Compositions and methods for treating plasma cell disorders, including multiple myeloma, with vaccine compositions and myeloma-specific CAR-T cells

Abstract

Described are multiple myeloma-specific CAR+ T cell compositions and vaccine compositions comprised of three cell lines, U266, H929, and K562. Methods are described for using the vaccine compositions in conjunction with MM-specific CAR+ T cell compositions in methods of immunizing against plasma cell disorders, including multiple myeloma and related disorders.

Description

[Technical field]

[0001] The present invention relates to compositions and methods useful for vaccination against and treatment of plasma cell disorders, including multiple myeloma (MM), using combination therapy with MM-specific chimeric antigen receptor T cells. [Background technology]

[0002] The emergence of novel therapeutic agents with low toxicity and high tumor specificity has significantly improved the clinical outcomes of patients with multiple myeloma (MM). In particular, triple combinations of proteasome inhibitors, immunomodulatory drugs (IMiDs), and steroids have led to improved response rates and more durable remissions (Attal et al.; N. Engl. J. Med. (2017) 376:1311-20;Durie et al.; Lancet (2017) 389:519-27). This translates into improved progression-free survival (PFS) and overall survival (OS) (Richardson et al.; Hematology (2014) 255-61). However, a significant proportion of patients eventually develop resistance to these agents and relapse. Approaches aimed at deepening and prolonging such responses include long-term maintenance therapy with or without consolidation regimens. Notably, lenalidomide (Len) maintenance in both transplant and non-transplant settings has shown significant clinical benefit from such approaches (McCarthy et al.; J Clin Oncol (2017) 35:3279-89). With the introduction of more effective treatments for MM, much of the overall improvement in clinical outcomes may be due to the ability to achieve deeper responses. Indeed, minimal residual disease (MRD) assessment by either flow cytometry or next-generation sequencing can currently detect as few as one cell in a million (ClonoSEQ Cleared for Residual Cancer Testing. Cancer Discov (2018) 8:OF6- OF6).

[0003] A growing body of literature has demonstrated a strong correlation between depth of response and improved clinical outcomes, to the point that MRD negativity is now considered a potentially acceptable clinical trial endpoint (Perrot et al.; Blood (2018) 132:2456-64). However, definitive clinical guidance and safe treatment options for the management of patients with MRD-positive MM are desperately needed in the art. Immunotherapy exploits the immune system's ability to specifically recognize and eliminate cancer cells. Indeed, immune checkpoint blockade (Hargadon et al.; Int. Immunopharmacol. (2018) 62:29-39) and genetically modified T cells with chimeric antigen receptors (CAR-T) (Raje et al.; N. Engl. J. Med. (2019) 380:1726-37) have demonstrated clinical efficacy in hematological malignancies and, more specifically, solid tumors. In contrast, cancer vaccines to date have not shown similar benefits (Hu et al.; Nat. Rev. Immunol. (2018) 18:168-82).

[0004] Chimeric antigen receptor T-cell therapy is considered a potential treatment for hematological cancers, including leukemia, lymphoma, and multiple myeloma. B-cell maturation antigen (BCMA) is a member of the tumor necrosis factor superfamily of proteins primarily expressed by malignant and normal plasma cells. A G protein-coupled receptor, group 6 member D, normally expressed in hair follicles, has been identified as having mRNA expression in bone marrow aspirates from MM patients, with a similar distribution to BCMA but expressed independently on CD138+ cells (Smith et al., Sci Transl. Med. 11:485 (2019)). The generation of a productive vaccine-specific immune response depends on the diversity and abundance of tumor-associated antigens, effective adjuvants, and concomitant immunostimulatory therapies. The established cell line, K562, is genetically modified to produce granulocyte-macrophage colony-stimulating factor (GM-CSF), a key immune stimulatory factor known to improve efficient antigen presentation (Borrello et al.; Hum. Gene Ther. (1999) 10:1983-91;Borrello et al.; Cytokine Growth Factor Rev. (2002) 13:185-93).

[0005] Although several factors such as poor tumor antigen selection, choice of vaccine adjuvant, and absence of concomitant immunomodulatory therapy may contribute to the lack of clinical efficacy, disease burden may have a significant impact on clinical outcomes. Although the ability of lenalidomide to increase vaccine-specific cellular and humoral immunity has been demonstrated (Noonan et al.; Clin. Cancer Res. (2012) 18:1426-34), in MRD-positive multiple myeloma, meaning that the disease burden is very low, multiple myeloma vaccination in combination with lenalidomide has not been found to be a potent therapeutic approach to enhance treatment efficacy without additional toxicity. Effective therapies for the treatment of MRD-positive MM and other plasma cell disorders are currently lacking, and there is no consensus on the management of this ever-increasing patient population. Herein, we describe a treatment regimen using an MM-specific vaccine in conjunction with an MM-specific CAR-T cell composition in successfully treated patients with minimal disease burden to not only improve disease response but also prevent disease progression. Summary of the Invention

[0006] According to a first aspect of the present invention, there is provided a composition for use in raising an immune response against a plasma cell disorder in a subject, the composition comprising an effective amount of multiple myeloma-specific CAR+ T cells. According to another aspect of the present invention, there is provided the above composition, wherein the multiple myeloma-specific CAR+ T cells are BCMA-specific CAR+ T cells or GPRC5D-specific CAR+ T cells. According to another aspect of the present invention, there is provided a composition as described above, which is allogeneic. According to another aspect of the present invention, there is provided a composition as described above, which when administered to said subject induces an immune response in the subject. According to another aspect of the present invention, there is provided a composition as described above, wherein the immune response induces a complete remission of said plasma cell disorder in the subject. According to another aspect of the present invention, there is provided a composition as described above, which extends progression-free survival in said subject. According to another aspect of the present invention, there is provided the above composition, wherein said complete response is determined as undetectable M spikes and positive immunofixation electrophoresis. According to another aspect of the present invention, there is provided the above composition, wherein the subject is a human. According to another aspect of the invention, there is provided a method of treating a plasma cell disorder in a subject, comprising administering to the subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells.

[0007] According to another aspect of the present invention, there is provided the above method, wherein said administering step also comprises providing an immunomodulatory drug to said subject. According to another aspect of the present invention, there is provided the above method, wherein said immunomodulatory agent is lenolidomide. According to another aspect of the present invention, there is provided the above method, wherein said immunomodulatory agent is provided to said subject before, during and / or after said administering step. According to another aspect of the present invention, there is provided the above method, wherein the vaccine composition is allogeneic. According to another aspect of the present invention, there is provided the above method, wherein the K562 cells express the GM-CSF gene. According to another aspect of the present invention, there is provided the above method, wherein the K562 cells are transfected with a gene encoding GM-CSF. According to another aspect of the present invention, the GM-CSF gene is up to about 1500 ng / 1×10 6 The above method is provided, wherein the amount of cells capable of expressing GM-CSF is 0.1 to 10% by volume. According to another embodiment of the present invention, the GM-CSF gene is about 35 to 1200 ng / 1×10 6 The above method is provided, wherein the amount of cells capable of expressing GM-CSF is 0.1 to 10% by volume.

[0008] According to another aspect of the present invention there is provided a method as described above, wherein the amount of GM-CSF produced is on average every 24 hours. According to another aspect of the present invention, there is provided the above method, wherein the GM-CSF is of human origin. According to another aspect of the present invention, there is provided the above method, wherein the ratio of the combination of U266 and H929 cells to K562 cells is about 20:1. According to another aspect of the present invention, there is provided the above method, wherein the dose of said composition is such that the ratio of tumor cells in said subject to K562 cells in said vaccine composition is greater than 2:1. According to another aspect of the present invention, there is provided a method as described above, wherein U266 and H929 cells are present in said vaccine composition in equal amounts. According to another aspect of the present invention, the U266 and H929 cells are about 5×10 7 and K562 cells are present in the vaccine composition in an amount of about 5×10 6 The method according to the invention provides a method as described above, wherein the composition is present in an amount of cells.

[0009] According to another aspect of the present invention, there is provided the above method, wherein a near complete remission or complete remission is achieved in said subject. According to another aspect of the present invention, there is provided the above method, wherein said complete remission is sustained in said subject for up to five years. According to another aspect of the present invention, there is provided the above method, wherein said complete remission is determined by measuring undetectable monoclonal spike and negative immunofixation electrophoresis. According to another aspect of the present invention, there is provided the above method, wherein said subject is positive for minimal residual disease. According to another aspect of the present invention, there is provided the above method, wherein said composition minimizes a non-specific immune response in the subject. According to another aspect of the present invention, there is provided the above method, wherein the vaccine composition is administered prior to the CAR+ T cell composition. According to another aspect of the invention, there is provided the above method, wherein the CAR+ T cell composition is administered prior to the vaccine composition. According to another aspect of the invention, there is provided the above method, wherein the vaccine composition is administered, followed by the CAR+ T cell composition, followed by a second dose of the vaccine composition.

