on CAMA cells at various time intervals after treatment with riboflavin and UV light (20% illumination intensity). [Diagram 5] The relative expression of surface markers EpCAM (front row of bars) and CD38 (back row of bars) within the viable population after treatment with various doses of UV light is shown. [Figure 6] Caspase-3 levels in CAMA cells after treatment with riboflavin and UV light are shown. [Figure 7] 1 shows the correlation between surface marker expression and viability of CAMA cells following treatment with riboflavin and UV light. [Figure 8] Figure 1 shows tumor growth curves in saline injected PyMT breast cancer tumor bearing mice (control, no vaccine) compared to mice administered inactivated whole cell vaccine and mice administered lysate vaccine (4T1 spheroid lysate vaccine as described in WO 2016 / 161309, which is incorporated by reference in its entirety). Results show a statistically significant reduction in tumor cell growth in the inactivated whole cell vaccine group versus the untreated control group starting 23 days post-injection (p=0.02 at day 23 and p<0.001 at day 25). [Figure 9] Overall survival in vaccinated (inactivated whole cell vaccine) vs. untreated / saline (control) groups of PyMT tumor-bearing mice. Mice administered the inactivated whole cell vaccine had significantly increased survival compared to the saline-treated control group (p=0.009, Mantel-Cox log-rank test). [Figure 10] Figure 1 shows the size of 4T1 tumors in mice prior to surgical removal. After surgery, mice were placed into three groups, and each group was shown to have similar mean tumor size (p=0.9) and spread. PBS ("control"), n=5 mice; adjuvant, n=8 mice; inactivated whole cell vaccine, n=8 mice. [Figure 11A]Figure 11 shows the results of an experiment in which mice were treated weekly with PBS (control), losartan and cationic liposome-DNA complexes (CLDC) (adjuvant), or inactivated whole cell vaccine (adjuvant + vaccine) starting 24 hours after surgical removal of the primary tumor. Metastatic disease in the lungs was quantified using IVIS imaging after intraperitoneal injection of 100 μl of luciferin. As shown in Figure 11A, mice treated with vaccine showed a significant reduction in measured metastatic burden compared to adjuvant-treated mice (day 14, p = 0.0157) and compared to both control and adjuvant-treated mice (day 16, p = 0.0119 and p = 0.0021, respectively). Figure 11B shows the photon flux data over time for each mouse in each group. [Figure 11B] Figure 11 shows the results of an experiment in which mice were treated weekly with PBS (control), losartan and cationic liposome-DNA complexes (CLDC) (adjuvant), or inactivated whole cell vaccine (adjuvant + vaccine) starting 24 hours after surgical removal of the primary tumor. Metastatic disease in the lungs was quantified using IVIS imaging after intraperitoneal injection of 100 μl of luciferin. As shown in Figure 11A, mice treated with vaccine showed a significant reduction in measured metastatic burden compared to adjuvant-treated mice (day 14, p = 0.0157) and compared to both control and adjuvant-treated mice (day 16, p = 0.0119 and p = 0.0021, respectively). Figure 11B shows the photon flux data over time for each mouse in each group. [Figure 12] 1 shows the frequency of primary tumor regrowth due to incomplete removal of the primary tumor in various treatment groups. [Figure 13]Figure 1 is a survival curve showing enhanced survival of mice in the group administered the inactivated whole cell vaccine. Mice were euthanized when moribund (i.e., weight loss >10%, seizures, decreased mobility, scruffy appearance, etc.). Median survival for control and adjuvant groups was 17.5 days, while the inactivated whole cell vaccine-treated group was 24 days. This difference in survival was not statistically significant (p=0.1), but is biologically important and indicates the aggressiveness of 4T1 tumors. [Figure 14] 1 is a graph showing tumor size doubling time as described in Example 3. Doubling time was longer in mice treated with the inactivated whole cell vaccine (p=0.01). [Figure 15] 1 is a graph showing tumor growth area in mice bearing subcutaneous Lewis Lung Carcinoma (LLC) tumors and treated with PBS control or inactivated LLC vaccine at days 3, 5, 7, 10, 13, and 19 after tumor injection. At days 13 (p=0.02) and 19 (p=0.001) after tumor cell injection, tumor growth was significantly reduced in mice vaccinated with the inactivated LLC vaccine. [Figure 16] Graphs showing T cell subtypes within tumors from control and vaccinated mice from LLC studies. FIG. 16A shows the percent of T cells that were either CD4+CD25+ (putative T regulatory T cells) or CD8+CD25+. There was a significant decrease in CD4+CD25+ T cells in vaccinated mice. FIG. 16B shows the percent CD8+ T cells expressing immunosuppressive proteins PD-1, Lag3, or Tim3. FIG. 16C shows the percent CD4+ T cells expressing immunosuppressive proteins PD-1, Lag3, or Tim3. In each data set shown in FIG. 16A-C, the control is shown on the left and the vaccine is shown on the right. [Figure 17]Figure 1 shows IFNg production (pg / ml) after splenocytes were isolated from healthy naive B6 mice vaccinated and boosted with an inactivated whole cell vaccine (derived from 4T1 mouse tumor cells) and various immune adjuvants. Splenocytes were then restimulated in vitro with inactivated 4T1 tumor cells for 72 hours and IFNg was measured by ELISA. The CLDC adjuvant system produced the best IFNg response. [Figure 18] Shown is the mean fluorescence intensity (MFI) of serum IgG antibodies at 1:1000 dilution from the blood of the mice shown in Figure 17 binding to live 4T1 cells. All of the vaccine / adjuvant systems provided significantly higher binding than the control or inactivated cells alone. [Figure 19] Results of an experiment are shown in which mice were injected with 4T1 breast tumor cells, which were then surgically removed and used to generate an inactivated whole cell vaccine, after which metastatic disease in the lungs was quantified using IVIS imaging. Mice shown in the right panel were treated with the inactivated whole cell vaccine, and mice shown in the left panel did not receive either vaccine. Scale bars in luminescence are also presented. 62% of vaccinated mice were negative for lung metastases at the same time point, and 80% of untreated mice developed lung tumors 16 days after tumor cell removal. [Figure 20] A representative scheme for inactivating cells using UV light and riboflavin to prepare a vaccine composition and treat a patient in need thereof is shown. [Figure 21] Shown is cell surface staining of mouse LLC cells after UV+Rf (UV light+riboflavin) inactivation. [Figure 22] Shows surface staining of mouse 4T1 breast cancer cells after UV+Rf inactivation. [Diagram 23] Shows GFP expression in mouse melanoma GFP+B16 tumor cells after inactivation by either UV+RF or γ-rays in vitro. [Figure 24] FIG. 1 shows expression of mouse tumor-associated antigen gp70 following UV+Rf inactivation of mouse colon carcinoma, CT26, tumor cells. [Diagram 25]Shown is surface protein staining of inactivated ex vivo dog tumor tissue 1 hour and 48 hours after UV+Rf inactivation. Cells were kept at 4° C. for 48 hours after inactivation. [Figure 26] UV+RF inactivation of two additional ex vivo canine tumor tissues is shown. [Figure 27] Shown is the staining of inactivated human hepatoma cells HepG2 for the surface marker GLUT1. Inactivated cells are shown in the left panel, and live cells are shown in the right panel. The chart below shows the percentage of antibody positive and antibody negative cells when HepG2 cells were stained for the surface markers GLUT1 and HLA1. [Figure 28] 1 shows proliferation of T cells from the spleen of untreated 4T1 tumor-bearing mice. T cells proliferated in vitro when cultured with inactivated 4T1 tumor cells. [Figure 29] 1 shows the lack of proliferation of inactivated 4T1 mouse mammary carcinoma cells in culture at various time points after inactivation. [Diagram 30] Shown is data on the lack of proliferation of inactivated human hepatoma cells HepG2 (left panel) and the lack of proliferation of HepG2 and human colon cancer cells CRL-2577 (right panel). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0011] Provided herein is a method for inactivating cells and preventing their replication using UV light and riboflavin. This chemical process is specific to the DNA / RNA present in the cells. Thus, the DNA and / or RNA of the cells are modified while the proteins (including cell surface antigens, enzymes, etc.) remain untouched in the process. By preventing the replication process while preserving the antigens and phenotype of the cells, the treated cancer cell preparation can be used as a vaccine composition. Due to the fact that the antigen is present in its natural state on the cells of the vaccine composition, the immune response may be enhanced to a degree that exceeds the level observed with a single antigen or protein preparation intended to induce the same response. The combination of the inactivated whole cells with an adjuvant further enhances this immunological effect.

[0012] This technology can be used in an autologous or allogeneic manner, using tumor cells isolated from the patient, cancer stem cell preparations, or grown in culture, etc. When administered to patients, whole cell vaccines reduce tumor growth, decrease metastasis, and increase survival.

[0013] Thus, the technology described herein provides a rapid method for isolating, preparing, and administering cancer cell vaccines to patients to generate responses in patients that compete with the use of standard chemotherapeutic agents.

[0014] 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 disclosure belongs. The terminology used in the description of the present invention is for the purpose of describing particular embodiments only and is not intended to be limiting.

[0015] definition The following terminology is used in the description and appended claims.

[0016] The singular forms "a," "an," and "the" are intended to include the plural forms as well, unless the context clearly indicates otherwise.

[0017] Additionally, the term "about" as used herein when referring to a measurable value, e.g., amount, dose, time, temperature, and the like, is meant to encompass variations of ±20%, ±10%, ±5%, ±1%, ±0.5%, or even ±0.1% of the particular amount.

[0018] Additionally, as used herein, "and / or" refers to and includes any and all possible combinations of one or more of the associated listed items, as well as the lack of combinations when interpreted in the alternative ("or").

[0019] It is specifically contemplated that the various features described herein can be used in any combination, unless the context indicates otherwise.

[0020] As used herein, the terms "reduce", "reduces", "reduction" and similar terms refer to a decrease of at least about 10%, about 15%, about 20%, about 25%, about 35%, about 50%, about 75%, about 80%, about 85%, about 90%, about 95%, about 97% or more.

[0021] As used herein, the terms "enhance," "enhances," "enhancement," and similar terms refer to an increase of at least about 10%, about 15%, about 20%, about 25%, about 50%, about 75%, about 100%, about 150%, about 200%, about 300%, about 400%, about 500% or more.

[0022] The terms "treat", "treating" or "treatment of" (and grammatical variations thereof) mean that the severity of a patient's condition is reduced or at least partially improved or stabilized, and / or some relief, alleviation, reduction or stabilization of at least one clinical symptom is achieved, and / or a delay in the progression of a disease or disorder is observed.

[0023] The terms "prevent," "preventing," and "prevention" (and grammatical variations thereof) refer to the prevention and / or delay of onset of a disease, disorder, and / or clinical condition in a patient and / or a reduction in the severity of onset of a disease, disorder, and / or clinical condition compared to that which would occur in the absence of the methods of the present disclosure. Prevention can be complete, e.g., the disease, disorder, and / or clinical condition does not occur at all. Prevention can also be partial, to the extent that the onset and / or severity of onset of a disease, disorder, and / or clinical condition in a patient is less than would occur in the absence of the present disclosure.

[0024] "Therapeutically effective amount" as used herein refers to an amount that, when administered to a patient for treating a disease or at least one of the clinical symptoms of a disease, is sufficient to affect such treatment of the disease or its symptoms. "Therapeutically effective amount" may vary depending, for example, on the disease and / or symptoms of the disease, the severity of the disease and / or symptoms of the disease or disorder, the age, weight, and / or health of the patient to be treated, and the judgment of the prescribing physician. The appropriate amount in any given instance may be ascertained by one skilled in the art or may be determined by routine experimentation.

[0025] Cancer vaccine composition A cancer vaccine composition is provided herein. The composition comprises, consists essentially of, or consists of inactivated cancer cells, optionally in combination with an adjuvant. Cancer cells are inactivated by modifying their DNA and / or RNA, rendering them replication-incompetent. The modification of the DNA and / or RNA of the cells does not kill the cells, i.e., the cancer vaccine is a live, replication-inactivated vaccine. Because cell viability is maintained, the vaccine presents a live antigen target to the patient's immune system. Peripheral inoculation stimulates an immune response against primary tumors and metastases.

[0026] In some embodiments, the cancer vaccine comprises, consists essentially of, or consists of cancer cells that have been inactivated using a photochemical process that inactivates tumor cell DNA and / or RNA replication while preserving protein type and phenotype. In some embodiments, the DNA and / or RNA of the cancer cells in the cancer cell vaccine comprises modified bases. For example, in some embodiments, the DNA of the cancer cells in the vaccine may comprise modified guanine bases, such as oxidized guanine bases.

[0027] In some embodiments, the cancer cells are autologous cancer cells. As used herein, "autologous" refers to cells removed or derived from the same patient to whom the vaccine is administered. In some embodiments, the cancer cells are allogeneic cells. As used herein, "allogeneic" refers to cells removed or derived from a donor other than the patient to whom the vaccine is administered.

[0028] In some embodiments, the cancer cells are derived from a patient suffering from one or more types of cancer. For example, the cancer cells may be isolated or derived from a patient suffering from cancer. The cancer may be a solid tumor or a liquid tumor. The cancer cells may be isolated or derived from a primary tumor or a metastatic tumor. The cancer may be stage I, stage II, stage III, or stage IV. In some embodiments, the cancer cells may be derived from a patient suffering from breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, stomach cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, or blood cancer. In some embodiments, the skin cancer is melanoma. In some embodiments, the blood cancer is leukemia, lymphoma, or myeloma. In some embodiments, the leukemia is acute lymphocytic leukemia or acute myeloid leukemia. In some embodiments, the lymphoma is Hodgkin's lymphoma or non-Hodgkin's lymphoma. In some embodiments, the myeloma is multiple myeloma.

