Exploring cellular immunotherapy platforms in multiple myeloma.

Despite major advances in therapeutic platforms, most patients with multiple myeloma (MM) eventually relapse and succumb to the disease. Among the novel therapeutic options developed over the past decade, genetically engineered T cells have a great deal of potential. Cellular immunotherapies, including chimeric antigen receptor (CAR) T cells, are rapidly becoming an effective therapeutic modality for MM. Marrow-infiltrating lymphocytes (MILs) derived from the bone marrow of patients with MM are a novel source of T cells for adoptive T-cell therapy, which robustly and specifically target myeloma cells. In this review, we examine the recent innovations in cellular immunotherapies, including the use of dendritic cells, and cellular tools based on MILs, natural killer (NK) cells, and CAR T cells, which hold promise for improving the efficacy and/or reducing the toxicity of treatment in patients with MM.

Efficacy of natural killer cell therapy combined with chemoradiotherapy in murine models of head and neck squamous cell carcinoma.

BACKGROUND AIMS: Natural killer (NK) cell-based cancer immunotherapy is effective when combined with other treatment modalities such as irradiation and chemotherapy. NK cell's antitumor function to treat solid tumor, including head and neck squamous cell carcinoma (HNSCC), has been targeted recently. This study assessed NK cell recruitment in response to chemoradiation therapy (CRT) in HNSCC. METHODS: Ex vivo expansion of NK cell, flow cytometry, cell viability assay, cytotoxicity assay, immunohistochemistry, and animal model were performed. RESULTS: Mouse NK cells were recruited to the tumor site by CRT in a nude mouse model. Furthermore, expanded and activated human NK cells (eNKs) were recruited to the tumor site in response to CRT, and CRT enhanced the anti-tumor activity of eNK in an NOD/SCID IL-2Rγnull mouse model. Various HNSCC cancer cell lines exhibited different NK cell ligand activation patterns in response to CRT that correlated with NK cell-mediated cytotoxicity. CONCLUSIONS: Identifying the activation patterns of NK cell ligands during CRT might improve patient selection for adjuvant NK cell immunotherapy combined with CRT. This is the first study to investigate the NK cell's antitumor function and recruitment with CRT in HNSCC mouse model.

Adoptive NK Cell Therapy - a Beacon of Hope in Multiple Myeloma Treatment.

Major advances in the treatment of multiple myeloma (MM) have been achieved by effective new agents such as proteasome inhibitors, immunomodulatory drugs, or monoclonal antibodies. Despite significant progress, MM remains still incurable and, recently, cellular immunotherapy has emerged as a promising treatment for relapsed/refractory MM. The emergence of chimeric antigen receptor (CAR) technology has transformed immunotherapy by enhancing the antitumor functions of T cells and natural killer (NK) cells, leading to effective control of hematologic malignancies. Recent advancements in gene delivery to NK cells have paved the way for the clinical application of CAR-NK cell therapy. CAR-NK cell therapy strategies have demonstrated safety, tolerability, and substantial efficacy in treating B cell malignancies in various clinical settings. However, their effectiveness in eliminating MM remains to be established. This review explores multiple approaches to enhance NK cell cytotoxicity, persistence, expansion, and manufacturing processes, and highlights the challenges and opportunities associated with CAR-NK cell therapy against MM. By shedding light on these aspects, this review aims to provide valuable insights into the potential of CAR-NK cell therapy as a promising approach for improving the treatment outcomes of MM patients.

Expanded natural killer cells potentiate the antimyeloma activity of daratumumab, lenalidomide, and dexamethasone in a myeloma xenograft model.

The development of new treatment agents in recent decades has significantly improved the survival of patients with multiple myeloma (MM). Nonetheless, MM remains an incurable disease; therefore, novel combination therapies are required. Natural killer (NK) cells are one of the safest immunotherapeutic options. In this study, we found that the anti-myeloma activity of expanded NK cells (eNKs) was improved by daratumumab, lenalidomide, and dexamethasone (DRd) in an MM xenograft mouse model. NK cells expanded from peripheral blood mononuclear cells collected from MM patients were highly cytotoxic against DRd pretreated tumor cells in vitro. To mimic the clinical protocol, a human MM xenograft model was developed using human RPMI8226-RFP-FLuc cells in NOD/SCID IL-2Rγnull (NSG) mice. MM bearing mice were randomly divided into six groups: no treatment, eNK, Rd, Rd + eNKs, DRd, and DRd + eNKs. DRd significantly enhanced the cytotoxicity of eNKs by upregulating NK cell activation ligands and effector function. DRd in combination with eNKs significantly reduced the serum M-protein level and prolonged mouse survival. In addition, DRd significantly increased the persistence of eNK and homing to MM sites. These results show that the anti-myeloma activity of ex vivo-expanded and activated NK cells is augmented by the immunomodulatory effect of DRd in MM-bearing mice, suggesting the therapeutic potential of this combination for MM patients.