[0010] According to another aspect of the present invention, there is provided the above method, wherein 1 to 5 doses of the vaccine composition are administered to the subject, with an interval of more than 1 day between each dose. According to another aspect of the present invention, there is provided the above method, wherein the vaccine composition is administered in two to four doses, with an interval of more than two weeks between each dose. According to another aspect of the present invention, there is provided the above method, wherein the vaccine composition is administered in two to four doses, with an interval of more than four weeks between each dose. According to another aspect of the present invention, there is provided the above method, wherein four doses are administered with an interval of about one month between each dose. According to another aspect of the present invention, there is provided the above method, wherein the first three administrations are equally spaced. According to another aspect of the present invention, there is provided the above method, wherein all doses are administered within one year of each other. According to another aspect of the present invention, there is provided the above method, wherein at least one dose of the vaccine composition is administered between days 7 and 18, inclusive, from the start of the lenalidomide course. According to another aspect of the present invention, there is provided the above method, wherein at least one dose is administered about 15 days after the start of the lenalidomide course.

[0011] According to another aspect of the present invention, there is provided the above method, wherein said plasma cell disorder is selected from the group consisting of MGUS, SMM, multiple myeloma, non-secretory multiple myeloma, indolent myeloma, light chain myeloma, plasma cell leukemia, and primary amyloidosis. According to another aspect of the present invention, there is provided the above method, wherein said plasma cell disorder is multiple myeloma. According to another aspect of the invention, there is provided a method of extending progression-free survival in a subject with multiple myeloma, comprising administering to the subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells. According to another aspect of the invention, there is provided a method of inducing clonal T cell expansion and increased myeloma-specific cytokine responses in a subject with multiple myeloma, comprising administering to the subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells.

[0012] According to another aspect of the present invention, there is provided the above method, wherein said increase in said subject is sustained for up to 7 years following said administering step. According to another aspect of the present invention, there is provided the above method, wherein said increase in said subject is sustained for up to 5 years following said administering step. According to another aspect of the invention, there is provided a method of inducing multiple myeloma-specific immunity in a subject, comprising administering to the subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells. According to another aspect of the present invention, there is provided the above method, wherein said subject is positive for minimal residual disease at the time of said administering step. According to another aspect of the invention, there is provided a method of preventing recurrence of multiple myeloma in a subject, comprising administering to the subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells.

[0013] According to another aspect of the present invention, there is provided the above method, wherein the subject is positive for minimal residual disease at the time of said administering step. According to another aspect of the present invention, there is provided the above method, wherein the subject is a human. According to another aspect of the present invention, there is provided the above method, wherein the multiple myeloma-specific CAR+ T cells are GPRC5D multiple myeloma-specific CAR+ T cells and / or BCMA-specific CAR+ T cells. Still other objects, features and attendant advantages of the present invention will become apparent to those skilled in the art upon reading and understanding the following detailed description of the embodiments made in accordance with the present invention, when taken in conjunction with the accompanying drawings. The invention of the present application will now be described in more detail with reference to exemplary embodiments of compositions and methods, given by way of example only, and with reference to the accompanying drawings, in which: [Brief description of the drawings]

[0014] [Figure 1] Schematic of the clinical trial. Patients received four doses of vaccine at the indicated time points (arrows) while maintaining Len. "*" indicates immune monitoring time points. [Diagram 2] FIG. 1 shows the frequency of T cell clones in the blood and bone marrow of all patients expanded in C3D14 and followed over time. [Diagram 3] Representative paired scatter plots from two patients showing clonal expansion of pre-existing T cell clones after vaccination as well as recruitment of novel clonotypes not previously present in either PB or BM. [Figure 4] Representative paired scatter plots comparing fold changes in frequency of expanded T cell clones in PB and BM. [Diagram 5] Figure 1 shows data depicting the change in Morishita index, which quantifies the degree of similarity between BM and PB T cell repertoires before, during (C3D14), and after vaccination. TCR = T cell receptor. [Figure 6]FIG. 1 shows representative plots showing IFNγ and TNFα production before, during (C3D14), and after vaccination in both CD8+ and CD4+ T cell compartments. [Figure 7] FIG. 1 shows that cytokine production was elevated in all patients after vaccination and was maintained for more than 4 years (p<0.0001 for both CD8+ and CD4+ compartments). [Figure 8] FIG. 1 shows box plots showing the frequency of occurrence of each individual cluster across patients and time points. [Figure 9] FIG. 1 shows T cell clones expanded after vaccination and followed up to 7 years after MM-GVAX administration in both PB and BM. [Figure 10] Representative plots showing IFNγ and TNFα production upon in vitro antigen stimulation of BM from vaccinated patients at the indicated long-term follow-up time points. *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001. [Figure 11] FIG. 1 shows that the frequency of CD69+ T cells is significantly higher in the CD8+ subset (p<0.001). [Figure 12] FIG. 13. Representative dot plots and histograms showing the classical phenotype of CD69+ BM T cells. [Figure 13] Representative histograms showing the expression of various markers on CD69+ (red) and CD69- (light blue) BM T cells. *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001. [Figure 14] FIG. 1 shows box plots depicting the relative abundance of the eight FlowSOM metaclusters in the two groups (relapsers and responders). [Figure 15]Representative dot plots showing manual gating analysis of DNAM1- / lowCD27-CD8+ T cells (left) and a summary of the frequency of this CD8+ T cell subset in both groups. *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0015] Described herein is an allogeneic whole cell GM-CSF secreting multiple myeloma (MM) vaccine (MM-GVAX) and methods of administering this vaccine in combination with lenalidomide, also referred to as "Len", and MM-specific CAR+ T cells to MM patients with minimal residual disease burden, defined as undetectable monoclonal spike but positive immunofixation electrophoresis (IFE), demonstrating eradication of residual disease and conversion to complete remission (CR). Vaccine / Len / CAR combination therapy has the potential to be effective against other plasma cell disorders with detectable or undetectable monoclonal spike proteins. Also described are the safety, response time, and immune monitoring of the vaccine and MM-specific T cell responses. To our knowledge, this is the first description of treating patients with minimal disease burden with MM-specific CAR+ in combination with MM-GVAX to attempt to prevent disease progression as well as further improve disease response.

[0016] As used herein, and unless otherwise indicated, the term "about" is intended to mean ±5% of the value it modifies. Thus, "about 100" means 95 to 105. In addition, the term "about" modifies a term in a series of terms, such as "about 1, 2, 3, 4, or 5." The term "about" should be understood to modify each member of a list, such that "about 1, 2, 3, 4, or 5" can be understood to mean "about 1, about 2, about 3, about 4, or about 5." The same applies to lists modified by the term "at least" or other quantitative modifiers, such as, but not limited to, "less than," "greater than," etc. As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise.

[0017] As used herein, the terms "comprising" (and all forms of comprising, e.g., "comprise," "comprises," and "comprised"), "having" (and all forms of having, e.g., "have" and "has"), "including" (and all forms of including, e.g., "includes" and "include"), or "containing" (and all forms of containing, e.g., "contains" and "contain") are inclusive or open-ended and do not exclude further, unrecited elements or method steps. As used herein, the terms "treat," "treated," or "treating" refer to both therapeutic treatments in which the objective is the slowing (reduction) of an undesired physiological condition, disorder, or disease, or the attainment of a beneficial or desired clinical outcome. For purposes of the embodiments described herein, beneficial or desired clinical outcomes include, but are not limited to, alleviation of symptoms; reduction in the extent of the condition, disorder, or disease; stabilization (i.e., non-worsening) of the condition of the condition, disorder, or disease; delay in the onset or slowing of progression of the condition, disorder, or disease; amelioration or remission (partial or complete), detectable or undetectable, of the condition of the condition, disorder, or disease; amelioration of at least one measurable physical parameter, not necessarily discernible by the patient; or enhancement or amelioration of the condition, disorder, or disease. Thus, "treatment of cancer" or "cancer therapy" or "treatment of multiple myeloma" or "multiple myeloma" treatment or "treatment of plasma cell disorder" or "plasma cell disorder treatment" refers to activity that reduces or ameliorates either the primary event or the secondary symptoms or symptoms associated with cancer, multiple myeloma, or any other condition described herein. In some embodiments, the cancer being treated is one of the cancers listed herein. In one embodiment, the cancer is multiple myeloma.

[0018] As used herein, the term "subject" may be used interchangeably with the term "patient." The subject may be a mammal, such as, for example, a dog, cat, monkey, horse, or cow. In some embodiments, the subject is a human. In some embodiments, the subject has been diagnosed with a blood cancer. In some embodiments, the subject has been diagnosed with multiple myeloma. In some embodiments, the subject is suspected of having multiple myeloma. As used herein, the term "expressing" when referring to cell surface receptors, such as, but not limited to, CD3, CD4, and CD8, can also refer to cells that are positive for those markers. For example, a cell that expresses CD3 is said to be CD3 positive (CD3 + ) cells. As used herein, the term "express" can refer to a gene that is located within a cell as part of chromosomal DNA or on some other vector. A cell "expresses" a gene when it is induced to produce the protein that the gene encodes. The protein that is produced can be contained within the cell or transported outside the cell.

[0019] As used herein, the term "vaccine" refers to a product or composition that stimulates a subject's immune system to provide immunity to a particular disease or condition, thereby protecting the subject from that disease or condition. A vaccine may be part of a composition, which may or may not include other components, including but not limited to an adjuvant. As used herein, the term "adjuvant" refers to a component that modifies the action of a primary component, e.g., a vaccine. When used in a vaccine composition, an adjuvant can help generate a stronger immune response in a subject to which the vaccine composition is administered. The term "cancer," as used herein, is defined as a disease characterized by the rapid and uncontrolled growth of abnormal cells. Cancer cells can spread locally or through the bloodstream or lymphatic system to other parts of the body.