[0029] In some embodiments, the cancer cells are derived from an immortalized cancer cell line. As used herein, "cancer cell line" refers to a transformed cell line derived from a cancer sample. Typically, a cancer cell line is capable of generating a tumor when explanted into a suitable host. A cancer cell line typically retains characteristics in vitro that are common to the cancer from which it is derived, including, for example, loss of differentiation, loss of contact inhibition, and will undergo essentially unlimited cell division in vitro. A cancer cell line may include, for example, a cell line that has been genetically modified to express a protein that allows the cell to be more fully recognized via antigen-presenting cells.

[0030] In some embodiments, the cancer cells are cancer stem cells.

[0031] In some embodiments, the cells are derived from a non-cancerous but abnormal growth, ie, a benign tumor or growth.

[0032] In some embodiments, the cancer vaccine comprises, consists essentially of, or consists of leukocytes (e.g., tumor-associated macrophages), tumor-associated endothelial cells, tumor-associated fibroblasts, or any other cell type present in the tumor microenvironment.

[0033] In some embodiments, the cancer vaccine composition further comprises an adjuvant. The effect of the adjuvant is to boost the immunological response. In some embodiments, the adjuvant modifies monocyte function.

[0034] Examples of suitable adjuvants include saponin preparations, virosomes, virus-like particles, non-toxic derivatives of enterobacterial lipopolysaccharide (LPS), immunostimulatory oligonucleotides (e.g., immunostimulatory oligonucleotides having CpG motifs), mineral-containing compositions, oil emulsions, polymers, micelle-forming adjuvants (e.g., liposomes), immunostimulatory complex matrices (e.g., ISCOMATRIX), particles, squalene, phosphates, cationic liposome-DNA complexes (CLDC), DDA, DNA adjuvants. , gamma-insulin, ADP-ribosylating toxins, detoxified derivatives of ADP-ribosylating toxins, Freund's complete adjuvant, Freund's incomplete adjuvant, muramyl dipeptide, monophosphoryl lipid A (MPL), poly IC, CpG oligodeoxynucleotide (ODN), imiquimod, adjuvant system AS01, adjuvant system AS02, adjuvant system AS03, MF59®, and aluminum or aluminum salts (e.g., alum, aluminum phosphate, aluminum hydroxide). Other suitable adjuvants include TLR agonists, NOD agonists, and lipid-DNA agonist complexes.

[0035] In some embodiments, the cancer vaccine composition further comprises one or more agonists or antagonists.

[0036] In some embodiments, the agonist comprises a Toll-like receptor (TLR) agonist. In some embodiments, the TLR agonist is an agonist of TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, TLR11, or TLR12. In certain embodiments, the agonist is a TLR3 and / or TLR9 agonist.

[0037] In some embodiments, the antagonist is a CC chemokine receptor type 2 (CCR2) antagonist.

[0038] In some embodiments, the antagonist is an angiotensin receptor blocker (ARB), such as losartan, telmisartan, irbesartan, azilsartan, candesartan, eprosartan, olmesartan, or valsartan. In some embodiments, the ARB is administered at a dose of between about 5 and about 100 mg / kg, e.g., about 5, about 10, about 15, about 20, about 25, about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, about 90, about 95, or about 100 mg / kg.

[0039] In some embodiments, the cancer vaccine comprises at least one (i.e., one, two, or all three) of a TLR agonist, a CCR2 antagonist, and an ARB.

[0040] In some embodiments, the agonist or antagonist (e.g., TLR3 and / or TLR9 agonist) is contained in or conjugated to liposomes. Liposomes are spherical, self-closed vesicles composed of amphiphilic lipids. Liposomes may be unilamellar, having one lipid bilayer membrane, or multilamellar, having two or more concentrically arranged bilayers. Suitable liposomes may have a selected average particle size (diameter) of about 200-500 nm. Various methods of preparing liposomes and encapsulating therapeutic agents therein are well documented (see, e.g., U.S. Pat. Nos. 3,932,657, 4,311,712, and 5,013,556, all of which are incorporated herein by reference). Known methods include reverse phase evaporation, as described in U.S. Pat. No. 4,235,871, incorporated herein by reference.

[0041] The lipids used to form the liposomes described herein include vesicle-forming lipids having two hydrocarbon chains, typically acyl chains, and a polar head group. This class includes phospholipids, such as phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidic acid (PA), phosphatidylinositol (PI), and sphingomyelin (SM), where the two hydrocarbon chains are typically between about 14-22 carbon atoms in length and have varying degrees of unsaturation. The selection of lipids and ratios can be varied to achieve the degree of flexibility or rigidity desired, control stability, and / or control the release rate of the encapsulated agent. When more than one lipid type is used, an appropriate amount of a relatively unsaturated lipid (such as PC) may be used to form a stable liposome. In one embodiment, at least 45-50 mol % of the lipids used to form the liposomes is PC.

[0042] Liposomes may also contain lipids derivatized with hydrophilic polymers such as polyethylene glycol (PEG). Suitable hydrophilic polymers include polyvinylpyrrolidone, polyvinylmethylether, polymethyloxazoline, polyethyloxazoline, polyhydroxypropyloxazoline, polyhydroxypropylmethacrylamide, polymethacrylamide, polydimethylacrylamide, polyhydroxypropylmethacrylate, polyhydroxyethylacrylate, hydroxymethylcellulose, hydroxyethylcellulose, polyethylene glycol, polyaspartamide, and hydrophilic peptide sequences. Methods for preparing lipids derivatized with hydrophilic polymers are known (see, for example, U.S. Pat. No. 5,395,619, incorporated herein by reference).

[0043] In some embodiments, the cancer vaccine comprises a cationic liposome-DNA complex (CLDC).

[0044] In some embodiments, the cancer vaccine further comprises a photosensitizer, such as riboflavin (vitamin B2). In some embodiments, the cancer vaccine is substantially free of photosensitizers.

[0045] In some embodiments, the cancer vaccine composition further comprises a carrier. In some embodiments, the cells and / or photosensitizer are suspended in the carrier. In some embodiments, the carrier comprises normal saline (e.g., 0.9% sodium chloride), dextrose saline (e.g., 5% dextrose in 0.9% sodium chloride), phosphate buffered saline (e.g., 137 mmol / L NaCl, 2.7 mmol / L KCl, 10 mmol / L Na2HPO4, 2 mmol / L KH2PO4).

[0046] In some embodiments, the cancer vaccine composition further comprises one or more additional pharma- ceutically acceptable components known to those skilled in the art, including, but not limited to, pharma- ceutically acceptable carriers, diluents, excipients, adjuvants, fillers, buffers, preservatives, antioxidants, lubricants, stabilizers, solubilizers, surfactants (e.g., wetting agents), masking agents, colorants, flavoring agents, and sweeteners. Suitable carriers, diluents, excipients, etc. can be found in standard pharmaceutical textbooks. See, for example, Handbook of Pharmaceutical Additives, 2nd Edition (eds. M. Ash and I. Ash), 2001 (Synapse Information Resources, Inc., Endicott, New York, USA), Remington's Pharmaceutical Sciences, 20th edition, pub. Lippincott, Williams & Wilkins, 2000; and Handbook of Pharmaceutical Excipients, 2nd edition, 1994.

[0047] Method for producing a cancer cell vaccine The cancer cell vaccine described herein is made by using harmless chemicals in a selective process that blocks cell replication process while preserving antigen protein structure. More specifically, the cancer cell vaccine is made by the combined application of photosensitizer and light to make cancer cells replication-deficient while preserving other biological functions of cells and proteins treated. An exemplary scheme for making and using the cancer cell vaccine is shown in Figure 20. The process for making the cancer vaccine of the present disclosure is detailed below.

[0048] First, cancer cells are provided. The cancer cells may be autologous, i.e., removed or derived from the subject to be vaccinated. In some embodiments, the cancer cells may be allogeneic. The cancer cells may also be derived from a cancer cell line.

[0049] In some embodiments, the cancer cells are cancer stem cells. In some embodiments, the cancer vaccine comprises, consists essentially of, or consists of leukocytes (e.g., tumor-associated macrophages), tumor-associated endothelial cells, tumor-associated fibroblasts, or any other cell type present in the tumor microenvironment.

[0050] In some embodiments, the cancer cells are provided as a single cell suspension during inactivation. In some embodiments, the cells are suspended in a medium during inactivation. Exemplary media that may be used include, but are not limited to, RPMI1640, MEM, DMEM, IMDM, DMEM-F12, Opti-MEM, Ham's F12, Media 199, or combinations thereof.

[0051] The cancer cells are then inactivated using photochemical techniques. This is accomplished using photosensitizers that can act as electron transfer agents. The application of photosensitizers that can be placed in an excited state in the vicinity of guanine bases in DNA or RNA constructs allows for selective modification of these bases (e.g., oxidation, crosslinking, fragmentation, deamination). Since electrochemical reactions can only occur over short distances, the photosensitizer needs to bind or associate with (i.e., intercalate) nucleic acids to carry out the desired chemical reaction.

[0052] In some embodiments, the photosensitizer is a flavin, such as riboflavin (vitamin B2), flavin mononucleotide, or flavin adenine dinucleotide. In some embodiments, the photosensitizer is a tertiary aliphatic amine (e.g., 1,4-diazabicyclo(2,2,2)octane), piperazine, (e.g., N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid and 1,4-dimethylpiperazine), amino acid (e.g., tyrosine, tryptophan, histidine, methionine), enzyme (e.g., superoxide dismutase), or EDTA (ethylenediaminetetraacetic acid). In some embodiments, the photosensitizer is riboflavin.

[0053] The cells are added to a solution containing a photosensitizer (eg, riboflavin), or a photosensitizer is added to a solution containing the cells (eg, a single cell suspension of the cells in medium).

[0054] In some embodiments, the concentration of photosensitizer used during inactivation is about 10 μM to about 100 μM, e.g., about 10 μM, about 15 μM, about 20 μM, about 25 μM, about 30 μM, about 35 μM, about 40 μM, about 45 μM, about 50 μM, about 55 μM, about 60 μM, about 65 μM, about 70 μM, about 75 μM, about 80 μM, about 85 μM, about 90 μM, about 95 μM, or about 100 μM. In some embodiments, the solution contains a photosensitizer at a concentration of about 1 μM to about 50 μM, such as about 2 μM, about 3 μM, about 4 μM, about 5 μM, about 6 μM, about 7 μM, about 8 μM, about 9 μM, about 10 μM, about 15 μM, about 20 μM, about 25 μM, about 30 μM, about 35 μM, about 40 μM, about 45 μM, or about 50 μM. In some embodiments, the photosensitizer concentration is less than about 10 μM, such as less than about 9 μM, about 8 μM, about 7 μM, about 6 μM, about 5 μM, about 4 μM, about 3 μM, about 2 μM, or about 1 μM.

[0055] The solution containing the photosensitizer and cells (optionally in culture medium) is then subjected to a light treatment. The light treatment may include treatment with visible, ultraviolet, and / or infrared light. The light treatment inactivates DNA and / or RNA in the cancer cells by modifying the bases of these nucleic acids. In some embodiments, guanine bases are selectively modified. In some embodiments, guanine bases are selectively oxidized. Oxidized guanine bases cannot be repaired by natural enzymes and cellular repair mechanisms. As such, there is no possibility for the induced modifications to revert to a form that would restore the cells' ability to replicate.

[0056] In some embodiments, the light treatment comprises, consists essentially of, or consists of treatment with ultraviolet (UV) light. The UV light may be UV-A, UV-B, or UV-C light. The UV light may have a wavelength of 170-400 nm, including all ranges and subranges therebetween. For example, in some embodiments, the UV light has a wavelength of 315-400 nm, 310-320 nm, 280-360 nm, 280-315 nm, or 180-280 nm. The UV light may be provided by a UV light source known in the art, such as a Mirasol® PRT illuminator (TerumoBCT, Lakewood, Colorado). In some embodiments, the cells may be treated with multiple wavelengths of light simultaneously.

[0057] In certain embodiments, when riboflavin is used as the photosensitizer, UV light having a wavelength of 310-320 nm is used. The inventors have determined that this wavelength prevents riboflavin from reacting in free solution and thus generating undesirable oxygen free radicals. At these wavelengths, riboflavin will selectively react when intercalated into nucleic acids.

[0058] The dose of UV light may vary depending on the volume of the solution being treated. For example, the dose of UV light may be between 200-400 Joules (e.g., 300 Joules) for a volume of solution of about 170-370 ml. As will be appreciated by those skilled in the art, the dose may be adjusted upwards or downwards if the volume being treated is above or below this range.

[0059] In some embodiments, the dose of UV light may be about 200 Joules to about 600 Joules, for example, about 200, about 225, about 250, about 275, about 300, about 325, about 350, about 375, about 400, about 425, about 450, about 475, about 500, about 525, about 550, about 575, or about 600 Joules. In some embodiments, the volume of the cancer cell preparation for luminescence may be about 200 ml to about 600 ml, for example, about 200, about 225, about 250, about 275, about 300, about 325, about 350, about 375, about 400, about 425, about 450, about 475, about 500, about 525, about 550, about 575, or about 600 ml. In some embodiments, the dose of UV light can be about 0.5 J / ml to about 3.0 J / ml. For example, the dose of UV light can be about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1.0, about 1.1, about 1.2, about 1.3, about 1.4, about 1.5, about 1.6, about 1.7, about 1.8, about 1.9, about 2.0, about 2.1, about 2.2, about 2.3, about 2.4, about 2.5, about 2.6, about 2.7, about 2.8, about 2.9, or about 3.0 Joules / ml.