Natural killer cells have a synergistic anti-tumor effect in combination with chemoradiotherapy against head and neck cancer.

BACKGROUND: The use of natural killer (NK) cells is a promising approach in the field of cancer immunotherapy; however, combination treatments are required to enhance the effects of NK cell immunotherapy. In this study, we assessed the potential of irradiation and cisplatin as a chemoradiotherapy (CRT) regimen to augment the effects of NK cell immunotherapy in head and neck squamous cell carcinoma (HNSCC). METHODS: NK cells were expanded using our recently established K562-OX40 ligand and membrane-bound interleukin (IL)-18 and IL-21 feeder cells in the presence of IL-2/IL-15 from peripheral blood of healthy donors. RESULTS: The results showed an increase in the purity of NK cells and expression of activation markers such as NKG2D and lymphocyte function-associated antigen 1 during the expansion process, which is positively correlated to the NK cell infiltration and overall survival in patients with HNSCC. CRT induced NK cell activation ligand (ULBP2) and adhesion molecules (ICAM-1, -2 and -3) on HNSCC, leading to enhanced cytotoxicity of NK cells against HNSCC. CONCLUSIONS: Our findings suggest that the NK cells have a potent anti-tumor effect in combination with CRT against HNSCC.

Novel IL-15 dendritic cells have a potent immunomodulatory effect in immunotherapy of multiple myeloma.

Dendritic cells (DCs) are the most potent antigen-presenting cells, and have thus been used in clinical cancer vaccines. However, the effects of DC vaccines are still limited, leading researchers to explore novel ways to make them effective. In this study, we investigated whether human monocyte-derived DCs generated via the addition of interleukin 15 (IL-15) had a higher capacity to induce antigen-specific T cells compared to conventional DCs. We isolated CD14+ monocytes from peripheral blood from multiple myeloma (MM) patients, and induced immature DCs with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-4 in the presence or absence of IL-15 for 4-6 days. Then we generated mature DCs (mDCs) with lipopolysaccharide for another 2 days [IL-15 mDCs (6 days), IL-15 mDCs (8 days), and conventional mDCs (8 days)]. IL-15 mDCs (6 days) showed higher expression of MHC I and II, CD40, CD86, and CCR7, and the secretion of IFN-γ was significantly higher compared to conventional mDCs. IL-15 mDCs (6 days) showed superior polarization of naïve T cells toward Th1 cells and a higher proportion of activated T cells, cytokine-induced killer (CIK) cells, and natural killer (NK) cells for inducing strong cytotoxicity against myeloma cells, and lower proportion of regulatory T cells compared to conventional mDCs. These data imply that novel multipotent mDCs generated by the addition of IL-15, which can be cultivated in 6 days, resulted in outstanding activation of T cells, CIK cells and NK cells, and may facilitate cellular immunotherapy for cancer patients.

Expansion of cytotoxic natural killer cells in multiple myeloma patients using K562 cells expressing OX40 ligand and membrane-bound IL-18 and IL-21.