[0020] The term "multiple myeloma" as used herein is defined as a cancer that arises in white blood cells. In some embodiments, the white blood cells are white blood cells in the bone marrow. In some embodiments, multiple myeloma arises in plasma cells. The term "plasma cell disorder", as used herein, is defined as a disorder characterized by elevated serum levels of a monoclonal immunoglobulin protein, also called "M protein" or "M spike", or elevated serum levels of bone marrow plasma cells. The term "antigen" as used herein is defined as a molecule that elicits an immune response. The immune response may include either the production of antibodies or the activation of specific immunocompetent cells, or both. Those skilled in the art will appreciate that any macromolecule, including virtually any protein or peptide, can act as an antigen. Moreover, antigens can be derived from recombinant or genomic DNA. Those skilled in the art will appreciate that any DNA that includes a nucleotide sequence or a portion of a nucleotide sequence that encodes a protein will elicit an immune response, thereby encoding an "antigen," as the term is used herein. Those skilled in the art will further appreciate that an antigen need not be encoded exclusively by a full-length nucleotide sequence of a gene. It will be readily apparent that embodiments include, but are not limited to, the use of portions of the nucleotide sequences of two or more genes, and further that such nucleotide sequences may be configured in various combinations to elicit a desired immune response. Moreover, those skilled in the art will appreciate that an antigen need not be encoded by a "gene" at all. It will be readily apparent that an antigen may be synthesized or obtained from a biological sample. Such biological samples may include, but are not limited to, tissue samples, tumor samples, cells or biological fluids.

[0021] The term "anti-tumor effect" as used herein refers to a biological effect that may be manifested by a reduction in tumor burden, a reduction in tumor cell count, a reduction in the number of metastases, an increase in life expectancy, or an amelioration of various physiological symptoms associated with a cancer condition. An "anti-tumor effect" may also be manifested by the ability of peptides, polynucleotides, cells, and antibodies to prevent tumor initiation. As used herein, the term "autologous" is intended to refer to any material originating from the same individual that is subsequently reintroduced into that individual. The term "chimeric antigen receptor" or "CAR" as used herein is defined as a molecule that combines antibody-based specificity for a desired antigen with a T cell receptor activating intracellular domain to produce a chimeric protein that exhibits specific anti-tumor cell-mediated immune activity.

[0022] The term "B cell maturation antigen" or "BCMA" refers to a protein that is a member of the tumor necrosis factor ("TNF") receptor superfamily, also called TNF receptor superfamily member 17, or "TNFRSF17." TNFRSF17 is a cell surface receptor that recognizes B cell activating factor. It has been found to be preferentially expressed in mature B lymphocytes and may be important in B cell development and autoimmune responses, as well as cell survival and proliferation. The term "G protein-coupled receptor 5D" or "GPRC5D" refers to an orphan G protein receptor that is normally expressed in hair follicles, but has been found to be expressed in the bone marrow of patients with multiple myeloma. "Effective amount" or "therapeutically effective amount," as used interchangeably herein, refer to an amount of a compound, formulation, substance, or composition described herein that is effective to achieve a particular biological result. Such result may include, but is not limited to, inhibition of cancer cell proliferation as determined by any means suitable in the art.

[0023] As used herein, GM-CSF refers to granulocyte-macrophage colony-stimulating factor, which is a known protein that is often used in cancer treatment. Initial studies with GM-CSF, also called "Leukine" in the literature, involved stimulating the proliferation of macrophages and neutrophils to reduce the hematopoietic toxicity associated with dose-intensive chemotherapy. When transfected into tumor cells and administered as a vaccine, the GM-CSF gene demonstrated tumor regression and prolonged survival in both animal models and early clinical trials (Nemunaitis, Expert Rev Vaccines; 2005; 4(3): 259-74). As used herein, the term "GVAX" refers to a cancer vaccine composed of whole tumor cells genetically modified to secrete the immune stimulating cytokine GM-CSF. One or more cell types may be included in the GVAX vaccine. One example is MM-GVAX, or a whole cell cancer vaccine for the treatment of multiple myeloma. As used herein, the term "immunomodulatory drug", also referred to as "IMiD", may refer to drugs that modify the immune system's response by increasing or decreasing serum antibody production. Immunostimulators may enhance the immune response to infectious diseases, tumors, and primary or secondary immune deficiencies. Immunosuppressants are used to decrease the immune response to transplanted organs and treat autoimmune diseases.

[0024] As used herein, the term "lenalidomide," also known by the trademark Revlimid, refers to an immunomodulatory drug used to treat multiple myeloma and myelodysplastic syndromes (MDS), which may be administered alone or with steroids, including but not limited to dexamethasone. As used herein, the term "minimal residual disease" or "MRD" refers to a small number of cancer cells remaining in the body after treatment. The number of remaining cells may be so small that they do not cause any physical signs or symptoms and are often impossible to detect by traditional methods, such as looking at the cells under a microscope and / or tracking abnormal serum proteins in the blood. A positive test result for MRD means that residual (residual) disease has been detected. A negative result means that no residual disease has been detected. As described herein, MRD is used to measure the effectiveness of treatment and predict patients at risk of recurrence. If a patient tests positive for MRD, it means that there are still residual cancer cells in the body after treatment. If MRD is detected, it is known as "MRD positive". If a patient tests negative, it means that no residual cancer cells were found. If MRD is not detected, it is known as "MRD negative".

[0025] Subjects who are candidates for receiving the vaccine compositions and CAR+T cells described herein may also have been or are currently receiving immunomodulatory agents, including but not limited to thalidomide, lenalidomide and pomalidomide, and proteasome inhibitors, including but not limited to bortezomib, carfilzomib and ixazomib. Subjects who are candidates for receiving the vaccine compositions described herein may have a plasma cell disorder. Subjects with a plasma cell disorder may have elevated serum levels of M spike protein, or "M spike", although this is not always the case, and such subjects may also be identified by the presence of a certain amount of bone marrow plasma cells in the serum at the time of diagnosis. Subjects with plasma cell disorders include, but are not limited to, subjects diagnosed with monoclonal gammopathy of undetermined significance ("MGUS"); multiple myeloma ("MM"), including smoldering myeloma ("SMM"), non-secretory multiple myeloma, indolent myeloma, and light chain myeloma; plasma cell leukemia, including basal cell leukemia; and primary amyloidosis. Next generation sequencing (NGS) technologies as well as flow cytometry have enabled quantification of "minimal residual disease" (MRD) burden in patients with MM. MRD negativity is a major prognostic factor for MM and is now even more achievable, making it a measurable endpoint for testing and treatment. However, guidance is currently lacking on how to optimally treat MM patients who are in complete remission (CR) but are still MRD positive. The vaccine compositions described herein provide a specific immune response against MM plasma cells that eradicates the minimal residual disease burden, which can translate into clinically meaningful outcomes.

[0026] The allogeneic GM-CSF-producing MM vaccine (MM-GVAX) described herein may comprise three or more distinct cell lines, including, but not limited to, the known xenogeneic MM cell lines H929 and U266, both publicly available from cell line depositories, e.g., ATCC (Manassas, VA; ATCC.org), and K562 cells, also publicly available. The K562 cell line can be transfected or transformed with a gene encoding GM-CSF in an expressible configuration. Expression constructs that can be used include those that contain typical known components, such as promoter, operator, origin of replication, etc., operably linked to the GM-CSF coding sequence, that allow optimal expression in the host cell.

[0027] The amount of cells of each cell line in the vaccine composition is not limited, and may be equal or unequal amounts of each cell line relative to each other. The ratio of H929 and U266 may be 1:1, but is not limited to this ratio and may be present in unequal amounts. The ratio of the amount of H929 / U266 mixed cells to K562 / GM-CSF may be about 40:1 to K562 / GM-CSF, or may be about 35:1, 30:1, 25:1, 20:1, 15:1, or 10:1. In one embodiment, the ratio is about 20:1. Regardless of the ratio of cell lines, in one embodiment, the ratio is about 50-1500 ng / 1×10 6 cells / 24 hr GM-CSF present. The absolute amount of cells present in the vaccine was approximately 1 × 10 for H929 and U266 cells, respectively. 7 ~Approx. 1×10 9 The cells may be 1, 5, 10, 50, or 100×10 7 In one embodiment, the composition comprises 5×10 each of H929 and U266 cells. 7 The K562 / GM-CSF cells were approximately 1 × 10 4 ~Approx. 1×10 7 The amount of cells may be 1, 5, 10, 50, or 100×10 7 In some embodiments, the composition comprises 1×10 6 This includes cases where the amount of K562 / GF-CSF cells is 100%.

[0028] The vaccine composition may contain components other than the three or more cell lines, including, but not limited to, other cell lines, adjuvants such as aluminum, e.g., aluminum hydroxide, aluminum phosphate, and aluminum potassium sulfate; squalene oil, e.g., MF59; preservatives, e.g., thiomersal or thimerosal; stabilizers, e.g., gelatin, sorbitol, sucrose, lactose, mannitol, glycerol, 199 medium, arginine hydrochloride, sodium glutamate, and urea; and emulsifiers, e.g., polysorbate 80, sorbitan trioleate, and sodium citrate. Other components commonly used in vaccine production may be present and may include antibiotics, ovalbumin, yeast proteins, latex, formaldehyde, glutaraldehyde; and regulators such as acidity regulators, e.g., sodium and / or potassium based salts, disodium adipate, succinic acid, sodium hydroxide, histidine, sodium borate, trometamol, and human serum albumin. Such components may be included in amounts typical of vaccine formulations, and may be in any amount, although typically very small amounts, so long as the presence of the component does not negatively affect the efficacy of the vaccine. Human serum albumin is typically used at 0-10%. CAR comprises an extracellular domain having an antigen recognition domain, a transmembrane domain, and a cytoplasmic cell signaling domain. In some embodiments, a transmembrane domain that naturally binds to one of the domains of the CAR is used. In some embodiments, the transmembrane domain can be selected or modified by amino acid substitution to avoid such domains binding to the transmembrane domain of the same or different surface membrane proteins and minimize interaction with other members of the receptor complex. For example, the transmembrane domain can be the CD8α hinge domain.