[0060] The cells may be treated with UV light for about 1 minute to about 60 minutes, e.g., about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 15, about 20, about 25, about 30, about 35, about 40, about 45, about 50, about 55, or about 60 minutes. In some embodiments, the cells are treated with UV light for about 1 minute to about 10 minutes, about 1 minute to about 5 minutes, or about 1 minute to about 3 minutes.

[0061] In some embodiments, the cancer cells are pre-incubated in a solution containing a photosensitizer (eg, riboflavin) for a period of time before subjecting the cells to light treatment.

[0062] In some embodiments, the cells do not undergo any additional purification or modification steps after light treatment. In other embodiments, the cancer cells are isolated and / or washed after light treatment. For example, the cells may be pelleted and optionally washed after light treatment. By pelleting and / or washing the cells, the photosensitizer (e.g., riboflavin) may be substantially removed from the composition. In some embodiments, the cancer cells are concentrated after light treatment.

[0063] In some embodiments, the cancer cells are resuspended or combined with one or more additional pharma- ceutically acceptable components, as described above, after light treatment, in some embodiments, the cancer cells are resuspended in a solution containing an adjuvant after light treatment.

[0064] In some embodiments, the cells remain viable for 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 days after light treatment, hi some embodiments, the cells die (e.g., by an apoptotic mechanism) 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 days after treatment.

[0065] The cells produced using this method are incapable of replicative processes, but substantially maintain and preserve the antigenic and epitope characteristics of the original native cells or antigens in the process. In some embodiments, the inactivation process does not substantially alter the metabolic processes, phenotype, or structure of the cancer cells. For example, in some embodiments, the inactivation process does not substantially alter the expression of cell surface markers in the cancer cells. In some embodiments, the inactivation process does not substantially alter the expression levels of cell surface markers such as EpCAM, CD38, CD34, CD117, CD44, CD24, Sca1, HLA, Glut1, MHC class I, PDL-L1, CD45, gp70, GFP, and / or CD90 in the cells. In some embodiments, the inactivation process does not compromise the integrity of the cell membrane and nuclear membrane of the cells.

[0066] The fact that the cells are replication-deficient provides protection against altered cellular compositions in the body that are involved in the formation of natural forms of disease (cancer) or neoplastic lesions. Thus, the inactivated cells generated by this process provide an improved source for antigen presentation, since the specificity of the chemicals preserves antigenic properties and cellular integrity, maintaining the protein structure in its native state.

[0067] Treatment method The cancer cell vaccine composition described herein can be used as a vaccine or stimulant for priming and recognition of the immune system to foster an immune response in cancer patients. This targeted therapy results in reduced side effects compared to traditional treatments such as chemotherapy or radiation. In particular, since the cancer cells in the vaccine maintain a normal phenotype, the possibility that they will induce undesired side effects is extremely low or nonexistent.

[0068] In some embodiments, the cancer cell vaccine may be administered to a patient to treat or prevent cancer in the patient. The cancer to be treated or prevented may be a solid tumor or a liquid tumor. For example, the cancer to be treated or prevented may be breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, stomach cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, or blood cancer. In some embodiments, the skin cancer is melanoma. In some embodiments, the blood cancer is leukemia, lymphoma, or myeloma. In some embodiments, the leukemia is acute lymphocytic leukemia or acute myeloid leukemia. In some embodiments, the lymphoma is Hodgkin's lymphoma or non-Hodgkin's lymphoma. In some embodiments, the myeloma is multiple myeloma.

[0069] In some embodiments, the vaccine may be administered to a patient to treat or prevent non-cancerous but abnormal growths, i.e., benign tumors or growths, in the patient. While most benign tumors / growths are treatable by surgery, some are in locations where surgery may not be possible and / or radiation may not be appropriate. Examples of non-cancerous growths that may be treated include, but are not limited to, adenomas, fibromas, neuromas, hemangiomas, seborrheic keratosis, dermatosis papularis nigricans, and sebaceous hyperplasia.

[0070] In some embodiments, the patient is evaluated for immune function and status prior to administration of the cancer vaccine. Such evaluation may include, but is not limited to, DTH skin tests, blood tests, lymph node aspirates, tumor tissue tests, and / or a determination of whether the patient is anergic, B cell responsiveness, etc. In some embodiments, the patient is not evaluated for immune function and status prior to administration of the cancer vaccine.

[0071] In some embodiments, the patient may be immunocompetent. In other embodiments, the patient may be immunocompromised. Optionally, the vaccine may be combined with genetic testing to quantitate the degree of immune responder or immune non-responder.

[0072] It will be understood by those skilled in the art that the appropriate number of cells in the cancer vaccine composition may vary between patients. In some embodiments, the cancer vaccine comprises about 1×10 3 , about 1×10 4 , about 1×10 5 , about 1×10 6 , about 1×10 7 , about 1×10 8 , about 1×10 9 , or about 1 × 10 10 In some embodiments, the cancer vaccine comprises about 1×10 cells. 5 ~Approx. 1×10 8 Contains cells.

[0073] In some embodiments, about 1 x 10 5 ~Approx. 1×108 For example, about 1×10 cells per administration may be administered to a patient. 5 , about 5×10 5 , about 1×10 6 , about 5×10 6 , about 1×10 7 , about 5×10 7 , or about 1 × 10 8 A total of 100 cells may be administered to the patient. In some embodiments, the administered dose is a split dose, where the total number of cells for administration is split into 2, 3, 4, 5, 6, 7, 8, 9, or 10 partial doses. One or more partial doses may be administered to the patient peripherally at different locations on the patient's body. Each partial dose may be administered at approximately the same time, or administration of the partial doses may be staggered. For example, the partial doses may be administered at intervals of 15 minutes, 20 minutes, 30 minutes, 45 minutes, 1 hour, or 3 hours.

[0074] In some embodiments, the cancer vaccine is administered to a patient once or more than once, hi some embodiments, the cancer vaccine is administered to a patient 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 times.

[0075] The cancer vaccine may be administered to a patient daily, about every 3 days, about every 7 days, about every 14 days, about once a month, or about once a year. In some embodiments, the cancer vaccine is administered at least once a week, at least once every 2 weeks, or at least once every 6 months. In some embodiments, the cancer vaccine is administered 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 20, or 25 times a year.

[0076] In some embodiments, a first cancer vaccine and a second cancer vaccine are administered to the patient. In some embodiments, the second cancer vaccine is administered after the first cancer vaccine to enhance the immune response. In some embodiments, the immune response and / or tumor growth in the patient is monitored between the administration of the first vaccine and the administration of the second vaccine. In some embodiments, the second cancer vaccine is administered when it is determined that the patient does not have a sufficient immune response after the administration of the first vaccine, or when it is determined that the tumor continues to grow or metastasize after the administration of the first vaccine. In some embodiments, the first cancer vaccine and the second vaccine comprise cells isolated or derived from a first tumor extract. For example, a tumor removed from a patient may be used to make the first and second vaccines, and after the administration of the first vaccine, the second vaccine is stored for later use. In some embodiments, the first vaccine and the second vaccine comprise cells isolated or derived from separate tumor extracts. For example, a tumor removed from a patient may be used to make a first vaccine, and after tumor recurrence or metastasis, the recurrent or metastatic tumor is removed and used to make a second vaccine.

[0077] The cancer vaccine may be delivered to a patient intramuscularly, intramucosally, intranasally, subcutaneously, intratumorally, intradermally, transdermally, intravaginally, intraperitoneally, intrarectally, intraarticularly or intralymphatically, orally or intravenously. In some embodiments, administration may be via sublingual, buccal, intravisceral (e.g., intrasplenic), or inhalation routes. For intravenous, cutaneous or subcutaneous injection, or injection into a tumor site, the cancer cell vaccine may be in the form of a parenterally acceptable aqueous solution with suitable pH, isotonicity and stability. Those skilled in the art are well able to prepare suitable solutions using isotonic vehicles, such as, for example, Sodium Chloride Injection, Ringer's Injection, Lactated Ringer's Injection, etc. Preservatives, stabilizers, buffers, antioxidants and / or other additives may be included as necessary.

[0078] In some embodiments, the vaccine is administered peripherally to the patient. In some embodiments, multiple aliquots of the cancer vaccine are administered peripherally to the patient at different locations.

[0079] In some embodiments, the cancer vaccine is administered simultaneously or sequentially (either before or after) with a vaccine booster. In some embodiments, the vaccine booster is an angiotensin receptor blocker (ARB) or a beta blocker (BB). Exemplary vaccine boosters include losartan, telmisartan, irbesartan, azilsartan, candesartan, eprosartan, olmesartan, valsartan, propranolol, acebutolol, atenolol, betaxolol, bisoprolol, carteolol, carvedilol, esmolol, labetalol, metoprolol, nadolol, nebivolol, penbutolol, pindolol, propranolol, sotalol, and timolol. In some embodiments, the vaccine booster is selected from the group consisting of losartan and propranolol. In some embodiments, the vaccine booster is losartan. In some embodiments, the vaccine booster is propranolol.

[0080] In some embodiments, the vaccination protocols described herein comprise administering a cancer cell vaccine composition comprising inactivated live cancer cells and a potent adjuvant comprising a TLR3 and / or TLR9 agonist attached to a liposome, and further comprise sequential or simultaneous administration of a vaccine enhancer (e.g., losartan) administered at or around the time of vaccination to reduce mobilization of immunosuppressive myeloid cells.

[0081] In some embodiments, the vaccination protocol described herein comprises administering to a patient in need thereof a cancer cell vaccine composition comprising inactivated live cancer cells. An adjuvant may optionally be administered at the time of vaccination. In some embodiments, the adjuvant is administered after vaccination, e.g., about 6 hours, about 12 hours, about 24 hours, about 36 hours, about 48 hours, about 60 hours, or about 72 hours after vaccination, to enhance the immune response. In some embodiments, the adjuvant comprises a liposome, e.g., CLDC. In some embodiments, a vaccine booster, such as losartan, may be administered at or around the time of vaccination. In some embodiments, a vaccine booster, such as losartan, may be administered after vaccination, e.g., about 6 hours, about 12 hours, about 24 hours, about 36 hours, about 48 hours, about 60 hours, or about 72 hours after vaccination. In some embodiments, a vaccine booster, such as losartan, may be administered to a patient, optionally daily, for a therapeutically effective number of days from the day the vaccine is administered. In some embodiments, the vaccine enhancer (e.g., losartan) is administered at a dose of between about 5 and about 100 mg / kg, e.g., about 5, about 10, about 15, about 20, about 25, about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, about 90, about 95, or about 100 mg / kg.

[0082] In some embodiments, the treatment reduces tumor growth or regrowth by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% compared to tumor growth in an unvaccinated patient. In some embodiments, the treatment extends patient survival by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% compared to an unvaccinated patient. In some embodiments, treatment reduces the incidence of metastasis by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% compared to unvaccinated patients.

[0083] The cancer cell vaccine may induce an immune response in a patient. In some embodiments, the immune response may include one or more of: (i) upregulation of immunoglobulins (e.g., IgG, IgM), (ii) T cell activation (e.g., production of multiple T cells that match multiple cancer neo-antigens), (iii) modulation of innate immune cells (e.g., myeloid cells), and (iv) restoration of "exhausted" T cell populations.

[0084] Suitable patients include both birds and mammals. The term "birds" as used herein includes, but is not limited to, chickens, ducks, geese, quail, turkeys, pheasants, parrots, parakeets, and the like. The term "mammals" as used herein includes, but is not limited to, humans, non-human primates, cows, sheep, goats, horses, cats, dogs, lagomorphs, and the like. Human subjects include neonatal, infant, juvenile, adult, and geriatric subjects. The terms "subject" and "patient" are used interchangeably herein.

[0085] The cancer cell vaccine may be administered to patients with a pre-existing condition, for example a pre-existing condition that has prevented treatment with other therapies, such as radiation, chemotherapy, or surgical resection.

[0086] Combination therapy The cancer cell vaccine may be administered alone or in combination with other treatments / therapies, simultaneously or sequentially, depending on the condition to be treated. Examples of treatments and therapies include, but are not limited to, chemotherapy (e.g., administration of active agents, including drugs, e.g., chemotherapeutic agents); surgery; and radiation therapy. Further examples of treatments and therapies include immune-based therapies, such as antibody therapy, adoptive cell therapy (ACT), and vaccine-based therapy. In some embodiments, the cancer cell vaccine described herein may be administered after another treatment / therapeutic to eliminate any remaining tumor cells.

[0087] In some embodiments, the cancer vaccine may be administered in combination with one or more of the following therapies: checkpoint inhibitors (e.g., PD-1 or PDL-1 inhibitors, antibody therapeutics, genetically modified dendritic cells, or genetically modified T cells (e.g., CAR-T cells).

[0088] In some embodiments, the cancer vaccine may be administered alone or in combination with a chemotherapeutic agent. A "chemotherapeutic agent" is a compound useful in the treatment of cancer, regardless of mechanism of action. Classes of chemotherapeutic agents include, but are not limited to, alkylating agents, antimetabolites, spindle poison plant alkaloids, cytotoxic / antitumor antibiotics, topoisomerase inhibitors, antibodies, photosensitizers, and kinase inhibitors. Chemotherapeutic agents include compounds used in "targeted therapy" and conventional chemotherapy.