BACKGROUND: Natural killer (NK) cell-based immunotherapy is a promising treatment approach for multiple myeloma (MM), but obtaining a sufficient number of activated NK cells remains challenging. Here, we report an improved method to generate ex vivo expanded NK (eNK) cells from MM patients based on genetic engineering of K562 cells to express OX40 ligand and membrane-bound (mb) IL-18 and IL-21. METHODS: K562-OX40L-mbIL-18/-21 cells were generated by transducing K562-OX40L cells with a lentiviral vector encoding mbIL-18 and mbIL-21, and these were used as feeder cells to expand NK cells from peripheral blood mononuclear cells of healthy donors (HDs) and MM patients in the presence of IL-2/IL-15. Purity, expansion rate, receptor expression, and functions of eNK cells were determined over four weeks of culture. RESULTS: NK cell expansion was enhanced by short exposure of soluble IL-18 and IL-21 with K562-OX40L cells. Co-culture of NK cells with K562-OX40L-mbIL-18/-21 cells resulted in remarkable expansion of NK cells from HDs (9,860-fold) and MM patients (4,929-fold) over the 28-day culture period. Moreover, eNK cells showed increased expression of major activation markers and enhanced cytotoxicity towards target K562, U266, and RPMI8226 cells. CONCLUSIONS: Our data suggest that genetically engineered K562 cells expressing OX40L, mbIL-18, and mbIL-21 improve the expansion of NK cells, increase activation signals, and enhance their cytolytic activity towards MM cells.

Expanded natural killer cells augment the antimyeloma effect of daratumumab, bortezomib, and dexamethasone in a mouse model.

The use of natural killer (NK) cells is a promising and safe immunotherapeutic approach in the field of cancer immunotherapy. However, combination treatments are required to enhance the effector functions and therapeutic efficacy of NK cells. In this study, we investigated the potential of daratumumab (Dara), bortezomib, and dexamethasone (Dvd) to augment the antitumor effects of NK cells in a multiple myeloma (MM) xenograft mouse model. NK cells were expanded and activated using the K562-OX40 ligand and membrane-bound IL-18 and IL-21 in the presence of IL-2 and IL-15 from peripheral blood mononuclear cells from MM patients. A human MM xenograft model was established using human RPMI8226-RFP-FLuc cells in NOD/SCID IL-2Rγnull (NSG) mice. Tumor-bearing mice were divided into six treatment groups: no treatment, expanded NK cells (eNKs), Dara, Dara + eNKs, Dvd, and Dvd + eNKs. Dvd treatment strongly enhanced the cytotoxicity of eNKs by upregulating expression of NK cell activation ligands, downregulating expression of NK cell inhibitory ligands, and promoting antibody-dependent cellular cytotoxicity. The combination of eNKs with Dvd significantly prolonged mouse survival and reduced the tumor burden and serum M-protein level. Furthermore, Dvd pretreatment significantly increased eNK persistence and homing to MM sites. Our findings suggest that Dvd treatment potentiates the antimyeloma effects of NK cells expanded and activated ex vivo by modulating immune responses in MM-bearing mice.

Pilot Study: Quantitative Photoacoustic Evaluation of Peripheral Vascular Dynamics Induced by Carfilzomib In Vivo.

Carfilzomib is mainly used to treat multiple myeloma. Several side effects have been reported in patients treated with carfilzomib, especially those associated with cardiovascular events, such as hypertension, congestive heart failure, and coronary artery disease. However, the side effects, especially the manifestation of cardiovascular events through capillaries, have not been fully investigated. Here, we performed a pilot experiment to monitor peripheral vascular dynamics in a mouse ear under the effects of carfilzomib using a quantitative photoacoustic vascular evaluation method. Before and after injecting the carfilzomib, bortezomib, and PBS solutions, we acquired high-resolution three-dimensional PAM data of the peripheral vasculature of the mouse ear during each experiment for 10 h. Then, the PAM maximum amplitude projection (MAP) images and five quantitative vascular parameters, i.e., photoacoustic (PA) signal, diameter, density, length fraction, and fractal dimension, were estimated. Quantitative results showed that carfilzomib induces a strong effect on the peripheral vascular system through a significant increase in all vascular parameters up to 50%, especially during the first 30 min after injection. Meanwhile, bortezomib and PBS do not have much impact on the peripheral vascular system. This pilot study verified PAM as a comprehensive method to investigate peripheral vasculature, along with the effects of carfilzomib. Therefore, we expect that PAM may be useful to predict cardiovascular events caused by carfilzomib.

Potent anti-myeloma efficacy of dendritic cell therapy in combination with pomalidomide and programmed death-ligand 1 blockade in a preclinical model of multiple myeloma.