[0029] A multiple myeloma-specific CAR may comprise an extracellular ligand-binding domain that binds to BCMA, GPRC5D, or other multiple myeloma-specific antigen; a transmembrane domain; a 4-1BB costimulatory signaling domain; and an intracellular CD3ζ signaling domain. The transmembrane domain may be the transmembrane domain of CD3ζ, CD4, CD8, or CD28. With respect to the cytoplasmic domain, for example, the CAR may be designed to have the CD28 and / or 4-1BB signaling domain itself, or may be combined with any other desired cytoplasmic domain useful in the context of the CAR. In some embodiments, the cytoplasmic domain of the CAR may be designed to further comprise the signaling domain of CD3ζ. For example, the cytoplasmic domain of the CAR may comprise, but is not limited to, CD3ζ, 4-1BB, and CD28 signaling modules, and combinations thereof. The CAR is expressed in the patient's T cells, and the cells are formulated for administration to the patient.

[0030] Multiple myeloma-specific CAR+ compositions for administration with the vaccines described herein can be generated as follows: Autologous peripheral blood mononuclear cells are transduced with a lentiviral vector containing an anti-multiple myeloma CAR, stimulated with antibodies to CD3 and CD28, and allowed to expand for a period of time, e.g., 8-10 days (see Friedman et al., Hum Gene Ther 2018; 29:585-601). Expression of multiple myeloma-specific CARs can be confirmed by methods known in the art. The compositions may be referred to as multiple myeloma-specific CAR+ T cells or MM-specific CAR+ T cells. The amount of MM-specific CAR+ T cell composition that can be administered to a subject is 50×10 6 , 150×10 6 , 450×10 6 , 800×10 6 Doses of cells, each dose having a variance of plus or minus 20%, including all doses in between. For further dosing information and dosing results of CAR+ T cell efficacy in MM, see Raje et al. NEJM 2019; 380:1726-37. The allogeneic GM-CSF-producing MM vaccine (MM-GVAX) described herein may be administered to subjects diagnosed with a plasma cell disorder, such as multiple myeloma (MM). Candidate MM patients may be MRD positive or negative. Candidate MM patients have the potential for low disease burden. Candidate MM patients have the potential for achieving stable near CR remission (nCR), defined as the absence of M spikes in either serum or urine for at least 4 months and IFE positivity. The rate of conversion from nCR to true CR was 53.3%, with 8 patients achieving improved clinical response within a mean time of 11.6 months from enrollment.

[0031] The allogeneic GM-CSF-producing MM vaccine (MM-GVAX) and MM-specific CAR+T cell compositions described herein may be administered as part of a number of different treatment regimens. In one example of an administration regimen, MM-GVAX is administered prior to harvesting of T cells, and then T cells are harvested from the patient. These are then used to generate MM-spiked CARs described herein, and then the MM-specific CAR+T cell composition is administered, followed finally by a second administration of MM-GVAX. In another example of an administration regimen, T cells are harvested from the patient to generate MM-specific CARs, the MM-specific CAR+T cells are administered to achieve remission in the patient, and then MM-GVAX is administered one or more times to enhance the long-term durability of the duration of remission. The time from CAR+ T cell administration to GVAX administration can be any time from 2 months up to 2 years after administration of the CAR T cells, including, but not limited to, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, and 24 months and all time points in between. Patients eligible for MM-GVAX after CAR T cell administration include those who are MRD- or those who are MRD+ but in remission or have undetectable stable disease.

[0032] The method of administration of the vaccine compositions and MM-specific CAR+ T cell compositions described herein is not particularly limited and may include oral, subcutaneous, intramuscular, intradermal, intranasal, or intravenous routes. The intravenous route is a particular example. The composition may be administered once, twice, three times, four times, or five or more times. The time between administrations of the doses of the vaccine compositions described herein is not limited and may be any time between one week and four months, for example, two weeks, three weeks, four weeks, one month, two months, three months, and any time between each administration. The time between multiple administrations may not be the same. In a particular example, there is one month between administrations. In an example administration schedule, all vaccine doses are administered within a period of time including up to one year, 11 months, 10 months, 9 months, 8 months, 7 months, 6 months, or 5 months, and all time points in between. The CAR T cells may be administered between any of the vaccine administrations or prior to administration of any vaccine.

[0033] Several factors influence the generation of active tumor-specific T cells as a result of administration of a vaccine and MM-specific CAR+ T cells, in particular the presence of appropriate antigen, effective antigen presentation, inhibition of the suppressive tumor microenvironment, and expression of appropriate chemokines all facilitate efficient trafficking of T cells to the tumor site. The vaccine compositions described herein take into account several of these key components. First, the vaccine composition described herein acts as a source of tumor-associated antigens (TAA) due to the presence of two established heterogeneous MM cell lines, H929 and U266. Such cell lines exhibit diverse patterns of somatic mutations that are often associated with high risk and relapse of MM. In particular, H929 harbors t(4;14) translocation and NRAS mutations, while U266 has several mutations including BRAF and TP53 pathways (Moreaux et al.; Haematologica; 2011;96:574-82). It has been shown that disease relapse occurs as a result of clonal evolution, which can lead to further aggressive genetic mutations. The vaccine composition described herein was designed to stimulate the immune system against some of these putatively high-risk antigens before they emerge in the process of clonal evolution associated with disease progression. Presentation of such high-risk antigens by the vaccine composition described herein is shown to significantly affect the timing and / or aggressiveness of disease relapse.

[0034] Second, the vaccine compositions described herein may include two unmodified MM cell lines, H929 and U266, as well as a genetically modified bystander GM-CSF-secreting cell line, K562 / GM-CSF. The GM-CSF gene used to transfect the K562 cells may be derived from any source, including but not limited to human. "Derived from," as used herein, may mean native to how or where GM-CSF occurs in nature. GM-CSF has been shown to be a key immune adjuvant. Importantly, the use of the K562 / GM-CSF cell line allows for titration of the amount of GM-CSF, thereby allowing delivery of an optimal dose in the vaccine compositions described herein. This GM-CSF dose is neither insufficient nor over-therapeutic, so that high doses of antigen could still be delivered but with reduced efficacy due to induction of myeloid-derived suppressor cells (MDSCs). It has been shown that a "therapeutic" dose of GM-CSF and sufficient amounts of antigen are required for an effective vaccine (Serafini et al.; Cancer Res. 2004; 64:6337-43). As described herein, K562 cells are administered at a dose of about 50 ng to about 1500 ng / 1×10 6 The amount of GM-CSF can be expressed per cell / 24 hours. The period during which GM-CSF can be produced can be up to about 72 hours, as measured by ELISA, but can be more or less than this, as necessary to maintain an effective amount of the vaccine composition. The amount can be produced on average every 24 hours. It is also required that the antigen cell source, i.e., tumor cells, be present in excess, such that the stoichiometry of tumor cells:bystander cells is at least greater than 2:1. The amount of GM-CSF can be measured by any known method, including, but not limited to, enzyme-linked immunosorbent assay ("ELISA").

[0035] The vaccine composition can be irradiated using known methods, which can inhibit the proliferation of tumor cell lines and induce immunogenic cell death to improve antigen delivery. The vaccine dose is typically at a 2:1 ratio to tumor cells, and in particular, the ratio of tumor cells to K562 / GM-CSF cells is 2:1. The determination of tumor cell burden can be determined by known methods, including but not limited to flow cytometry. Third, immunomodulatory drugs (IMiDs), including but not limited to lenalidomide, can significantly enhance T cell responses in cancer patients and enhance the vaccine efficacy of the vaccine compositions described herein. IMiDs that can be administered by the vaccine compositions described herein include, but are not limited to, lenalidomide, thalidomide, and pomalidomide. Lenalidomide is a specific example. Lenalidomide (sometimes referred to in the literature as "Len") may be used as a vaccine adjuvant or may be co-administered with the vaccine composition in the methods described herein. Lenalidomide may be administered any time prior to administration of the vaccine composition, co-administered with the vaccine composition, or administered after the vaccine composition. The dose of lenalidomide may range from 2.5-25 mg / dose. The time before or after administration of the vaccine composition is not limited, including up to 10 years either before or after administration, and may be up to 4 years before or after, may be 3 years before or after, may be 2 years before or after, or may be 1 year before or after, and any time between such time points, including but not limited to 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, and 1 month before or after. The administration of lenalidomide may be continuous, or in several separate doses. Administration of the vaccine composition described herein, in combination with continuous administration of lenalidomide and low tumor burden, is shown to result in effective and long-lasting anti-MM immunity.