[0089] Examples of suitable chemotherapeutic agents include erlotinib (TARCEVA®, Genentech / OSI Pharm.), docetaxel (TAXOTERE®, Sanofi-Aventis), 5-FU (fluorouracil, 5-fluorouracil, CAS number 51-21-8), gemcitabine (GEMZAR®, Lilly), PD-0325901 (CAS number 391210-10-9, Pfizer), cisplatin (cis-diamine, dichloroplatinum(II), CAS number 15663-27-1), carboplatin (CAS number 41575-94-4), paclitaxel (TAXOL®, Bristol-Myers Squibb), and the like. Oncology, Princeton, NJ), trastuzumab (HERCEPTIN®, Genentech), temozolomide (4-methyl-5-oxo-2,3,4,6,8-pentazabicyclo[4.3.0]nona-2,7,9-triene-9-carboxamide, CAS number 85622-93-1, TEMODAR®, TEMODAL®, Schering Plough), tamoxifen ((Z)-2-[4-(1,2-diphenyl-1-butenyl)phenoxy]-N,N-dimethylethanamine, NOLVADEX®, ISTUBAL®, VALODEX®), and doxorubicin (ADRIAMYCIN®), Akti-1 / 2, HPPD, and rapamycin.

[0090] Further examples of chemotherapeutic agents include oxaliplatin (ELOXATIN®, Sanofi), bortezomib (VELCADE®, Millennium Pharm.), Sutent (SUNITINIB®, SU11248, Pfizer), letrozole (FEMARA®, Novartis), imatinib mesylate (GLEEVEC®, Novartis), XL-518 (Mek inhibitor, Exelixis, WO 2007 / 044515), ARRY-886 (Mek inhibitor, AZD6244, Array BioPharma, Astra Zeneca), SF-1126 (PI3K inhibitor, Semaphores), and EGFR-1126 (PI3K inhibitor, Semaphores). Pharmaceuticals), BEZ-235 (PI3K inhibitor, Novartis), XL-147 (PI3K inhibitor, Exelixis), PTK787 / ZK222584 (Novartis), fulvestrant (FASLODEX®, AstraZeneca), leucovorin (folinic acid), rapamycin (sirolimus, RAPAMUNE®, Wyeth), lapatinib (TYKERB®, GSK572016, Glaxo Smith Kline), lonafarnib (SARASAR™, SCH66336, Schering Plough), sorafenib (NEXAVAR®, BAY43-9006, Bayer Labs), gefitinib (IRESSA®, AstraZeneca), irinotecan (CAMPTOSAR®, CPT-11, Pfizer), tipifarnib (ZARNESTRA™, Johnson & Johnson), ABRAXANE™ (Cremophor Free), an albumin-modified nanoparticle formulation of paclitaxel (American Pharmaceutical Partners, Schaumberg, Ill), vandetanib (rINN, ZD6474, ZACTIMA®, AstraZeneca), chlorambucil, AG1478, AG1571 (SU5271;Sugen), temsirolimus (TORISEL®, Wyeth), pazopanib (GlaxoSmithKline), canfosfamide (TELCYTA®, Telik), thiotepa and cyclosphosphamide (CYTOXAN®, NEOSAR®); alkyl sulfonates such as busulfan, improsulfan and piposulfan; aziridines such as benzodopa, carboquone, meturedopa and uredopa; ethylenimines and methylamelanamines such as altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide and trimethylomelanamine; acetogenins (especially bullatacin and bullatacinone); camptothecins (including the synthetic analog topotecan); bryostatin; kallistatin; CC-1065 (including its adzelesin, carzelcin) and synthetic analogs of biceresin); cryptophycins (particularly cryptophycin 1 and cryptophycin 8); dolastatins; duocarmycins (including synthetic analogs, KW-2189 and CB1-TM1); eleutherobin; pancratistatin; sarcodictyin; spongistatins; nitrogen mustards, such as chlorambucil, chlornaphazine, chlorophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembitine, phenesterine, prednimustine, trofosfamide, uracil mustard; nitrosoureas, such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, and ranimnustine; antibiotics, such as enediyne antibiotics (e.g., calicheamicin, calicheamicin γ1I, calicheamicin ω11 (Angew Chem. Intl. Ed. Engl. (1994) 33: 183-186); dynemicin, dynemicin A; bisphosphonates, clodronate, etc.; esperamicin;and neocarzinostatin chromophores and related chromoprotein enediyne antibiotic chromophores), aclacinomycin, actinomycin, ausramycin, azaserine, bleomycin, cactinomycin, carabicin, carminomycin, carzinophilin, chromomycinis, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin), epirubicin, esorubicin, idarubicin, nemorubicin, marcelomycin, mitomycins such as mitomycin C, mycophenolic acid, nogalamycin, olivomycin, peplomycin, porfiromycin, puromycin, keramycin, rhodomycin, antimetabolites such as rubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; antimetabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogues such as denopterin, methotrexate, pteropterin, trimetrexate; purine analogues such as fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogues such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine; androgens such as calsterone, dromostanolone propionate, epithiostanol, mepitiostane, testolactone; anti-adrenals such as aminoglutethimide, mitotane, trilostane; frolinic acid acid);aceglatone;aldophosphamide glycosides;aminolevulinic acid;eniluracil;amsacrine;bestrabutyl;bisantrene;edatraxate;defofamine;demecolcine;diaziquone;elfornitine;elliptinium acetate;epothilone;etoglucide;gallium nitrate;hydroxyurea;lentinan;lonidynin;maytansinoids such as maytansine and ansamitocin;mitoguazone;mitoxantrone;mopidamol;nitraelin;pentostatin;phenamet;pirarubicin;rosoxantrone;podophyllic acid;2-Ethylhydrazide; Procarbazine; PSK® polysaccharide complex (JHS Natural Products, Eugene, OR); Razoxane; Rhizoxin; Sizofiran; Spirogermanium; Tenuazonic acid; Triazicon; 2,2',2"-Trichlorotriethylamine; Trichothecines (especially T-2 toxin, veracrine A, roridin A, and anguidine); Urethane; Vindesine; Dacarbazine; Mannomustine; Mitobronitol; Mitolactol; Pipobroman; Gacytosine; Arabinoside ("Ara-C"); Cyclophosphamide; Thiotepa; 6-Thioguanine; Mercaptopurine; Methotrexate; Cisplatin and Carboplatin. platinum analogs such as vincristine; vinblastine; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; vinorelbine (NAVELBINE®); novantrone; teniposide; edatrexate; daunomycin; aminopterin; capecitabine (XELODA®, Roche); ibandronic acid; CPT-11; the topoisomerase inhibitor RFS2000; difluoromethylornithine (DMFO); retinoids such as retinoic acid; and pharmaceutically acceptable salts, acids, and derivatives of any of the above;

[0091] Further included within the definition of "chemotherapeutic agent" are: (i) anti-hormonal drugs that act to regulate or inhibit hormone action on tumors, e.g., antiestrogens and selective estrogen receptor modulators (SERMs), such as tamoxifen (NOLVADEX®; including tamoxifen citrate), raloxifene, droloxifene, 4-hydroxytamoxifen, trioxyphene, keoxyphene, LY117018, onapristone, and FARESTON® (toremifine citrate); (ii) aromatase inhibitors that inhibit the enzyme aromatase, which controls estrogen production in the adrenal glands, such as, for example, 4(5)-imidazole, aminoglutethimide, MEGASE® (megestrol acetate), AROMASIN® (exemestane; Pfizer), formestane (for mestanie), fadrozole, RIVISOR® (vorozole), FEMARA® (letrozole; Novartis), and ARIMIDEX® (anastrozole; AstraZeneca); (iii) antiandrogens such as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin; and troxacitabine (1,3-dioxolane nucleoside cytosine analogues); (iv) protein kinase inhibitors such as MEK inhibitors (WO 2007 / 044515); (v) lipid kinase inhibitors; (vi) antisense oligonucleotides, particularly those that inhibit the expression of genes in signal transduction pathways involved in abnormal cell proliferation, such as PKC-α, Raf and H-Ras, e.g., oblimersen (GENASENSE®, Genta Inc.(vii) ribozymes, such as VEGF expression inhibitors (e.g., ANGIOZYME®) and HER2 expression inhibitors; (viii) vaccines, such as gene therapy vaccines, e.g., ALLOVECTIN®, LEUVECTIN®, and VAXID®; PROLEUKIN® rIL-2; topoisomerase 1 inhibitors, such as LURTOTECAN®; ABAREFIX® rmRH; (ix) antiangiogenic agents, such as bevacizumab (Avastin®, Genentech); and pharmaceutically acceptable salts, acids, and derivatives of any of the above.

[0092] Further included within the definition of "chemotherapeutic agent" are therapeutic antibodies, such as alemtuzumab (Campath), bevacizumab (AVASTIN®, Genentech); cetuximab (ERBITUX®, Imclone); panitumumab (VECTIBIX®, Amgen), rituximab (RITUXAN®, Genentech / Biogen Idec), ofatumumab (ARZERRA®, GSK), pertuzumab (PERJETA™, OMNITARG™, 2C4, Genentech), trastuzumab (HERCEPTIN®, Genentech), tositumomab (Bexxar, Corixia), and the antibody drug conjugate gemtuzumab ozogamicin (MYLOTARG®, Wyeth).

[0093] Humanized monoclonal antibodies with therapeutic potential as chemotherapeutic agents in combination with the vaccines of the present disclosure include alemtuzumab, apolizumab, acelizumab, atlizumab, bapineuzumab, bevacizumab, bivatuzumab mertansine, cantuzumab mertansine, cedelizumab, certolizumab pegol, cidofusituzumab, cidtuzumab, daclizumab, eculizumab, efalizumab, epratuzumab, erlizumab, felvizumab, fontolizumab, gemtuzumab ozogamicin, inotuzumab ozogamicin, ipilimumab, labetuzumab, lintuzumab, matuzumab, mepolizumab, motavizumab, Motovizumab, natalizumab, nimotuzumab, norobizumab, numavizumab, ocrelizumab, omalizumab, palivizumab, pascolizumab, pecfusituzumab, pectuzumab, pertuzumab, pexelizumab, ralibizumab, ranibizumab, reslivizumab, reslizumab, resivizumab, rovelizumab These include, but are not limited to, ruplizumab, sibrotuzumab, siplizumab, sontuzumab, tacatuzumab tetraxetan, tadocizumab, talizumab, tefibazumab, tocilizumab, toralizumab, trastuzumab, tucotuzumab celmoleukin, tuxituzumab, umavizumab, urtoxazumab, and visilizumab.