Dendritic cell (DC)-based vaccines are recognized as a promising immunotherapeutic strategy against cancer; however, the efficacy of immunotherapy with DCs is controlled via immune checkpoints, such as programmed death-ligand 1 (PD-L1). PD-L1 expressed on DC and tumor cells binds to programmed death-1 (PD-1) receptors on the activated T cells, which leads to the inhibition of cytotoxic T cells. Blocking of PD-L1 on DC may lead to improve the efficacy of DC therapy for cancer. Here we demonstrated that DC vaccination in combination with pomalidomide and programmed death-ligand 1 (PD-L1) blockade inhibited tumor growth of a multiple myeloma (MM) mouse model. DCs + pomalidomide with dexamethasone + PD-L1 blockade significantly inhibited immune immunosuppressive factors and promoted proportions of immune effector cells in the spleen and tumor microenvironment. Additionally, functional activities of cytotoxic T lymphocytes and NK cells in spleen were enhanced by DCs + pomalidomide with dexamethasone + PD-L1 blockade. Taken together, this study identifies a potential new therapeutic approach for the treatment of MM. These results also provide a foundation for the future development of immunotherapeutic modalities to inhibit tumor growth and restore immune function in MM.

A novel TLR4 binding protein, 40S ribosomal protein S3, has potential utility as an adjuvant in a dendritic cell-based vaccine.

BACKGROUND: Dendritic cells (DCs) are professional antigen presenting cells (APCs), which can activate antigen-specific CD8+ T cell immunity, resulting in tumor clearance. Immature DCs are usually stimulated by various adjuvants through their immune receptors. Among them, Toll-like receptor 4 (TLR4) has an important role in activating DCs to cause their maturation. In fact, TLR4 is well-known to induce innate and adaptive immune responses against various external microbial or internal damage associated molecular patterns (DAMP). LPS is widely regarded as a strong stimulator of TLR4 signaling. However, LPS is inappropriate for use in humans since it is an endotoxin. Unfortunately, other TLR4 ligands such as HMGB1 or heat shock proteins have weak adjuvant effects. Therefore, there is a need to identify novel, biocompatible, strong, TLR4 ligands. METHODS: 40S ribosomal protein S3 (RPS3) was screened through pull-down assay using TLR4. BMDCs from wild type (WT) and TLR4 knock-out mice were treated by RPS3 to identify the activation and maturation of DCs. T cell generation including memory T cells, tumor prevention, and treatment experiments were performed with BMDCs based vaccination. Also, human DCs originated from patients were treated by RPS3 to confirm the activation and maturation of DCs. RESULTS: In this study, we identified 40S ribosomal protein S3 (RPS3) through a pull-down assay using a variety of human cancer cell-derived proteins that could bind to TLR4. RPS3 was released from tumor cells following treatment with an anticancer drug, and it was shown that the released RPS3 binds to TLR4. Recombinant RPS3 induced maturation and activation of DCs, and following pulsing with tumor specific antigens, these DCs could be used as a vaccine to significantly increase tumor specific CD8+IFN-γ+ T cells, and provide both tumor prevention and tumor treatment effects. The effect of RPS3 on DC maturation and its utility as a vaccine were shown to be dependent on TLR4 using TLR4 knockout mice. CONCLUSIONS: This study therefore proved that human cancer cell-derived RPS3, a novel TLR4 ligand, has great potential as an adjuvant in tumor-specific antigen DC-based vaccines.

Cellular immunotherapy in multiple myeloma.

In multiple myeloma (MM), the impaired function of several types of immune cells favors the tumor's escape from immune surveillance and, therefore, its growth and survival. Tremendous improvements have been made in the treatment of MM over the past decade but cellular immunotherapy using dendritic cells, natural killer cells, and genetically engineered T-cells represent a new therapeutic era. The application of these treatments is growing rapidly, based on their capacity to eradicate MM. In this review, we summarize recent progress in cellular immunotherapy for MM and its future prospects.

A novel function of API5 (apoptosis inhibitor 5), TLR4-dependent activation of antigen presenting cells.