[0036] The expansion and persistence of T cell clonotypes by bulk analysis of TCR repertoires can be observed in both blood and bone marrow (BM) after administration of the vaccine compositions described herein. Thus, both the local expansion of MM-specific clones induced by the vaccine compositions described herein in the bone marrow and the potential for infiltration by peripherally derived clonotypes can result in the development and maintenance of long-lasting immune responses. Moreover, the increase in polyfunctional cytokine-producing T cells in response to stimulation by both vaccine composition-associated and non-vaccine-associated MM antigens supports the development of broad immune responsiveness against a variety of heterologous shared MM antigens. Thus, the allogeneic whole cell vaccines described herein overcome the barriers imposed by patient-specific tumor diversity to induce effective T cell responses against shared tumor-specific antigens and, moreover, confer long-lasting antitumor immunity. The vaccine compositions described herein are shown to enhance the ability to detect T cell clonotypes expanded following vaccination and characterize polyfunctional cytokine T cell responses for up to seven years after vaccination.

[0037] Thus, the vaccine compositions described herein allow for the reversal and maintenance of myeloma-monoclonal gammopathy of unknown significance, known as "MGUS," an early stage multiple myeloma that is not actually cancer at all. MGUS is a benign condition manifested by low levels of M protein, low levels of abnormal plasma cells in the bone marrow, and the absence of indicators of active disease. This condition can be countered by the continued activation of T cell-mediated immunity induced by the vaccine compositions described herein, and tissue-resident-like CD8 T cells in the bone marrow of such patients. + Identified by the presence of T cell population. Therefore, patients who have been diagnosed with MGUS but have not progressed to multiple myeloma are also candidates for the vaccine described herein. Maintenance of MGUS patients by administering the vaccine described herein may prevent progression to myeloma. Complete remission in patients with multiple myeloma, as measured by patients having undetectable M spike protein and negative immunofixation electrophoresis readings, can be achieved by administration of the vaccine composition according to the methods and dosing schedules described herein, and thus methods of inducing complete remission in such patients are feasible. Complete remission may persist in patients for up to 5 years, up to 6 years, or up to 7 years.

[0038] Prolonged progression-free survival in subjects with multiple myeloma, as measured by determining the time to diagnosis until the date of progression, relapse or recurrence, can be achieved by administering the vaccine composition according to the methods and administration schedules described herein, thus making a method of prolonging progression-free survival in such patients feasible. Progression-free survival can be measured for up to 5 years, 6 years, or up to 7 years. Increasing clonal T cell expression and myeloma-specific cytokine responses in patients with multiple myeloma can be achieved by administration of a vaccine composition in combination with CAR T cells according to the methods and dosing schedules described herein, and thus a method of increasing clonal T cell expression and myeloma-specific cytokine responses in such patients is feasible. Induction of multiple myeloma-specific immunity in patients with multiple myeloma can be achieved by administration of vaccine compositions and CAR T cells according to the methods and administration schedules described herein, and thus a method of inducing multiple myeloma-specific immunity and achieving progression-free survival in such patients is feasible.

[0039] Prevention of multiple myeloma relapse in patients who have previously had a positive diagnosis of multiple myeloma but who have previously achieved MRD negativity can be achieved by administration of the vaccine composition either before or after administration of the MM-specific CAR T cells according to the methods and dosing schedules described herein, and thus a method of preventing multiple myeloma relapse in such patients is feasible. Thus, we herein provide evidence for the existence of a bone marrow (tissue) resident resting T cell population that lacks features of exhaustion and senescence and exhibits high PD1 levels and an effector memory-like phenotype. These findings support the concept of the BM as a reservoir of antigen-experienced memory T cells and provide evidence for a putative mechanism by which this occurs. + Unsupervised clustering of T cells allows identification of subpopulations preferentially enriched in patients in long-term disease remission. low CD27 - CD8 + This population of T cells was virtually absent in the bone marrow of patients in early post-vaccination relapse. Thus, CD27 T cells have a heterogeneous partially dysfunctional phenotype, defined by the combined expression of both exhaustion and activation markers. - CD8 + T cells have been identified as a source of MM-reactive lymphocytes. Their abundance induced by the vaccine composition described herein represents a positive prognostic significance in newly diagnosed multiple myeloma patients. Based on the MM-MGUS model of persistent immune surveillance, tumor-reactive CD8 + The depletion of T cell subpopulations significantly contributes to immune evasion and clinically meaningful disease progression. Evidence presented herein supports the notion that potentially tumor-reactive CD8 + It is clearly demonstrated that a decrease in T cell subpopulations precedes clinically evident disease relapse, and furthermore, its persistence correlates with long-term disease remission (FIGS. 14 and 15).

[0040] Moreover, evidence presented herein supports the conclusion that the mechanism by which vaccination confers antitumor immunity is through the generation of additional MM-specific T cells and stem-like resting T cells in the bone marrow. RM This supports the conclusion that the CD8 +Heterogeneous populations of T cells are identified. The phenotypic characterization of the immune phenotype of BM-resident memory T cells described herein provides further insight into the important role that bone marrow T cells play in maintaining MM-specific immunity for years after vaccination with the vaccine compositions described herein. The data described herein fully support a positive correlation between depth of MRD response and clinical outcome. The stable re-emergence of monoclonal proteins that do not meet the criteria for disease progression described herein provides evidence that the vaccine compositions administered in combination with MM-specific CAR+ T cells described herein, when administered in conditions of low disease burden, are effective in controlling progression and treating multiple myeloma by both increasing clinical responses and / or establishing an MM-MGUS equilibrium that significantly delays disease progression. EXAMPLES

[0041] The embodiments and illustrations set forth herein are explained in further detail below with reference to the following non-limiting examples. Example 1 Patient Selection and Eligibility Patients eligible to receive MM-GVAX alone were at least 18 years of age and had a diagnosis of multiple myeloma and an Eastern Cooperative Oncology Group (ECOG) performance status of 0-2 with adequate hematopoietic, hepatic, and renal function. Patients were eligible regardless of the number of previous therapies. Autologous hematopoietic stem cell transplantation could not have occurred within the past 12 months, and prior allogeneic bone marrow transplantation was not possible. To be enrolled, patients had to maintain a sustained near complete remission during at least 4 months of observation with a Len-containing regimen. Key exclusion criteria were disease progression after steroid cessation, defined as a detectable M-spike >0.5 g / dL during the observation period, or conversion to true complete remission (defined as absence of M-spike and negative serum / urine immunofixation).

[0042] Fifteen patients with multiple myeloma (MM) on Len-containing regimens who had achieved stable near complete remission (nCR) for at least 4 months were enrolled in the study. nCR was defined as undetectable M spike and positive serum and / or urine immunofixation (IFE) (BladE et al. Br J Haematol 1998;102:1115-23). ​​At the time of diagnosis, 53% of patients had stage I (International Staging System, ISS) disease (Palumbo et al. J Clin Oncol 2015;33:2863-9). Of note, none of the enrolled patients had features of high-risk MM as defined by IMWG (International Myeloma Working Group) cytogenetic criteria.

[0043] At the time of enrollment, Len was continued but all other anti-MM therapies were discontinued. Patients received 4 doses of MM-GVAX vaccination at 1, 2, 3, and 6 months in combination with Len at their current dose with a median dose of 15 mg (range 2.5-25 mg) (Figure 1) for at least 1 year. Median age at enrollment was 69 years (range 45-81 years). Patients eligible for the combination therapy of MM-GVAX and MM-specific CAR+T cells include those who have previously received CAR-T cells and have recovered from any CAR-T toxicity. Additionally, eligible patients have an Eastern Cooperative Oncology Group (ECOG) performance status of 0-2 with adequate recovery of the hematopoietic system and either MRD-negative status as determined by NGS sequencing or flow cytometry, complete remission with detectable MRD, or near complete remission defined as no detectable M spike but positive immunofixation in serum and / or urine, or have stable measurable disease for at least 3 months defined as less than 25% change on repeated measurements.

[0044] Example 2 Vaccine effectiveness All 15 patients were available for clinical response evaluation 1 year after administration of the first MM-GVAX dose. Eight of the 15 patients (53.3%) achieved a disease response consistent with true complete remission (CR), defined as absence of M spike and negative IFE, with a median time from enrollment to CR achievement of 11.6 months (range 1.4-13.9 months). This CR conversion rate of 53.3% (95% CI: 26.6-78.7%) was significantly higher than the null hypothesis of a 25% rate of CR conversion defined by design (p=0.011), thereby demonstrating clinical activity and meeting the primary endpoint of the study. Strikingly, only 6 of the 15 enrolled patients experienced disease progression, defined as the appearance of an M spike of at least 0.5 g / dL and / or an increase in free light chain content above 10 mg / dL, confirmed by repeat measurements. Pt6, Pt7 and Pt9 experienced early disease relapse within the first year of study enrollment. Seven patients (46.7%) subsequently developed detectable M-spikes that did not meet criteria for disease relapse. This was variably persistent over time but did not require any change in treatment. At the time of analysis, the estimated median overall survival (OS) was 11.5 years from MM diagnosis (95% CI: 5.9-n / a years) and 7.8 years from enrollment (95% CI: 4.2-7.8 years, n=6 / 15, 40%). Subgroup analysis of patients who achieved CR showed no statistically significant differences in terms of PFS and OS compared with patients who maintained a stable nCR or who subsequently developed a measurable M-spike. This finding suggests that vaccination can induce an immune equilibrium capable of maintaining long-term disease control even if the malignant MM plasma cell clone is not completely eradicated.