[0094] Numbered Embodiments of the Invention 1. A cancer vaccine composition comprising inactivated cancer cells that are incapable of replicating. 2. The cancer vaccine composition of embodiment 1, wherein the cancer cells are derived from a patient suffering from one or more types of cancer. 3. The cancer vaccine composition of embodiment 2, wherein the patient is suffering from one or more of breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, gastric cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, and blood cancer. 4. The cancer vaccine composition of embodiment 3, wherein the skin cancer is melanoma. 5. The cancer vaccine composition of embodiment 3, wherein the hematological cancer is leukemia, lymphoma, or myeloma. 6. The cancer vaccine composition of embodiment 5, wherein the leukemia is acute lymphocytic leukemia or acute myeloid leukemia. 7. The cancer vaccine composition of embodiment 5, wherein the lymphoma is Hodgkin's lymphoma or non-Hodgkin's lymphoma. 8. The cancer vaccine composition of embodiment 5, wherein the myeloma is multiple myeloma. 9. The cancer vaccine composition of embodiment 2, wherein the patient is suffering from a benign tumor. 10. The cancer vaccine composition of embodiment 2, wherein the cancer is a metastatic cancer. 11. The cancer vaccine composition of any one of embodiments 1 to 10, wherein the cancer cells are derived from an immortalized cell line. 12. The cancer vaccine composition of any one of embodiments 1 to 11, wherein the cancer cells are autologous. 13. The cancer vaccine composition of any one of embodiments 1-11, wherein the cells are allogeneic. 14. The composition comprises about 1×10 5 ~Approx. 1×10 8 14. The cancer vaccine composition according to any one of embodiments 1 to 13, comprising a cancer cell. 15. The cancer vaccine composition of any one of embodiments 1-14, wherein the DNA of the cancer cell comprises a modified guanine base. 16. The cancer vaccine composition of any one of embodiments 1-15, wherein the composition further comprises an adjuvant. 17. The cancer vaccine composition of embodiment 16, wherein the adjuvant modifies monocyte function. 18. The cancer vaccine composition of embodiment 16, wherein the adjuvant comprises aluminum hydroxide. 19. The cancer vaccine composition of embodiment 16, wherein the adjuvant comprises CLDC. 20. The cancer vaccine composition of embodiment 16, wherein the adjuvant comprises PolyIC, CpG oligodeoxynucleotide (ODN), or imiquimod. 21. The cancer vaccine composition of embodiment 16, wherein the adjuvant comprises a liposome. 22. The cancer vaccine composition of embodiment 21, wherein the liposome is conjugated to an agonist. 23. The cancer vaccine composition of embodiment 22, wherein the agonist is an agonist of at least one of TLR3 and TLR9. 24. The cancer vaccine composition of any one of embodiments 1-23, further comprising a pharma- ceutically acceptable carrier. 25. The cancer vaccine composition of embodiment 24, wherein the pharma- ceutically acceptable carrier is normal saline, dextrose saline, or phosphate buffered saline. 26. The cancer vaccine composition of any one of embodiments 1-25, wherein the cancer cells are inactivated using light treatment. 27. The cancer vaccine composition of embodiment 26, wherein the light treatment lasts for about 1 minute to about 3 minutes. 28. The cancer vaccine composition of embodiment 26 or 27, wherein the structure of the antigen protein on the cancer cell is not substantially changed by light treatment. 29. The cancer vaccine composition of any one of embodiments 26 to 28, wherein the DNA of the cancer cells is modified by light treatment. 30. The cancer vaccine composition of embodiment 29, wherein light treatment selectively oxidizes guanine bases in the DNA of cancer cells. 31. The cancer vaccine composition of any one of embodiments 26 to 30, wherein the light treatment does not substantially alter the metabolic process, phenotype, or structure of the cancer cells. 32. The cancer vaccine composition according to any one of embodiments 26 to 31, wherein light treatment does not substantially alter the expression or activity of surface markers in cancer cells. 33. The cancer vaccine composition of embodiment 32, wherein light treatment does not alter the expression levels of EpCAM, CD38, CD34, CD117, CD44, CD24, Sca1, HLA, Glut1, MHC class I, PDL-L1, CD45, gp70, GFP, or CD90 in the cells. 34. The cancer vaccine composition of any one of embodiments 26 to 33, wherein the integrity of the cell membrane or nuclear membrane of the cells is not compromised by light treatment. 35. The cancer vaccine composition of any one of embodiments 26 to 34, wherein the light treatment comprises treatment with UV light. 36. The cancer vaccine composition of embodiment 35, wherein the UV light has a wavelength of 170 to 400 nm. 37. The cancer vaccine composition of embodiment 35, wherein the UV light has a wavelength of 315 to 400 nm. 38. The cancer vaccine composition of embodiment 35, wherein the UV light has a wavelength of 310 to 320 nm. 39. The cancer vaccine composition of embodiment 35, wherein the UV light has a wavelength of 280 to 360 nm. 40. The cancer vaccine composition of embodiment 35, wherein the UV light has a wavelength of 280 to 315 nm. 41. The cancer vaccine composition of embodiment 35, wherein the UV light has a wavelength of 180 to 280 nm. 42. The cancer vaccine composition of embodiment 35, wherein the UV light has a wavelength of 170 to 200 nm. 43. The cancer vaccine composition of any one of embodiments 35 to 42, wherein the dose of UV light is from about 200 Joules to about 600 Joules. 44. The cancer vaccine composition of embodiment 43, wherein the dose of UV light is about 200 Joules to 400 Joules. 45. The cancer vaccine composition of embodiment 44, wherein the dose of UV light is about 300 Joules. 46. ​​The cancer vaccine composition of any one of embodiments 26 to 45, wherein the light treatment is carried out by contacting the cancer cells with light in the presence of a photosensitizer. 47. The cancer vaccine composition of embodiment 46, wherein the concentration of the photosensitizer is from about 1 μM to about 50 μM. 48. The cancer vaccine composition of embodiment 46 or 47, wherein the concentration of the photosensitizer is less than about 10 μM. 49. The cancer vaccine composition of any one of embodiments 46-48, wherein the photosensitizer is riboflavin. 50. A method for treating cancer in a patient in need thereof, comprising administering to the patient a cancer vaccine composition according to any one of embodiments 1 to 49. 51. The method of embodiment 50, wherein the cancer vaccine composition is administered simultaneously or sequentially with the vaccine booster. 52. The method of embodiment 51, wherein the vaccine booster is an angiotensin receptor blocker (ARB) or a beta blocker (BB). 53. The method of embodiment 51 or 52, wherein the vaccine booster is losartan. 54. The method of embodiment 53, wherein the dose of losartan is between about 5 and about 100 mg / kg. 55. The method of embodiment 54, wherein the dose of losartan is about 60 mg / kg. 56. The method of embodiment 51 or 52, wherein the vaccine booster is propranolol. 57. The method of any one of embodiments 50-56, wherein the cancer vaccine composition is administered once to the patient. 58. The method of any one of embodiments 50-56, wherein the cancer vaccine composition is administered to the patient more than once. 59. The method of embodiment 58, wherein the cancer vaccine composition is administered to the patient 2, 3, 4, 5, 6, 7, 8, 9, or 10 times. 60. The method of embodiment 58 or 59, wherein the cancer vaccine composition is administered to the patient at least once every 7 days. 61. The method of embodiment 58 or 59, wherein the cancer vaccine composition is administered to the patient at least once every 14 days. 62. The method of embodiment 58 or 59, wherein the cancer vaccine composition is administered to the patient at least once every 6 months. 63. The method of any one of embodiments 50-62, wherein the cancer vaccine composition is administered by a route selected from subcutaneous, intramuscular, intravenous, intranasal, sublingual, buccal, inhalation, intradermal, intratumor, intravisceral, oral, and intraperitoneal. 64. The method of embodiment 63, wherein the cancer vaccine composition is administered by subcutaneous injection. 65. The method of embodiment 63, wherein the cancer vaccine composition is administered by intravenous injection. 66. The method of embodiment 63, wherein the cancer vaccine composition is administered by intramuscular injection. 67. The method of any one of embodiments 50-62, wherein the patient is immunocompetent. 68. The method of any one of embodiments 50-62, wherein the patient is immunocompromised. 69. The method of any one of embodiments 50-68, wherein the treatment reduces tumor growth by at least 10% compared to tumor growth in an unvaccinated patient. 70. The method of embodiment 69, wherein the treatment reduces tumor growth by at least 20% compared to tumor growth in an unvaccinated patient. 71. The method of embodiment 70, wherein the treatment reduces tumor growth by at least 50% compared to tumor growth in an unvaccinated patient. 72. The method of any one of embodiments 50-71, wherein the treatment extends patient survival by at least 10% compared to unvaccinated patients. 73. The method of embodiment 72, wherein the treatment extends patient survival by at least 20% compared to unvaccinated patients. 74. The method of embodiment 73, wherein the treatment extends patient survival by at least 50% compared to unvaccinated patients. 75. The method of any one of embodiments 50-74, wherein the treatment upregulates IgG and / or IgM in the patient. 76. The method of any one of embodiments 50-75, wherein the treatment activates T cells in the patient. 77. The method of any one of embodiments 50-76, wherein the cancer is breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, gastric cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, or blood cancer. 78. The method of embodiment 77, wherein the skin cancer is melanoma. 79. The method of embodiment 77, wherein the blood cancer is leukemia, lymphoma, or myeloma. 80. The method of embodiment 79, wherein the leukemia is acute lymphocytic leukemia or acute myeloid leukemia. 81. The method of embodiment 79, wherein the lymphoma is Hodgkin's lymphoma or non-Hodgkin's lymphoma. 82. The method of embodiment 79, wherein the myeloma is multiple myeloma. 83. The method of any one of embodiments 50-82, wherein the cancer is a metastatic cancer. 84. The method of any one of embodiments 50-83, wherein the cancer vaccine composition is administered to the patient in combination with one or more additional therapeutic agents. 85. The method of embodiment 84, wherein the one or more additional therapeutic agents are selected from the group consisting of checkpoint inhibitors, antibody therapeutic agents, genetically modified dendritic cells, genetically modified T cells, and chemotherapeutic agents. 86. A method for making a cancer vaccine, comprising contacting cancer cells with UV light in the presence of riboflavin. 87. The method of embodiment 86, wherein the DNA of the cancer cells is modified by UV light. 88. The method of embodiment 87, wherein UV light selectively oxidizes guanine bases in the DNA of cancer cells. 89. The method of embodiment 86, wherein the light treatment does not substantially change the structure of the antigen protein on the cancer cell. 90. The method of any one of embodiments 86-89, wherein UV light does not substantially alter the metabolic processes, phenotype, or structure of the cancer cells. 91. The method of any one of embodiments 86-90, wherein UV light does not substantially alter the expression or activity of surface markers in the cancer cells. 92. The method of embodiment 91, wherein UV light does not substantially alter the expression levels of EpCAM, CD38, CD34, CD117, CD44, CD24, Sca1, HLA, Glut1, MHC class I, PDL-L1, CD45, gp70, GFP and / or CD90 in the cells. 93. The method of any one of embodiments 86-92, wherein the inactivation does not compromise the integrity of the cell's cellular and nuclear membranes. 94. The method of any one of embodiments 86-93, wherein the cancer cells are contacted with UV light in the presence of riboflavin for about 1 minute to about 3 minutes. 95. The method of any one of embodiments 86-94, wherein the UV light has a wavelength of 170-400 nm. 96. The method of any one of embodiments 86-94, wherein the UV light has a wavelength of 315-400 nm. 97. The method of any one of embodiments 86-94, wherein the UV light has a wavelength of 310-320 nm. 98. The method of any one of embodiments 86-94, wherein the UV light has a wavelength of 280-360 nm. 99. The method of any one of embodiments 86-94, wherein the UV light has a wavelength of 280-315 nm. 100. The method of any one of embodiments 86-94, wherein the UV light has a wavelength of 180-280 nm. 101. The method of any one of embodiments 86-94, wherein the UV light has a wavelength of 170-200 nm. 102. The method of any one of embodiments 86-101, wherein the dose of UV light is from about 200 Joules to about 600 Joules. 103. The method of embodiment 102, wherein the dose of UV light is from about 200 Joules to about 400 Joules. 104. The method of embodiment 103, wherein the dose of UV light is about 300 Joules. 105. The method of any one of embodiments 86-104, wherein the cancer cells are present in a single cell suspension. 106. The method of embodiment 105, wherein riboflavin is added to the single cell suspension. 107. The method of any one of embodiments 86-106, wherein the cancer cells are preincubated in a solution containing riboflavin prior to contacting the cells with UV light. 108. The method of embodiment 107, wherein the solution contains 10 to 100 μM riboflavin. 109. The method of embodiment 107, wherein the solution comprises from about 1 μM to about 50 μM riboflavin. 110. The method of embodiment 107, wherein the solution comprises less than about 10 μM riboflavin. 111. A cancer vaccine composition according to any one of embodiments 1 to 49 for use as a medicament. 112. A cancer vaccine composition according to any one of embodiments 1 to 49, for use as a medicament for treating cancer. 113. A cancer vaccine composition according to any one of embodiments 1 to 49, for use in a method for treating cancer. 114. Use of a cancer vaccine composition according to any one of embodiments 1 to 49 in the manufacture of a medicament for treating cancer. 115. The cancer vaccine composition of any one of embodiments 111 to 113 or the use of embodiment 114, wherein the cancer is breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, gastric cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, or blood cancer. 116. The cancer vaccine composition or use of embodiment 115, wherein the skin cancer is melanoma. 117. The cancer vaccine composition or use of embodiment 115, wherein the blood cancer is leukemia, lymphoma, or myeloma. 118. The cancer vaccine composition or use of embodiment 117, wherein the leukemia is acute lymphocytic leukemia or acute myeloid leukemia. 119. The cancer vaccine composition or use of embodiment 117, wherein the lymphoma is Hodgkin's lymphoma or non-Hodgkin's lymphoma. 120. The cancer vaccine composition or use of embodiment 117, wherein the myeloma is multiple myeloma. 121. The cancer vaccine composition of any one of embodiments 111 to 113 or the use of embodiment 114, wherein the cancer is a metastatic cancer. EXAMPLES

[0095] The following examples are included herein for illustrative purposes only and are not intended to be limiting.

[0096] Example 1. Inactivation of tumor cells derived from tumor cell lines CAMA cells (human breast tumor line) were inactivated using UV light treatment in the presence of riboflavin. Cells were treated with a Mirasol® PRT illuminator at illumination intensities of 10% (190 Joules), 20% (380 Joules), 30% (570 Joules), 40% (760 Joules), 50% (950 Joules) or 100% (1896 Joules). Cells not treated with UV light (live) were included as controls. Proliferation (Figure 1), viability (Figure 2), cell surface marker expression (Figures 3, 4, 5), caspase activity (Figure 6), and cell and nuclear membrane integrity were examined (day 0) and 2, 4, 6, and 8 days after treatment.

[0097] As shown in Figure 1, treatment with riboflavin / UV light rendered the cells unable to replicate in culture. This effect was immediately evident on the day of treatment, even at doses as low as 190-380 Joules (10-20% illumination intensity). Even though they were inactivated, the cells remained substantially viable after treatment (Figure 3). Specifically, the cells remained intact and metabolically functional (Figure 4). After 4 days, the concentration of caspase-3 increased significantly (Figure 6), consistent with cell death by an apoptotic mechanism. Thus, after treatment, the cells no longer proliferate and slowly die over time.

[0098] Cell surface marker expression (EpCAM and CD38) was maintained at relatively consistent levels after treatment from doses as low as 190-380 Joules (10-20% illumination intensity) to as high as 1896 Joules (100% illumination intensity) (Figures 3 and 5). Cell viability correlated positively with surface marker expression (Figure 7). This indicates that after UV treatment, cells maintained the cell surface antigens required to generate an immune response, even over a wide range of UV doses.

[0099] This data establishes a dynamic therapeutic window for preparing inactivated cells without damaging the cell surface marker proteins required to stimulate antibody production.

[0100] Example 2. Vaccine Safety: Inactivation of Autologous Tumor Cells and Injection into Healthy Test Mice PyMT tumor cells were injected into wild-type C57Bl6 mice. After tumor growth, tumor tissues were harvested and used to generate cancer cell vaccines. A total of 2 × 10 tumor cells were harvested from seven C57Bl6 mice. 8 PyMT ex vivo tumor cells were resuspended in a solution containing (i) 265 ml of DMEM medium supplemented with 20% fetal bovine serum and glutamine (no antibiotics) and (ii) 35 ml of riboflavin. The cells were treated with a total dose of 300 Joules of UV light.

[0101] 1 x 10 of treated cells6 The cells were placed in culture medium and incubated under optimal conditions. After one month in culture, no evidence of growth or proliferation was observed, which corresponded to 100% inactivation of replicative capacity in the treated cells.

[0102] In addition, a total of 10 C57 / Bl6 mice were treated with 1 × 10 6 Inactivated cells were injected subcutaneously. After 1, 2, and 3 weeks, additional doses were administered (4 doses in total, 1 × 10 6 The animals were monitored for 160 days after the first injection. During the 160-day monitoring period, no tumors were observed in any of the test subjects, and no side effects of the injection were observed, which corresponded to the complete inactivation of the cells.

[0103] Finally, eight immunodeficient NOD / SCID mice were injected with 1 × 10 6 Inactivated PyMT cells were injected into the mice. The animals were monitored for 5 months after injection. No tumor growth was observed during this monitoring period.