[0045] Example 3 Vaccine Formulation and Administration The cell lines used for vaccine formulation were produced by the Cell Processing and Gene Therapy Facility at the Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins in compliance with GMP. U266 and H929 were originally obtained from ATCC. K562 / GM-CSF was produced as previously described (Borrello et al.; Hum Gene Ther. 1999;10:1983-91). Briefly, K562 cells were cultured in vitro in RPMI 1640 medium (tumor medium) supplemented with 20% fetal calf serum (FCS) and penicillin-streptomycin (50 U / ml) and grown in suspension culture at 37°C and 5% CO2.

[0046] Electroporation of K562 cells was used for transfection in generating the K562GM-CSF line. K562 cells (1 × 10 7 ) were washed once with serum-free RPMI, then resuspended in 1 ml of tumor medium and mixed with 40 μg of GM-CSF plasmid DNA. The plasmid pCEP4hGM-CSF vector (Invitrogen, San Diego, CA) contains the human GM-CSF gene under the control of the cytomegalovirus (CMV) promoter as well as the hygromycin resistance gene and the EBNA-1 origin of replication sequence. The construct was digested with ClaI and AvrII to excise the EBNA-1 sequence. Cells were electroporated by shock (0.4 V, 960 mF) using a Gene Pulser cuvette (0.4-cm electrode gap) from Bio-Rad (Hercules, CA) and cultured in complete tumor medium for 24 h before drug selection.

[0047] K562-GM cells were selected and grown in tumor medium supplemented with hygromycin (400 μg / ml) and 0.1 M HEPES. These cells were further selected in high doses of hygromycin (1200 μg / ml) and subcloned by limiting dilution assay. Subclones with high GM-CSF expression were then adapted to serum-free medium AIM-V in the presence of hygromycin (Calbiochem, LaJolla, CA). Same number (5 x 10 each) 7 ) MM cell lines U266 and H929 were cultured at 5 × 10 6 The vaccine cells were mixed with the bystander cell line K562GM-CSF. After irradiation, the vaccine cells were cryopreserved and stored in liquid nitrogen until the day of use. On the day of vaccination, the individual cells were thawed, mixed at the appropriate concentrations, and drawn up into three different syringes. The final vaccine syringe was kept on ice until administration, which was done within 60 minutes of thawing.

[0048] Patients were vaccinated four times on day 14 at months 1, 2, 3, and 6 after enrollment (cycles 1, 2, 3, and 6, respectively), and Len was continued for at least 1 year at the dose administered before enrollment. Len was administered for 21 of 28 days. At single vaccination visits, patients received intradermal MM-GVAX in a total volume of 1 mL or less in three limbs, and Pneumococcal Conjugate Vaccine 13 (PCV-13, Prevnar®) was injected intramuscularly in one arm. At the end of the study, patients continued treatment with Len, and evidence of clinical efficacy was assumed. All subjects underwent BM and PB sampling for safety assessments, disease response determinations, and biomarker correlation analysis immediately prior to treatment (baseline), on day 14 of cycle 3 (C3D14, before the third vaccine dose), and 1 year after enrollment. Some patients had further follow-up BM and PB samples taken beyond 1 year and up to 7 years after study enrollment.

[0049] Example 4 MRD burden predicts vaccine response To quantify and track MM disease burden, minimal residual disease (MRD) testing by next-generation sequencing was performed in seven patients (46.7%) for whom baseline MM DNA was available (Rawstron et al. “A complementary role of multiparameter flow cytometry and high-throughput sequencing for minimal residual disease detection in chronic lymphocytic leukemia: an European Research Initiative on CLL study”; Leukemia [Internet]. 2016;30:929-36. Available at nature.com / articles / leu2015313). Test samples in all but one subject showed a B-cell repertoire MRD frequency of 10 -6 (10 -4 %), the bone marrow -3 (10 -1 %) above which (high-level MRD + ) below this threshold (low-level MRD + ) that would allow assessment of the clinical significance of disease burden.

[0050] Dominant IGH and IGK / L cancer clones were identified from the immunosequencing results in pretreatment bone marrow using the following criteria: 1) the sequence must have a frequency of >5%. 2) the sequence must be present in >0.1% of total nucleated cells. 3) the sequence must be non-contiguous (sequence frequency of ≤4 sequences over the next 10 years). 4) the sample must have >200 putative templates. These dominant clones identified in bone marrow were followed over time to determine the frequency of the cancer clones at subsequent time points after treatment. To account for somatic hypermutation (SHM), IGH clones with ≤2 mismatches to the dominant clone in bone marrow were also followed over time. MRD frequency in each sample was measured as the frequency of the cancer clone in all proliferative rearrangements of the locus examined. All high-level MRDs + Patients experienced disease recurrence within 1 year of enrollment (median = 4.8 months, range: 2.8-9.5 months) while patients had low-level MRD + Patients had a significantly longer PFS (median=84.15 months, range: 51.9-97.3 months, p=0.01). Detection of high-level MRD compared with low-level MRD was associated with an increased likelihood of clinical relapse (hazard ratio, HR=25.79, 95% CI: 2.17-306.4). Of note, Pt2 and Pt4 had prolonged clinical responses despite the occurrence of variably detectable M-spikes, which never met criteria for disease progression. In contrast, Pt12 and Pt13 achieved true CR and maintained responses over time. Thus, MRD burden at the time of vaccination can predict long-term clinical outcomes.

[0051] Example 5 Vaccination with MM-GVAX induces systemic immunity to myeloma To determine the impact of MM-GVAX vaccination on the entire T cell receptor (TCR) repertoire, deep sequencing analysis of TCR chain Vβ (TRBV) was performed on matched peripheral blood (PB) and bone marrow (BM) samples from all patients at baseline, before the third dose of MM-GVAX (C3D14), and at 1 year. Immunosequencing of human TCRβ and CDR3 regions of IGH and IGK / L chains on T and B cells was performed on genomic DNA extracted from BM or PB samples, respectively, using the immunoSEQ® assay (Adaptive Biotechnologies, Seattle, WA). Extracted genomic DNA was amplified by bias-controlled multiplex PCR followed by high-throughput sequencing.By collapsing and filtering sequences, the absolute abundance of each unique CDR3 region was identified and quantified and further analyzed as previously described (Robins et al. "Comprehensive assessment of T-cell receptor β-chain diversity in αβ T cells"; Blood [Internet]. 2009;114:4099-107. Available at ashpublications.org / blood / article / 114 / 19 / 4099 / 26771; Carlson et al. "Using synthetic templates to design an unbiased multiplex PCR assay"; Nat Commun [Internet]. 2013;4:2680. Available at nature.com / articles / ncomms3680; Robins et al. "Ultra- sensitive detection of rare T cell clones"; J Immunol Methods [Internet]. 2012;375:14-9. Available at: linkinghub.elsevier.com / retrieve / pii / S0022175911002468; Kirsch et al. “T-cell receptor profiling in cancer:; Mol Oncol [Internet]. 2015;9:2063-70. Available at: doi.wiley.com / 10.1016 / j.molonc.2015.09.003; DeWitt et al. “A Public Database of Memory and Naive B-Cell Receptor Sequences”; Turner SJ, editor. PLoS One [Internet]. 2016;11:e0160853. Available at: dx.plos.org / 10.1371 / journal.pone.0160853). Only proliferative rearrangements were used for repertoire analysis.

[0052] TCR clonotype composition varied before vaccination, ranging from minimal repertoire bias to prominent oligoclonal expansion. Despite the hypothesis that vaccination should result in an asymmetry of the TCR repertoire relative to clonality expansion, we did not observe major changes in the relative proportion of TCR proliferative rearrangements in either compartment. Proliferative clonality varied greatly over time among different patients, but no vaccine-associated patterns could be identified. Overall, proliferative clonality appeared relatively stable over time in most subjects and did not correlate with clinical outcomes. Given that asymmetries in the minimal TCR repertoire were observed with vaccination, we examined changes in clonal abundance before and after vaccination by comparing the frequency of appearance of each clone. Of note, after the first two doses of MM-GVAX (C3D14), there was a significant expansion of T cell clones in both PB and BM of all subjects, and such clones reached and persisted at high frequencies after the initial expansion after vaccination (Figure 2). Despite the lack of overall clonality changes associated with vaccination, there is clear evidence of clonal expansion and contraction in both PB and BM samples. Interestingly, many of the clones that expanded significantly after vaccination were not observed before vaccination, suggesting either the expansion of previously undetected clones or the recruitment of novel clonal types at the time of vaccination (Figure 3). The expansion of T cell clones in both PB and BM highlights the ability of MM-GVAX to induce broad systemic immune responses. Most clones were present at comparable frequencies in PB and BM, but some clonotypes were preferentially enriched in BM (Figure 4). To better understand the clonal dynamics associated with vaccination, the Morishita index was used to determine the similarity between the TCR repertoires in these two compartments. In some patients (Pt7, Pt10, and Pt15), the TCR repertoires of BM and PB became more different from each other after vaccination compared to baseline, but in most subjects, there was little change in the similarity between the two compartments after vaccination (Figure 5).Further analysis of expanded T cell clones in BM and PB after vaccination revealed diverse clonal expansion behavior across patients. In some, the frequency of expanded clonotypes increased in both PB and BM, suggesting the existence of parallel processes in both compartments. Conversely, in others, some clones expanded in one compartment but not the other, or expanded in both compartments but displayed preferential enrichment in either BM or PB. Collectively, these results indicate that MM-GVAX induced a systemic T cell response and that clonal expansion was measurable in both BM and PB.