[0104] Taken together, these data suggest that there are no safety concerns with the injected cell preparation.

[0105] Example 3. Vaccine efficacy: Injection of inactivated tumor cells into test mice bearing breast cancer inhibits tumor growth 2.5 × 10 5 viable PyMT cells were injected into the mice. Three days later, mice were treated with either saline (control, n=10), vaccine with inactivated tumor cells (n=10), or a pre-tested lysate vaccine (positive control, n=6). Vaccines were administered in the form of 1x10 6The vaccines contained 100 inactivated cells / mouse and were administered subcutaneously into both forelimbs under anesthesia (typically 100-110 μl / limb). Mice were further administered losartan (60 mg / kg) by intraperitoneal injection once daily for 3 days, starting on the day of vaccination, for a total of 3 doses. Finally, 24 hours after vaccination, mice were given a booster of CLDC adjuvant (100 μl, intraperitoneal). Vaccinations were repeated weekly for a total of 5 vaccine / losartan / CLDC boost cycles. Tumor growth was measured over time (length × width) using calipers. The extent of mortality observed over a 2-month period following injection was also monitored.

[0106] Figure 8 shows the tumor growth curves for saline injection (control, no vaccine) compared to administration of the inactivated whole cell vaccine and lysate vaccine (4T1 Spheroid Lysate Vax). Starting 23 days after injection, there was a statistically significant reduction in tumor cell growth in the inactivated whole cell vaccine group versus the untreated control group (p=0.02 at day 23 and p<0.0001 at day 25).

[0107] Figure 9 shows overall survival. In this experiment, mice were euthanized when the longest tumor diameter exceeded 15 mm. Mice administered the inactivated whole cell vaccine had a significantly extended survival time compared to the saline-treated control group (p=0.0038). The median overall survival time of mice administered the inactivated whole cell vaccine was 34 days compared to the control of 26 days (an increase of about 30%).

[0108] Figure 14 shows the doubling time. In mice treated with the inactivated whole cell vaccine, the doubling time of tumor growth was significantly longer compared to control mice (p=0.01).

[0109] Example 4. Vaccine Efficacy: Injection of inactivated tumor cells into test mice with invasive breast cancer reduces lung metastases, limits tumor regrowth, and enhances survival 4T1 breast cancer is a highly tumorigenic, invasive, transplantable tumor cell line that, unlike most tumor models, can spontaneously metastasize from the primary tumor in the mammary gland to multiple distant sites, including lymph nodes, blood, liver, lung, brain, and bone. 1 × 10 6 4T1 luciferase tumor cells were injected into each mouse. Eleven days after injection (mean tumor area was 52 mm 2 When tumor size was 0.01, primary tumors were measured and then surgically removed. One mouse died during surgery. The remaining 21 mice were grouped according to pre-surgery tumor size with equal mean tumor size (Figure 10). The following groups were determined: PBS ("control"), n=5 mice; adjuvant, n=8 mice; inactivated whole cell vaccine, n=8 mice.

[0110] The surgically removed tumor tissue was then placed in culture medium overnight at 4°C. The following day, the tumor tissue was divided and then treated with collagenase. The cells were filtered to remove tissue debris and then quantified. A portion of the cells was used to create the first vaccine. Mice were injected with inactivated 4T1 tumor cells (1.7 x 10 6 The mice were vaccinated with 1000x1000 cells / mL + adjuvant once a week, boosted with adjuvant 24 hours after vaccination, and given 3 daily doses of 60 mg / kg losartan starting on the day of vaccination. This cycle was repeated weekly.

[0111] Mice were imaged periodically using the IVIS device to detect the development of metastatic disease. Mice were injected with 100 μl of luciferin and then 10 min later, subjected to IVIS. Photon flux counts were calculated using IVIS-based software and compared between groups (Figure 11A-B).

[0112] Starting 24 hours after surgical removal of the primary tumor, mice were treated weekly with PBS alone (control), cationic liposome-DNA complexes (CLDC) and losartan (adjuvant), or inactivated whole cell vaccine (adjuvant+vaccine). After intraperitoneal injection of 100 μl of luciferin, metastatic disease in the lungs was quantified using IVIS imaging (FIG. 19). As shown in FIG. 11A, there was a significant reduction in the measured metastatic burden in vaccine-treated mice compared to adjuvant-treated mice (day 14, p=0.0157) and compared to both control and adjuvant-treated mice (day 16, p=0.0119 and p=0.0021, respectively). FIG. 11B shows the photon flux data over time for each mouse in each group.

[0113] Some mice showed primary tumor regrowth due to incomplete resection. The number of mice with primary regrowth was recorded 17 days after surgery (Figure 12). Interestingly, fewer mice treated with inactivated whole cell vaccine had their primary tumor regrowth compared to the other treatment groups. 60% of mice in the control group, 63% in the adjuvant alone group, and only 38% in the inactivated whole cell vaccine group had primary tumor regrowth.

[0114] The effect of the vaccine on survival was also examined. Mice were euthanized when they showed signs of morbidity (loss of 10% or more of body weight, low activity, seizures, etc.) and were assessed the day after surgery (Figure 13). Median survival of mice treated with the inactivated whole cell vaccine was 24 days compared to 18 days in control mice and 17.5 days after surgical removal of the primary tumor in adjuvant alone mice. Thus, treatment with the vaccine increased median survival by approximately 6 days. All mice in all groups eventually succumbed to metastatic disease. Any observed effects are noteworthy, as the 4T1 metastatic model is highly aggressive and typically non-responsive to many conventional therapies.

[0115] Example 5. Testing efficacy of inactivated whole cell vaccines in different mouse tumor models (LLC) To test the efficacy of the inactivated whole cell vaccine in different mouse tumor models, healthy B6 mice were injected with Lewis Lung Carcinoma (LLC) cells. When the primary tumors developed, they were excised and the tumor cells were inactivated with 300 J. Another 19 B6 mice were inactivated with 5 × 10 5 LLC cells were injected subcutaneously into the flank. Three days after tumor cell injection, mice received 1.7 × 10 LLC cells as their first vaccine. 6 Mice were administered 10 inactivated cells / mouse / vaccine. Additionally, mice were administered a CLDC adjuvant, e.g., losartan, and a CLDC boost 24 hours after vaccination. Mice were administered their vaccines weekly for a total of three vaccine doses. On day 19, all mice were euthanized and tumor tissues were collected and stained for immune cells.

[0116] As shown in FIG. 15, tumor growth was significantly reduced / delayed in inactivated whole cell vaccine-treated mice (p=0.02 on day 13 and p=0.001 on day 19). Two mice in the vaccine-treated group were tumor-free on day 19 (complete remission=20%). The final tumor weight of tumor-bearing mice was reduced in vaccine-treated mice, but not significantly (p=0.0547). The tumor doubling time was also reduced non-significantly, but was shorter in vaccine-treated mice (p=0.0570). In the tumors, there was a significant reduction in CD4+CD25+ T cells in vaccinated mice (p=0.004, putative immunosuppressive regulatory T cells), as well as a reduction in CD4+ T cells expressing GITR as another marker of regulatory T cells (p=0.02) (FIG. 16A). In association with this, a significant decrease in CD8+ T cells expressing the immunosuppressive proteins Lag3 (p=0.01) and Tim3 (p=0.05) (Figure 16B) and a significant decrease in CD4+ T cells expressing the proteins Lag3 (p=0.005) and Tim3 (p=0.02) (Figure 16C) was observed in vaccinated mouse tumors.

[0117] Example 6. Determine whether ex vivo canine tumor tissue has tumor cells that maintain surface expression of proteins Two tumor tissues were surgically obtained from dogs with autochthonous cancer undergoing treatment. One tissue was obtained from anal gland carcinoma (ASA) and the other from thyroid carcinoma (TC). The tissues were digested with collagenase and the resulting single cell suspension was frozen at -80°C in multiple vials. The tumor cells were later thawed and one bullet of cells was used as control cells and the other as inactivated cells. 300J was used to inactivate the dog cells. The number of cells used was less than previously used to inactivate the tumor cells. It took 1 minute 38 seconds to inactivate each group of cells. The tumor cells were then stained for expression of canine CD44, CD90, PD-L1 and CD45. CD45 was used to gate out all hematopoietic cells (Figure 25A-D). Analysis of "tumor" cells was performed for CD45 negative cells (including tumor cells, fibroblasts, endothelial cells, and other cell types).

[0118] In ASA, 1 hour after inactivation, 15% of cells expressed CD44 (compared to 20% in control), 1% expressed CD90 (compared to 0.3%), and 7.7% expressed PD-L1 (compared to 3.1%). In TC, after inactivation, 7.1% expressed CD44 (compared to 6.8% of control cells), 0.6% expressed CD90 (compared to 0.4%), and 0% expressed PD-L1 (compared to 0.2%). The remaining cells were placed at 4°C for 48 hours and then stained again. After 48 hours, in ASA, CD44=10% (11% in control), CD90=2% (3.2% in control), and PD-L1=9.5% (3% in control). In TC, CD44=5.8% (8.9% of controls), CD90=3.1% (3.2% of controls), and PD-L1=3% (2.9% of controls).

[0119] Additional tumor tissue pieces were obtained from two different dogs undergoing surgery; one was a GI population and the other was a lung population. The tumor tissue was digested with collagenase, washed, and then stained to create a single cell suspension. All cells were then frozen in cell freezing medium. Later, the cells were thawed and half of the cells were inactivated using a UV+RF (UV light + riboflavin) protocol. The other half was kept on ice. The cells were then stained for surface expression of canine MHC class I, CD44, CD90, and PD-L1 (Figure 26).

[0120] Taken together, these data indicate that surface markers are maintained on canine tumor cells in vitro following inactivation.

[0121] Example 7. Testing different adjuvants to generate significant cellular and humoral immune responses against inactivated tumor cells Healthy Balb / c mice were either vaccinated with PBS or inactivated cells alone, or with the inactivated 4T1 vaccine (used in the metastatic mouse tumor study described in Example 4) in the CLDC adjuvant system, with topical imiquimod (2 mg / kg) applied to the skin prior to vaccination, or mixed with CpG ODN (50 μg / vaccine). Vaccines were administered subcutaneously in equal amounts to both the left and right forelimbs proximal to the paws of the mice. Mice received 1.7×10 6 Cells / mouse / vaccine were administered. Vaccine was administered on day 1 and then re-administered on day 14. Mice were euthanized 8 days after booster vaccine administration and spleens and blood were collected. Splenocytes were cultured with inactivated 4T1 cells (to block replication) and IFNg production was measured after 72 hours (Figure 17). Splenocytes were cultured at a ratio of 25 splenocytes to 1 inactivated 4T1 cell.

[0122] As shown in Figure 17, the CLDC adjuvant system had the best recall response in terms of IFNg production, followed by CpG ODN. Vaccine alone also produced some IFNg in 3 out of 4 mice, but this was not significantly different from the control.

[0123] Example 8. Further testing of adjuvants to generate significant cellular and humoral immune responses against inactivated tumor cells Healthy Balb / c mice were either vaccinated with PBS or inactivated cells alone, or with the inactivated 4T1 vaccine (used in the metastatic mouse tumor study described in Example 4) in the CLDC adjuvant system, with topical imiquimod (2 mg / kg) applied to the skin prior to vaccination, or mixed with CpG ODN (50 μg / vaccine). Vaccines were administered subcutaneously in equal amounts to both the left and right forelimbs proximal to the paws of the mice. Mice received 1.7×10 6 Cells / mouse / vaccine were administered. Vaccine was administered on day 1 and then re-administered on day 14. Mice were euthanized 8 days after booster vaccine administration and spleens and blood were collected. Serum from these mice was screened for IgG antibody binding against live 4T1 tumor cells by incubating serum with the cells at either 1:500 or 1:1000 dilution, staining with fluorescently labeled donkey anti-mouse secondary antibody, and then subtracting background staining (normal mouse serum at 1:500 and 1:1000).

[0124] As shown in Figure 18, unvaccinated and inactivated whole cell vaccine alone did not produce any IgG specific to 4T1 tumor cells. Imiquimod produced variable amounts, with 3 out of 5 mice having very high staining. CpG ODN produced a reliable and significant increase in staining at a dilution of 1:1000. Staining not much higher than background was detected at a dilution of 1:500.

[0125] Example 9. Preservation of surface antigens The mouse lung cancer line LLC maintained expression of surface antigens after inactivation (Figures 21A-21B). LLC cells were treated with riboflavin (RF, 50 μM) and UV light (300 J) and stained for surface expression of CD34, CD117, CD44, and CD90. All four antigens were maintained on the surface of the cells. The mouse breast cancer cell line 4T1 was also assessed for surface marker expression of CD44, Sca1, and EpCAM before and after UV+RF inactivation (Figure 22).

[0126] The mouse melanoma tumor cell line B16 transfected with green fluorescent protein was injected into C57B16 mice, removed from the mice, reconstituted into single cell suspensions, and then analyzed for expression of GFP+ tumor cells before and after UV+RF inactivation and after gamma irradiation (100 Gy). Neither form of cell inactivation negatively affected expression of GFP by tumor cells in ex vivo tumor tissue (Figure 23).

[0127] Finally, the murine colon cancer cell line CT26 was inactivated with UV+RF and then analyzed for surface expression of the known tumor-associated antigen gp70 (Figures 24A-24B). UV+RF inactivation enhanced the expression of gp70 on CT26 cells.

[0128] Furthermore, the human hepatoma cell line HepG2 was inactivated and stained for human HLA and GLUT1 expression after UV+RF inactivation (300 J, FIG. 27) and imaged on an adherent cell cytometer.

[0129] Results from all of these studies indicate that inactivation can be achieved without significant modifications to cell-specific antigens on the cell surface. Moreover, these markers are maintained in morphologically intact cells over long storage periods following treatment, and these observations have been observed in cells derived from all three species tested (mouse, dog, and human).

[0130] Example 10. Induction of the immune system by inactivated cells Splenocytes were removed from mice with 4T1 tumor growth but not receiving any treatment, and placed in culture to test the T cell immune response against either cells from the 4T1 live cell line compared to cells from the UV+RF inactivated 4T1 cell line (Figure 28). Splenocytes were cultured at 1x10 6 Plate at 4 x 10 cells / well. 5 Cells / well were co-cultured with (live or inactivated) tumor cells for 72 hours. After 48 hours of culture, the proliferation dye EdU was added. Splenocytes were then harvested and stained for CD4+ and CD8+ T cell proliferation. A significant increase in both CD4+ and CD8+ T cell proliferation was observed when the cells were co-cultured with UV+RF inactivated 4T1 tumor cells.

[0131] Example 11: Pharmacokinetics: Tumor cell proliferation and persistence in vitro and in vivo To test the safety and efficacy of the inactivation process, PyMT tumor cells were removed from solid tumors on B6 mice and inactivated using UV light (300 J). 6 5 healthy B6 mice were subcutaneously injected with 1x10 "viable" inactivated PyMT cells. This was repeated on days 7, 14, and 21 for a total of four doses. Approximately 160 days after the initial inactivated tumor cell injection, the mice were euthanized and excised. No evidence of tumor growth was observed. To test the safety of the inactivated cells in immunodeficient mice, 8 NOD / SCID mice were injected with 1x10 6 Inactivated PyMT cells were injected subcutaneously into the right flank. Mice were monitored thereafter. Three mice died during the study, presumably from unrelated causes (probably viral or bacterial infection). Necropsy was performed on one mouse, which revealed no tumors, and skin analysis was performed on the other two mice, which also showed no tumors. 252 days after tumor cell injection, the mice were euthanized, and the remaining five mice were tumor-free.

[0132] Cell proliferation in culture after inactivation was also evaluated (Figure 29). 4T1 cells were injected into the fat pad of Balb / c mice and tumor growth was monitored until the tumors reached approximately 10 mm in diameter. The tissue was removed, digested with collagenase, washed, inactivated with UV+Rf, and then cultured at 37°C for 24 and 72 hours. No proliferation of 4T1 cells was observed 24 and 72 hours after inactivation. This was also compared to non-inactivated ex vivo tumor cells (partial proliferation) and the 4T1 tumor cell line (proliferation). Similar studies were performed with the human liver cancer cell line HepG2 and the human colon cancer cell line CRL-2577 (Figure 30). Proliferation studies were performed using Click-iT EdU, which labels newly formed DNA. The fluorescence of the labeled new DNA is compared to total DNA staining using an adherent cell cytometer. This study identified thousands of replicating cells, up to 20-35% of high EdU cells in live controls in CRL-2577 and approximately 75% in the longer-term labeled HepG2 population.

[0133] Although the teachings of the present disclosure have been described with respect to various applications, methods, and compositions, it will be understood that various changes and modifications can be made without departing from the teachings herein and the scope of the following claims. The foregoing examples are provided to more fully illustrate the teachings of the present disclosure and are not intended to limit the scope of the teachings presented herein. While the present teachings have been described in terms of these exemplary embodiments, those skilled in the art will readily appreciate that numerous variations and modifications of these exemplary embodiments are possible without undue experimentation. All such variations and modifications are within the scope of the present teachings.

[0134] All references cited herein, e.g., patents, patent applications, articles, textbooks, GenBank™ or accession numbers, etc., and references cited therein, unless they have already been incorporated herein, are hereby incorporated by reference in their entirety. In the event that one or more of the incorporated documents and similar material differs or conflicts with this application, including, but not limited to, defined terms, term usage, techniques discussed, etc., this application controls.

[0135] The foregoing description and examples detail certain specific embodiments of the invention and set forth the best mode contemplated by the inventors. However, no matter how detailed the foregoing description and examples are in the text, it will be understood that the invention may be practiced in many ways and that the invention must be construed in accordance with the appended claims and any equivalents thereof. In certain embodiments, for example, the following items are provided: (Item 1) A cancer vaccine composition comprising inactivated cancer cells that are incapable of replicating. (Item 2) 2. The cancer vaccine composition of item 1, wherein the cancer cells are derived from a patient suffering from one or more types of cancer. (Item 3) 3. The cancer vaccine composition of item 2, wherein the patient is suffering from one or more of breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, gastric cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, and blood cancer. (Item 4) 4. The cancer vaccine composition of item 3, wherein the skin cancer is melanoma. (Item 5) 4. The cancer vaccine composition of item 3, wherein the blood cancer is leukemia, lymphoma, or myeloma. (Item 6) Item 6. The cancer vaccine composition according to item 5, wherein the leukemia is acute lymphocytic leukemia or acute myeloid leukemia. (Item 7) 6. The cancer vaccine composition of item 5, wherein the lymphoma is Hodgkin's lymphoma or non-Hodgkin's lymphoma. (Item 8) 6. The cancer vaccine composition of item 5, wherein the myeloma is multiple myeloma. (Item 9) 3. The cancer vaccine composition of item 2, wherein the patient is suffering from a benign tumor. (Item 10) 3. The cancer vaccine composition of item 2, wherein the cancer is a metastatic cancer. (Item 11) 11. The cancer vaccine composition according to any one of items 1 to 10, wherein the cancer cells are derived from an immortalized cell line. (Item 12) 12. The cancer vaccine composition according to any one of items 1 to 11, wherein the cancer cells are autologous. (Item 13) The cancer vaccine composition according to any one of items 1 to 11, wherein the cells are allogeneic. (Item 14) The composition is about 1×10 5 ~Approx. 1×10 8 14. The cancer vaccine composition according to any one of items 1 to 13, comprising a cancer cell. (Item 15) 15. The cancer vaccine composition according to any one of items 1 to 14, wherein the DNA of the cancer cell comprises a modified guanine base. (Item 16) 16. The cancer vaccine composition according to any one of items 1 to 15, wherein the composition further comprises an adjuvant. (Item 17) 17. The cancer vaccine composition of item 16, wherein the adjuvant modifies monocyte function. (Item 18) Item 17. The cancer vaccine composition of item 16, wherein the adjuvant comprises aluminum hydroxide. (Item 19) 17. The cancer vaccine composition of item 16, wherein the adjuvant comprises CLDC. (Item 20) 17. The cancer vaccine composition of item 16, wherein the adjuvant comprises polyIC, CpG oligodeoxynucleotide (ODN), or imiquimod. (Item 21) 17. The cancer vaccine composition of item 16, wherein the adjuvant comprises a liposome. (Item 22) 22. The cancer vaccine composition of claim 21, wherein the liposome is conjugated to an agonist. (Item 23) 23. The cancer vaccine composition of item 22, wherein the agonist is an agonist of at least one of TLR3 and TLR9. (Item 24) 24. The cancer vaccine composition according to any one of items 1 to 23, further comprising a pharma- ceutically acceptable carrier. (Item 25) 25. The cancer vaccine composition of item 24, wherein the pharma- ceutically acceptable carrier is normal saline, dextrose saline, or phosphate buffered saline. (Item 26) 26. The cancer vaccine composition according to any one of items 1 to 25, wherein the cancer cells are inactivated using light treatment. (Item 27) 27. The cancer vaccine composition according to item 26, wherein the light treatment lasts for about 1 minute to about 3 minutes. (Item 28) 28. The cancer vaccine composition according to item 26 or 27, wherein the light treatment does not substantially change the structure of the antigen protein on the cancer cell. (Item 29) 29. The cancer vaccine composition according to any one of items 26 to 28, wherein DNA of the cancer cells is not altered by the light treatment. (Item 30) 30. The cancer vaccine composition according to item 29, wherein the light treatment selectively oxidizes guanine bases in the DNA of the cancer cells. (Item 31) 31. The cancer vaccine composition according to any one of items 26 to 30, wherein the light treatment does not substantially change the metabolic process, phenotype, or structure of the cancer cells. (Item 32) 32. The cancer vaccine composition according to any one of items 26 to 31, wherein the light treatment does not substantially change the expression or activity of a surface marker in cancer cells. (Item 33) 33. The cancer vaccine composition according to item 32, wherein the light treatment does not change the expression levels of EpCAM, CD38, CD34, CD117, CD44, CD24, Sca1, HLA, Glut1, MHC class I, PDL-L1, CD45, gp70, GFP, or CD90 in the cells. (Item 34) 34. The cancer vaccine composition according to any one of items 26 to 33, wherein the light treatment does not impair the integrity of the cell membrane or nuclear membrane of the cells. (Item 35) 35. The cancer vaccine composition according to any one of items 26 to 34, wherein the light treatment comprises treatment with UV light. (Item 36) Item 36. The cancer vaccine composition according to item 35, wherein the UV light has a wavelength of 170 to 400 nm. (Item 37) Item 36. The cancer vaccine composition according to item 35, wherein the UV light has a wavelength of 315 to 400 nm. (Item 38) Item 36. The cancer vaccine composition according to item 35, wherein the UV light has a wavelength of 310 to 320 nm. (Item 39) Item 36. The cancer vaccine composition according to item 35, wherein the UV light has a wavelength of 280 to 360 nm. (Item 40) Item 36. The cancer vaccine composition according to item 35, wherein the UV light has a wavelength of 280 to 315 nm. (Item 41) Item 36. The cancer vaccine composition according to item 35, wherein the UV light has a wavelength of 180 to 280 nm. (Item 42) Item 36. The cancer vaccine composition according to item 35, wherein the UV light has a wavelength of 170 to 200 nm. (Item 43) 43. The cancer vaccine composition according to any one of items 35 to 42, wherein the dose of UV light is from about 200 Joules to about 600 Joules. (Item 44) Item 44. The cancer vaccine composition according to item 43, wherein the dose of UV light is about 200 Joules to 400 Joules. (Item 45) Item 45. The cancer vaccine composition of item 44, wherein the dose of UV light is about 300 Joules. (Item 46) 46. ​​The cancer vaccine composition according to any one of items 26 to 45, wherein the light treatment is carried out by contacting the cancer cells with light in the presence of a photosensitizer. (Item 47) Item 47. The cancer vaccine composition according to item 46, wherein the photosensitizer has a concentration of about 1 μM to about 50 μM. (Item 48) 48. The cancer vaccine composition of item 46 or 47, wherein the concentration of the photosensitizer is less than about 10 μM. (Item 49) 49. The cancer vaccine composition according to any one of items 46 to 48, wherein the photosensitizer is riboflavin. (Item 50) 50. A method for treating cancer in a patient in need thereof, comprising administering to said patient a cancer vaccine composition according to any one of items 1 to 49. (Item 51) 51. The method of claim 50, wherein the cancer vaccine composition is administered simultaneously or sequentially with a vaccine booster. (Item 52) 52. The method of claim 51, wherein the vaccine enhancer is an angiotensin receptor blocker (ARB) or a beta blocker (BB). (Item 53) 53. The method of claim 51 or 52, wherein the vaccine booster is losartan. (Item 54) The method according to Item 53, wherein the dose of losartan is between about 5 and about 100 mg / kg. (Item 55) 55. The method of claim 54, wherein the dose of losartan is about 60 mg / kg. (Item 56) 53. The method of claim 51 or 52, wherein the vaccine booster is propranolol. (Item 57) 57. The method according to any one of items 50 to 56, wherein the cancer vaccine composition is administered once to the patient. (Item 58) 57. The method of any one of items 50 to 56, wherein the cancer vaccine composition is administered to the patient two or more times. (Item 59) 59. The method of claim 58, wherein the cancer vaccine composition is administered to the patient 2, 3, 4, 5, 6, 7, 8, 9, or 10 times. (Item 60) 60. The method of item 58 or 59, wherein the cancer vaccine composition is administered to the patient at least once every 7 days. (Item 61) 60. The method of item 58 or 59, wherein the cancer vaccine composition is administered to the patient at least once every 14 days. (Item 62) 60. The method of item 58 or 59, wherein the cancer vaccine composition is administered to the patient at least once every 6 months. (Item 63) 63. The method according to any one of items 50 to 62, wherein the cancer vaccine composition is administered by a route selected from subcutaneous, intramuscular, intravenous, intranasal, sublingual, buccal, inhalation, intradermal, intratumor, intravisceral, oral, and intraperitoneal. (Item 64) 64. The method of claim 63, wherein the cancer vaccine composition is administered by subcutaneous injection. (Item 65) 64. The method of claim 63, wherein the cancer vaccine composition is administered by intravenous injection. (Item 66) 64. The method of claim 63, wherein the cancer vaccine composition is administered by intramuscular injection. (Item 67) 63. The method of any one of items 50 to 62, wherein the patient is immunocompetent. (Item 68) 63. The method according to any one of items 50 to 62, wherein the patient is immunocompromised. (Item 69) 69. The method of any one of items 50 to 68, wherein the treatment reduces tumor growth by at least 10% compared to tumor growth in an unvaccinated patient. (Item 70) 70. The method of claim 69, wherein the treatment reduces tumor growth by at least 20% compared to tumor growth in an unvaccinated patient. (Item 71) 71. The method of claim 70, wherein the treatment reduces tumor growth by at least 50% compared to tumor growth in an unvaccinated patient. (Item 72) 72. The method of any one of items 50 to 71, wherein the treatment extends survival of the patient by at least 10% compared to an unvaccinated patient. (Item 73) 73. The method of claim 72, wherein the treatment extends survival of the patient by at least 20% compared to an unvaccinated patient. (Item 74) 74. The method of claim 73, wherein said treatment extends survival of said patient by at least 50% compared to an unvaccinated patient. (Item 75) 75. The method of any one of items 50 to 74, wherein the treatment upregulates IgG and / or IgM in the patient. (Item 76) 76. The method of any one of items 50 to 75, wherein the treatment activates T cells in the patient. (Item 77) 77. The method according to any one of items 50 to 76, wherein the cancer is breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, gastric cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, or blood cancer. (Item 78) 78. The method of claim 77, wherein the skin cancer is melanoma. (Item 79) 78. The method of claim 77, wherein the blood cancer is leukemia, lymphoma, or myeloma. (Item 80) 80. The method of item 79, wherein the leukemia is acute lymphocytic leukemia or acute myeloid leukemia. (Item 81) 80. The method of claim 79, wherein the lymphoma is Hodgkin's lymphoma or non-Hodgkin's lymphoma. (Item 82) 80. The method of claim 79, wherein the myeloma is multiple myeloma. (Item 83) 83. The method of any one of items 50 to 82, wherein the cancer is a metastatic cancer. (Item 84) 84. The method of any one of items 50 to 83, wherein the cancer vaccine composition is administered to the patient in combination with one or more additional therapeutic agents. (Item 85) 85. The method of item 84, wherein the one or more additional therapeutic agents are selected from the group consisting of checkpoint inhibitors, antibody therapeutics, genetically modified dendritic cells, genetically modified T cells, and chemotherapeutic agents. (Item 86) 1. A method for making a cancer vaccine, comprising contacting cancer cells with UV light in the presence of riboflavin. (Item 87) 87. The method of claim 86, wherein the UV light modifies DNA of the cancer cells. (Item 88) 88. The method of claim 87, wherein the UV light selectively oxidizes guanine bases in the DNA of the cancer cells. (Item 89) 87. The method of claim 86, wherein the light treatment does not substantially change the structure of the antigen protein on the cancer cells. (Item 90) 90. The method of any one of items 86 to 89, wherein the UV light does not substantially alter the metabolic process, phenotype, or structure of the cancer cells. (Item 91) 91. The method according to any one of items 86 to 90, wherein the UV light does not substantially alter the expression or activity of a surface marker in the cancer cells. (Item 92) 92. The method of claim 91, wherein the UV light does not substantially alter the expression levels of EpCAM, CD38, CD34, CD117, CD44, CD24, Sca1, HLA, Glut1, MHC class I, PDL-L1, CD45, gp70, GFP, and / or CD90 in the cells. (Item 93) 93. The method according to any one of items 86 to 92, wherein the inactivation does not compromise the integrity of the cell membrane and nuclear membrane of the cell. (Item 94) 94. The method according to any one of items 86 to 93, wherein the cancer cells are contacted with UV light in the presence of riboflavin for about 1 minute to about 3 minutes. (Item 95) 95. The method according to any one of items 86 to 94, wherein the UV light has a wavelength of 170 to 400 nm. (Item 96) 95. The method according to any one of items 86 to 94, wherein the UV light has a wavelength of 315 to 400 nm. (Item 97) 95. The method according to any one of items 86 to 94, wherein the UV light has a wavelength of 310 to 320 nm. (Item 98) 95. The method according to any one of items 86 to 94, wherein the UV light has a wavelength of 280 to 360 nm. (Item 99) 95. The method according to any one of items 86 to 94, wherein the UV light has a wavelength of 280 to 315 nm. (Item 100) 95. The method according to any one of items 86 to 94, wherein the UV light has a wavelength of 180 to 280 nm. (Item 101) 95. The method according to any one of items 86 to 94, wherein the UV light has a wavelength of 170 to 200 nm. (Item 102) 102. The method according to any one of items 86 to 101, wherein the dose of UV light is from about 200 Joules to about 600 Joules. (Item 103) Item 103. The method according to item 102, wherein the dose of UV light is from about 200 Joules to about 400 Joules. (Item 104) Item 104. The method of item 103, wherein the dose of UV light is about 300 Joules. (Item 105) 105. The method according to any one of items 86 to 104, wherein the cancer cells are present in a single cell suspension. (Item 106) 106. The method of claim 105, wherein the riboflavin is added to the single cell suspension. (Item 107) 107. The method of any one of items 86 to 106, wherein the cancer cells are preincubated in a solution containing riboflavin prior to contacting the cells with the UV light. (Item 108) Item 108. The method according to item 107, wherein the solution contains 10 to 100 μM riboflavin. (Item 109) Item 110. The method according to Item 107, wherein the solution contains about 1 μM to about 50 μM riboflavin. 108. The method of claim 107, wherein the solution contains less than about 10 μM riboflavin. (Item 111) 50. The cancer vaccine composition according to any one of items 1 to 49 for use as a medicament. (Item 112) 50. The cancer vaccine composition according to any one of items 1 to 49, for use as a medicament for treating cancer. (Item 113) 50. The cancer vaccine composition according to any one of items 1 to 49 for use in a method for treating cancer. (Item 114) 50. Use of a cancer vaccine composition according to any one of items 1 to 49 in the manufacture of a medicament for treating cancer. (Item 115) 115. The cancer vaccine composition according to any one of items 111 to 113, or the use according to item 114, wherein the cancer is breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, gastric cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, or blood cancer. (Item 116) Item 116. The cancer vaccine composition or use according to item 115, wherein the skin cancer is melanoma. (Item 117) Item 116. The cancer vaccine composition or use according to item 115, wherein the blood cancer is leukemia, lymphoma, or myeloma. (Item 118) Item 118. The cancer vaccine composition or use according to item 117, wherein the leukemia is acute lymphocytic leukemia or acute myeloid leukemia. (Item 119) Item 118. The cancer vaccine composition or use according to item 117, wherein the lymphoma is Hodgkin's lymphoma or non-Hodgkin's lymphoma. (Item 120) Item 118. The cancer vaccine composition or use according to item 117, wherein the myeloma is multiple myeloma. (Item 121) 115. The cancer vaccine composition according to any one of items 111 to 113, or the use according to item 114, wherein the cancer is a metastatic cancer.