[0053] Example 6 Vaccination induces MM-specific polyfunctional T cell responses in bone marrow T cell responses were functionally characterized in BM to both vaccine-relevant and non-relevant MM antigens. Samples from all patients and time points were stimulated in vitro with lysates of one of the MM-GVAX cell lines (U266 and H929) and analyzed for intracellular cytokine production. MM-GVAX-specific interferon-γ (IFNγ) and TNFα responses were significantly increased in C3D14 and 1-year CD8 + and CD4 + Both CD4+ and CD4+ T cell subsets produced IFNγ and / or TNFα in response to vaccine-associated and non-associated MM antigens. + or CD8 + T cell frequencies were significantly elevated after just two vaccinations and maintained a sustained increase for up to 4 years (p<0.0001, Figure 7). BM-derived mononuclear cells obtained at the indicated time points before and after vaccination were stimulated with either 2% human AB serum alone or AIM-V medium supplemented with SW780 (bladder cancer cell line) lysate or U266 / H929 (MM-GVAX cell line) lysate, respectively. After 5 days, cells were harvested and stained for flow cytometric analysis of intracellular cytokine production.

[0054] Notably, MM-GVAX inhibited the expression of polyfunctional CD4 + and CD8 +There was a significant increase in the frequency of T cells, defined as the percentage of T cells simultaneously producing IFNγ and TNFα, as well as single cytokines, albeit to a lesser extent. Interestingly, the most striking changes were in the frequency of CD8 T cells producing either TNFα or IFNγ / TNFα. + The results suggest that the MM-GVAX-induced immune response is polyfunctional, directed against a broad range of commonly shared MM-associated antigens, and is mediated by CD8 T cells (Figure 8). Pt6, Pt7, and Pt9, who relapsed early after vaccination, generated vaccine-specific T cell cytokine responses comparable to those of patients who achieved long-term disease remission. These results suggest that the MM-GVAX-induced immune response is polyfunctional, directed against a broad range of commonly shared MM-associated antigens, and mediated by CD8 T cells (Figure 8). + and CD4 + These findings demonstrate that effective vaccine-induced antitumor immunity involves both CD8 and CD8 T cells. + and CD4 + This supports data from other studies showing that tumor-specific T cells are required for both CD8 and CD8+ T cell proliferation (Bennett et al. "Induction of a CD8 + + Cytotoxic T Lymphocyte Response by Cross-priming Requires Cognate CD4 + + T Cell Help”; J Exp Med [Internet]. 1997;186:65-70. Available at rupress.org / jem / article / 186 / 1 / 65 / 7193; and Alspach E, Lussier et al.; “MHC-II neoantigens shape tumour immunity and response to immunotherapy”; Nature [Internet]. 2019;574:696-701. Available at nature.com / articles / s41586-019-1671-8).

[0055] Example 7 Vaccine-induced MM-specific T cell immunity persists for years after vaccination An important feature of adaptive immunity is its persistence over time and the ability to mount an effective response upon re-encounter with antigen. Therefore, we detected and characterized MM-GVAX-specific responses in available samples taken several years after vaccination.

[0056] Bone marrow and peripheral blood samples were collected at pre-established time points, enriched for mononuclear cells using a Lymphoprep (STEMCELL Technologies®) gradient, and cryopreserved in freezing medium (50% complete AIM-V medium, 40% complement-depleted human AB serum, and 10% DMSO). Samples were then thawed and washed twice with pre-warmed (37C) AIM-V supplemented with 0.02mg / mL DNase and phosphate-buffered saline (PBS), respectively. Flow cytometry reagents were purchased from BioLegend, BD Biosciences, and Invitrogen. Monoclonal antibodies were previously titrated to optimal concentrations. Surface staining was performed at 37C for 20 min, while intracellular detection of cytokines was performed by incubating cells with specific mAb cocktails for 20 min at room temperature after fixing the cells with CytoFix / CytoPerm kit (BD Biosciences) according to the manufacturer's instructions. All data were acquired on a Gallios® flow cytometer (Beckmann-Coulter) equipped with three lasers (violet, 405 nm; blue, 488 nm; red, 633 nm) and capable of detecting 10 parameters. Flow cytometry data were corrected using single stained cell control and compensation beads (BioLegend) with FlowJo. After pretreatment with biexponential transformation and standard gating to remove aggregates and dead cells, CD3 + CD8 +T cells were subsequently exported from FlowJo and further analyzed by R (version 4.0.1) with custom scripts using Bioconductor libraries and R packages. Briefly, data were analyzed using the FlowSOM algorithm for unsupervised clustering and visualized by UMAP. Differential discovery analysis was performed in R using the diffcyt framework and CATALYST workflow (Nowicka et al.; “CyTOF workflow: differential discovery in high-throughput high-dimensional cytometry datasets”; F1000Research [Internet]. 2019;6:748. Available at f1000research.com / articles / 6-748 / v3). Data were then generated as one new file for each cluster and further analyzed by FlowJo to determine the positive cell frequency and mean fluorescence intensity (MFI) for each marker.

[0057] In three patients for whom longitudinal samples were available, the expanded clonotypes detected in C3D14 were still present up to 7 years after vaccination. Surprisingly, the frequency of expanded clones in each compartment remained relatively stable from 1-year after vaccination in both BM and PB, suggesting the establishment of an immune steady state (Figure 9). Intracellular cytokine staining of such BM samples stimulated with both MM-GVAX-related and unrelated MM antigens revealed persistent polyfunctional CD4 + and CD8 + T cell responses were demonstrated up to 7 years after vaccination (Figure 10). In summary, the persistence of vaccine-specific clones in PB and BM as well as MM-specific polyfunctional T cells for 7 years after vaccination, even in the presence of detectable disease, suggests the establishment of an immune equilibrium, which may be responsible for the long-term disease control observed in this study.

[0058] Example 8 T cells in the BM exhibit effector phenotype and tissue-resident-like properties The phenotypic composition of BM T cells was examined for expression of checkpoint molecules, costimulatory molecules, and chemokine receptors. BM T cell composition was remarkably similar across time points (data not shown). A CD69-expressing tissue-resident-like T cell population (T RM ) was identified as being consistently present in all BM samples. + T RM The proportion of CD8 + T RM CD4 + Classical phenotypic identification of T cell subsets revealed a higher proportion of CD4 + T cells are transformed into a central memory phenotype (T CM ), whereas CD8 + In T cells, effector memory (T EM ) and effector (T EMRA ) subset was found to be typical. + In T cells, CD4 + The frequency of stem cell memory-like T cells (T SCM ) was present (Figure 12). RM is T EM and T EMRA Although the phenotype was predominantly observed in BM, TCM and TSCM-like TRM could only be detected to a low extent. + CD8 +T cells are representative of memory populations and have tissue-resident characteristics. Interestingly, they lack exhaustion markers such as TIGIT and TIM3, as well as senescence markers (CD57), but express activation markers CD27 and PD1. Interestingly, BM TRM express higher levels of both BM-homing chemokine receptors CXCR4 and CXCR6, which are believed to be characteristic of tissue-resident T cells (Kumar et al., "Human Tissue- Resident Memory T Cells Are Defined by Core Transcriptional and Functional Signatures in Lymphoid and Mucosal Sites"; Cell Rep [Internet]. ElsevierCompany.; 2017;20:2921-34. Available at dx.doi.org / 10.1016 / j.celrep.2017.08.078) (Figure 13). A significant proportion of patients maintained disease control despite re-emergence of low levels of monoclonal protein. This re-emergence persisted over time, but did not meet the criteria for relapse. This stable low-level disease suggests the establishment of a state of immune equilibrium. The identification of a BM-resident T cell population consistently present in this cohort highlights the role of T cells in maintaining immune equilibrium and tumor surveillance. RM This supports previous findings that tissue-resident memory CD8 + + T cells promote melanoma-immune equilibrium in skin”; Nature [Internet]. 2019;565:366-71. Available at: nature.com / articles / s41586-018- 0812-9.

[0059] Example 9 Immune correlates of clinical outcomes following MM-GVAX vaccination BM CD8 after vaccination +To gain insight into the T cell immunophenotype and their association with clinical outcomes, BM CD8 + T cells were analyzed by FlowSOM, an unsupervised clustering algorithm, and dimensionality reduction techniques, such as Uniform Manifold Approximation and Projection (UMAP), were used to simplify the visualization of the different T cell clusters. Hierarchical metaclustering of FlowSOM clusters revealed that CD8 + T cell subpopulations were grouped into similar immunophenotypes, CD8 + Two subsets of T cells were identified: C1 and C2. These subsets were enriched in patients with long-lasting disease control (responders) compared to patients with early progressive disease (relapsers) within the first year after vaccination (Figure 14, *, p<0.05; **, p<0.01). Strikingly, clusters C1 and C2 were defined by low to absent expression of DNAM1 and lack of CD27, and were enriched for senescent, effector and exhausted CD8 + Further characterization of cluster C2 revealed that it contained a relatively heterogeneous subpopulation of T cells that moderately expressed PD1 and CD69 + CD57- subpopulation was identified and enriched in responders with CD8 + It was suggested that T cells reside in the BM and may play a role in long-term MM control. Interestingly, cluster C1 was characterized by elevated CD57 expression, suggesting that such effector-senescent cells may still be functional, despite lack of proliferative capacity. The results obtained by the FlowSOM algorithm were then reproduced by standard methods of flow cytometry, where manually gated CD27-DNAM1low / -CD8 + T cells were enriched in vaccine responders (Figure 15). + Changes in T cell composition occur earlier than clinically evident disease relapse and may represent a reservoir of tumor-specific T cells, potentially playing a role in the establishment and maintenance of vaccine-induced immune equilibrium. +Evidence is provided that T cell subsets can be identified.

[0060] Example 10 Generation of MM-specific CAR+ T cells Chimeric antigen receptors can be generated by taking T cells from a patient, which are autologous (allogeneic) T cells, and transducing such cells with a lentiviral vector encoding a second generation CAR with an anti-MM specific single chain variable fragment antibody, e.g., anti-BCMA, CD137 (4-1BB) or CD28 costimulatory motif, and a CD3 zeta signaling domain. After transduction of the T cells, the cells can be expanded ex vivo for a period of at least 7 days up to 30 days. The cells can be formulated into a composition suitable for injection using known methods.

[0061] Example 11 Administration of MM-specific CAR+ T cells before and / or after vaccination MM-specific CAR+ T cell compositions can be administered to eligible patients either before or after administration of MM-GVAX. Patients can be lymphodepleted with fludarabine (30 mg per square meter of body surface area per day) and cyclophosphamide (300 mg per square meter per day) on days -5, -4, and -3, followed by infusion of MM-specific CAR+ T cells on day 0. 50 x 10 total CAR-positive T cells during the dose escalation phase. 6 , 150×10 6 , 450×10 6 , or 800 x 10 6 cells (+ / - 20%) and total CAR+ T cells in the expansion phase: 150 x 10 6 ~450×10 6 A dose of cells can be administered. Clinical response and disease progression can be assessed according to the IMWG Uniform Response Criteria for Multiple Myeloma. Endpoints can include assessment of MRD, which can be determined by next generation sequencing, overall survival, and progression-free survival, measurement of cytokines and chemokines, and quantification of MM-specific antigens in the blood.

[0062] Following MM-specific CAR+ T cell administration, MM-GVAX can be administered again as a second, third, or fourth dose. Alternatively, the patient can be administered MM-specific CAR+T cells first, followed by MM-GVAX as described above. One, two, three, or more doses of MM-GVAX can be administered after administration of the MM-specific CAR+T cells. MM-GVAX can be administered a minimum of two months and up to two years after administration of the MM-specific CAR+T cells. MM-GVAX can be administered at least once within this time frame, and can also be administered multiple times. To enhance the efficacy of MM-GVAX, it can be co-administered with several immune enhancing agents, including, but not limited to, lenalidomide and pomalidomide.

[0063] The efficacy of this combined administration of MM-GVAX and MM-specific CAR+ T cells results in endpoints such as MRD negativity and progression-free survival. Although the present invention has been described in detail with reference to exemplary embodiments thereof, it will be apparent to those skilled in the art that various modifications can be made and equivalents can be utilized without departing from the scope of the invention. Each of the aforementioned documents is incorporated herein by reference in its entirety.

Claims

1. A composition for use in raising an immune response to a plasma cell disorder in a subject, comprising multiple myeloma-specific CAR+ T cells.

2. 2. The composition of claim 1, wherein the multiple myeloma-specific CAR+ T cells are BCMA-specific CAR+ T cells.

3. 2. The composition of claim 1, wherein the multiple myeloma-specific CAR+ T cells are GDRC5D-specific CAR+ T cells.

4. The composition of claim 2 or 3, which is allogeneic.

5. The composition of claim 1, which when administered to a subject induces an immune response in the subject.

6. The composition of claim 5 , wherein the immune response induces a complete remission of the plasma cell disorder in the subject.

7. The composition of claim 5, which extends progression-free survival in the subject.

8. The composition of claim 6, wherein the complete response is determined as no detectable M spike and negative immunofixation electrophoresis.

9. The composition of claim 1 , wherein the subject is a human.

10. 1. A composition comprising multiple myeloma-specific CAR+ T cells for use in a method of treating a plasma cell disorder in a subject, the method comprising administering to the subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells.

11. The composition of claim 10 , wherein the administering step also comprises administering an immunomodulatory agent to the subject.

12. 11. The composition of claim 10, wherein the immunomodulatory agent is lenalidomide.

13. The composition of claim 10 , wherein the immunomodulatory agent is administered to the subject before, during, and / or after the administering step.

14. The composition of claim 10, wherein the vaccine composition is allogeneic.

15. The composition of claim 10, wherein K562 cells express the GM-CSF gene.

16. The composition of claim 10, wherein the K562 cells are transfected with a gene encoding GM-CSF.

17. The GM-CSF gene is at a maximum of about 1500 ng / 1×10 6 The composition of claim 16, capable of expressing the amount of GM-CSF per cell / 24 hours.

18. The amount of GM-CSF expressed is about 35 to 1200 ng / 1×10 6 The composition of claim 17, wherein the cell / 24 hours.

19. 18. The composition of claim 17, wherein the amount of GM-CSF produced is on average every 24 hours.

20. The composition of claim 15, wherein the GM-CSF is of human origin.

21. 11. The composition of claim 10, wherein the ratio of a combination of U266 and H929 cells to K562 cells is about 20:

1.

22. 11. The composition of claim 10, wherein the dose of the vaccine composition is such that the ratio of tumor cells in the subject to K562 cells in the composition is greater than 2:

1.

23. The composition of claim 10, wherein U266 and H929 cells are present in the vaccine composition in equal amounts.

24. The U266 and H929 cells were approximately 5×10 7 and K562 cells are present in the vaccine composition in an amount of about 5×10 6 The composition of claim 10, wherein the composition is present in the vaccine composition in an amount of cells.

25. The composition of claim 10, wherein a near complete or complete response is achieved in the subject.

26. 26. The composition of claim 25, wherein the near complete or complete remission is sustained in the subject for up to five years.

27. 26. The composition of claim 25, wherein the complete response is determined by measuring undetectable monoclonal spike and negative immunofixation electrophoresis.

28. The composition of claim 10, wherein the subject is positive for minimal residual disease.

29. The composition of claim 10, wherein the vaccine composition minimizes a non-specific immune response in a subject.

30. 11. The composition of claim 10, wherein the vaccine composition is administered prior to the CAR+ T cell composition.

31. 11. The composition of claim 10, wherein the CAR+ T cell composition is administered prior to the vaccine composition.

32. 11. The composition of claim 10, wherein the vaccine composition is administered followed by the CAR+ T cell composition followed by a second dose of the vaccine composition.

33. The method of claim 10, wherein 1 to 5 doses of the vaccine composition are administered to the subject with an interval of more than 1 day between each dose.

34. The method of claim 33, wherein 2 to 4 doses of the vaccine composition are administered with an interval of more than 2 weeks between each dose.

35. 34. The composition of claim 33, wherein more than four weeks pass between each administration.

36. 34. The composition of claim 33, wherein four doses are administered with an interval of about one month between each dose.

37. 34. The composition of claim 33, wherein the first three administrations are equally spaced.

38. 34. The composition of claim 33, wherein all doses are administered within one year of each other.

39. 13. The composition of claim 12, wherein at least one dose of the vaccine composition is administered between days 7 and 18, inclusive, from the start of a course of lenalidomide.

40. 13. The composition of claim 12, wherein at least one dose is administered about 15 days after the start of a lenalidomide course.

41. 11. The composition of claim 10, wherein the plasma cell disorder is selected from the group consisting of MGUS, SMM, multiple myeloma, non-secretory multiple myeloma, indolent myeloma, light chain myeloma, plasma cell leukemia, and primary amyloidosis.

42. 42. The composition of claim 41, wherein the plasma cell disorder is multiple myeloma.

43. 1. A composition comprising multiple myeloma-specific CAR+ T cells for use in a method of extending progression-free survival in a subject with multiple myeloma, the method comprising administering to a subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells.

44. 1. A composition comprising multiple myeloma-specific CAR+ T cells for use in a method of inducing clonal T cell expansion and increased myeloma-specific cytokine response in a subject with multiple myeloma, the method comprising administering to the subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells.

45. 45. The composition of claim 44, wherein said increase in said subject persists for up to seven years after said administering step.

46. 45. The composition of claim 44, wherein said increase in said subject persists for up to five years after said administering step.

47. 1. A composition comprising multiple myeloma-specific CAR+ T cells for use in a method of inducing multiple myeloma-specific immunity in a subject, the method comprising administering to a subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells.

48. The composition of claim 10, wherein the subject is positive for minimal residual disease at the time of the administering step.

49. 1. A composition comprising multiple myeloma-specific CAR+ T cells for use in a method of preventing recurrence of multiple myeloma in a subject, the method comprising administering to a subject a CAR+ T cell composition comprising multiple myeloma-specific CAR+ T cells and a vaccine composition comprising U266, H929, and K562 cells.

50. 50. The composition of claim 49, wherein the subject is positive for minimal residual disease at the time of the administering step.

51. The composition of claim 10 , wherein the subject is a human.

52. The composition of claim 10, wherein the multiple myeloma-specific CAR+ T cells are BCMA-specific CAR+ T cells and / or GPRC5D-specific CAR+ T cells.

Images