Claims

1. A cancer vaccine composition comprising inactivated cancer cells, The inactivated cancer cells are obtained by treating cancer cells with UV light, the DNA of the inactivated cancer cells comprises oxidized guanine bases, and the inactivated cancer cells are (i) is replication-deficient; (ii) is viable; and (iii) characterized by conserved expression of one or more cell surface antigen proteins; Cancer vaccine compositions.

2. 2. The cancer vaccine composition of claim 1, wherein the one or more antigenic proteins comprise EpCAM, CD38, CD34, CD117, CD44, CD24, Sca1, HLA, Glut1, MHC class I, PD-L1, CD45, gp70, or CD90.

3. The cancer vaccine composition of claim 2 , wherein the one or more antigenic proteins include EpCAM and / or CD38.

4. The cancer vaccine composition of any one of claims 1 to 3, wherein the inactivated cancer cells are obtained by treating cancer cells with UV light in the presence of riboflavin.

5. The cancer vaccine composition of claim 4, wherein the cancer cells are treated with UV light comprising a wavelength of 310 to 320 nm.

6. 6. The cancer vaccine composition of claim 4 or 5, wherein the treatment with UV light lasts from about 1 minute to about 3 minutes.

7. 7. The cancer vaccine composition of claim 4, wherein the UV light is at a dose ranging from about 200 joules to about 600 joules, or from about 0.6 joules / ml to 2 joules / ml.

8. 8. The cancer vaccine composition of claim 7, wherein the UV light is at a dose of about 300 joules, or 190 joules to 380 joules, or about 1 joule ml.

9. The cancer vaccine composition of any one of claims 4 to 8, wherein the riboflavin is at a concentration of about 1 µM to about 50 µM.

10. 10. The cancer vaccine composition of claim 4, wherein the inactivated cancer cells are viable for at least 1 day, at least 2 days, at least 3 days, at least 4 days, or at least 5 days after treatment with UV light.

11. The cancer vaccine composition according to any one of claims 1 to 10, wherein the cancer cells are breast cancer cells, lung cancer cells, liver cancer cells, bladder cancer cells, gynecological cancer cells, brain cancer cells, stomach cancer cells, prostate cancer cells, skin cancer cells, thyroid cancer cells, pancreatic cancer cells, colon cancer cells, or blood cancer cells.

12. The cancer vaccine composition of any one of claims 1 to 11, wherein the cancer cells are isolated from a subject with cancer.

13. The cancer vaccine composition according to any one of claims 1 to 11, wherein the cancer cells are derived from an immortalized cell line.

14. The cancer vaccine composition of any one of claims 1 to 11, wherein the cancer cells are isolated from a benign tumor.

15. The composition is about 1×10 ５ ~Approx. 1×10 ８ The cancer vaccine composition according to any one of claims 1 to 14, comprising cancer cells.

16. The composition is about 1×10 ６ The cancer vaccine composition of claim 15, comprising cancer cells.

17. The cancer vaccine composition of any one of claims 1 to 16, further comprising an adjuvant.

18. 18. The cancer vaccine composition of claim 17, wherein the adjuvant is a Toll-like receptor (TLR) 3 agonist, a TLR9 agonist, a TLR4 agonist, or a TLR7 agonist.

19. 19. The cancer vaccine composition of claim 18, wherein the adjuvant is a TLR9 agonist.

20. 20. The cancer vaccine composition of claim 19, wherein the adjuvant comprises a cationic liposome-DNA complex (CLDC).

21. 20. The cancer vaccine composition of claim 19, wherein the adjuvant comprises a CpG oligodeoxynucleotide (ODN).

22. The cancer vaccine composition of any one of claims 17 to 21, wherein the adjuvant modifies monocyte function.

23. The cancer vaccine composition of any one of claims 1 to 22 for use as a medicament for treating cancer in a subject in need thereof.

24. 24. The cancer vaccine composition for use according to claim 23, wherein the cancer is breast cancer, lung cancer, liver cancer, bladder cancer, gynecological cancer, brain cancer, stomach cancer, prostate cancer, skin cancer, thyroid cancer, pancreatic cancer, colon cancer, or blood cancer.

25. 25. The cancer vaccine composition for use according to claim 24, wherein the cancer is a gynecological cancer.

26. The cancer vaccine composition for use according to any one of claims 23 to 25, wherein the cancer is a metastatic cancer.

27. The cancer vaccine composition for use according to any one of claims 23 to 26, wherein the inactivated cancer cells are autologous.

28. 28. The cancer vaccine composition for use according to any one of claims 23 to 27, wherein the cancer vaccine composition is formulated for subcutaneous, intravenous, or intramuscular injection.

29. The cancer vaccine composition for use according to any one of claims 23 to 28, wherein the cancer vaccine composition is for a single administration to the subject.

30. 29. The cancer vaccine composition for use according to any one of claims 23 to 28, wherein the cancer vaccine composition is for administration to the subject 2, 3, 4, 5, 6, 7, 8, 9, or 10 times.

31. 31. The cancer vaccine composition for use according to any one of claims 23 to 30, wherein the cancer vaccine composition is for administration at least once every 7 days, at least once every 14 days, or at least once every 6 months.

32. 32. The cancer vaccine composition for use according to any one of claims 23 to 31, wherein the adjuvant is administered after administration of the cancer vaccine composition, and wherein the adjuvant modifies monocyte function.

33. The cancer vaccine composition for use according to any one of claims 23 to 32, wherein the subject is a human.

34. 1. An in vitro method for producing a cancer vaccine comprising viable, replication-incompetent cells, the method comprising: (1) pre-incubating cancer cells in a solution containing riboflavin, wherein the solution contains 1 to 50 μM riboflavin; (2) inactivating the cancer cells by contacting the cancer cells with UV light in the presence of riboflavin, wherein the UV light comprises a wavelength of 310-320 nm, and the UV light is at a dose of about 0.5 Joules / ml to about 3 Joules / ml, or about 200 Joules to about 600 Joules, and the UV light selectively oxidizes guanine bases in DNA of the cancer cells; A method comprising:

35. 35. The method of claim 34, wherein the solution contains about 50 μM riboflavin.

36. The cancer cells are about 1×10 ５ ~Approx. 1×10 ８ 36. The method of claim 34 or 35, wherein the concentration is 100%.

37. The cancer cells are about 1×10 ５ ~Approx. 1×10 ７ 36. The method of claim 34 or 35, wherein the concentration is 100%.

38. 38. The method of any one of claims 34 to 37, wherein the cancer cells are contacted with the UV light in the presence of the riboflavin for about 1 minute to about 3 minutes.

39. 39. The method of any one of claims 34 to 38, wherein the UV light is at a dosage of about 200 joules to about 400 joules, or about 200 joules to about 380 joules, or about 0.6 joules / ml to 2 joules / ml.

40. 40. The method of claim 39, wherein the UV light is at a dose of about 300 joules, or about 1 joule ml.

41. 41. The method of any one of claims 34 to 40, wherein the inactivated cancer cells are viable for at least 1 day, at least 2 days, at least 3 days, at least 4 days, or at least 5 days after treatment with UV light.

42. 42. The method of any one of claims 34 to 41, wherein the cancer cells are breast cancer cells, lung cancer cells, liver cancer cells, bladder cancer cells, gynecological cancer cells, brain cancer cells, gastric cancer cells, prostate cancer cells, skin cancer cells, thyroid cancer cells, pancreatic cancer cells, colon cancer cells, or blood cancer cells.

43. 43. The method of any one of claims 34 to 42, wherein the inactivation does not substantially alter the expression levels of EpCAM, CD38, CD34, CD117, CD44, CD24, Sca1, HLA, Glut1, MHC class I, PD-L1, CD45, gp70, and / or CD90 on the cancer cells, and / or wherein the inactivation does not substantially alter the structure of one or more antigen proteins on the cancer cells.

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