Dendritic cell (DC)-based vaccines are recognized as a promising immunotherapeutic strategy against cancer. Various adjuvants are often incorporated to enhance the modest immunogenicity of DC vaccines. More specifically, many of the commonly used adjuvants are derived from bacteria. In the current study, we evaluate the use of apoptosis inhibitor 5 (API5), a damage-associated molecular pattern expressed by many human cancer cells, as a novel DC vaccine adjuvant. We showed that API5 can prompt activation and maturation of DCs and activate NFkB by stimulating the Toll-like receptor signaling pathway. We also demonstrated that vaccination with API5-treated DCs pulsed with OVA, E7, or AH1-A5 peptides led to the generation of OVA, E7, or AH1-A5-specific CD8 + T cells and memory T cells, which is associated with long term tumor protection and antitumor effects in mice, against EG.7, TC-1, and CT26 tumors. Additionally, we determined that API5-mediated DC activation and immune stimulation are dependent on TLR4. Lastly, we showed that the API5 protein sequence fragment that is proximal to its leucine zipper motif is responsible for the adjuvant effects exerted by API5. Our data provide evidence that support the use of API5 as a promising adjuvant for DC-based therapies, which can be applied in combination with other cancer therapies. Most notably, our results further support the continued investigation of human-based adjuvants.

Synergistic Antimyeloma Activity of Dendritic Cells and Pomalidomide in a Murine Myeloma Model.

We have previously shown that immunization with tumor antigen-loaded dendritic cells (DCs) and the immunomodulating drug, lenalidomide, synergistically potentiates the enhancing antitumor immunity in a myeloma mouse model. In this study, we investigated the immunogenicity of DCs combined with pomalidomide and dexamethasone in a myeloma mouse model. MOPC-315 cells were injected subcutaneously to establish myeloma-bearing mice. Four test groups were used to mimic clinical protocol: (1) PBS control, (2) DCs, (3) pomalidomide + dexamethasone, and (4) DCs + pomalidomide + dexamethasone. The combination of DCs plus pomalidomide and dexamethasone displayed greater inhibition of tumor growth compared to the other groups. This effect was closely related with reduced numbers of immune suppressor cells including myeloid-derived suppressor cells, M2 macrophages, and regulatory T cells, with the induction of immune effector cells such as CD4+ and CD8+ T cells, memory T cells, natural killer (NK) cells, and M1 macrophages, and with the activation of T lymphocytes and NK cells in the spleen. Moreover, the level of the immunosuppressive factor vascular endothelial growth factor was significantly reduced in the tumor microenvironment. The collective findings in the murine myeloma model suggest that tumor antigen-loaded DCs combined with pomalidomide and dexamethasone synergistically enhance antitumor immunity by skewing the immune-suppressive status toward an immune-supportive status.

Lenalidomide and Programmed Death-1 Blockade Synergistically Enhances the Effects of Dendritic Cell Vaccination in a Model of Murine Myeloma.

The therapeutic efficacy of dendritic cell (DC)-based immunotherapy may be potentiated in combination with other anticancer therapies that enhance DC function by modulating immune responses and the tumor microenvironment. In this study, we investigated the efficacy of DC vaccination in combination with lenalidomide and programmed death (PD)-1 blockade in a model of murine myeloma. MOPC-315 cell lines were injected subcutaneously to establish myeloma-bearing mice and the following five test groups were established: PBS control, DCs, DCs + lenalidomide, DCs + PD-1 blockade, and DCs + lenalidomide + PD-1 blockade. The combination of DCs plus lenalidomide and PD-1 blockade more potently inhibited tumor growth compared to the other groups. This effect was associated with a reduction in immune suppressor cells (such as myeloid-derived suppressor cells, M2 macrophages, and regulatory T cells) and an increase in immune effector cells [such as CD4+ and CD8+ T cells, natural killer (NK) cells, and M1 macrophages] in the spleen. Functional activities of cytotoxic T lymphocytes and NK cells were also enhanced by the triple combination. Levels of immunosuppressive cytokines, such as TGF-ß and IL-10, were significantly reduced in the tumor microenvironment. These findings suggest that the combination of DCs plus lenalidomide and PD-1 blockade synergistically establishes a robust anti-myeloma immunity through a two-way mechanism, which inhibits immunosuppressive cells while activating effector cells with superior polarization of the Th1/Th2 balance in favor of the tumor immune response. This result should provide an experimental ground for incorporating check point inhibitors to existing immunotherapeutic modalities against multiple myeloma.

Combination therapy with dendritic cells and lenalidomide is an effective approach to enhance antitumor immunity in a mouse colon cancer model.

In this study, we investigated efficacy of lenalidomide in combination with tumor antigen-loaded dendritic cells (DCs) in murine colon cancer model. MC-38 cell lines were injected subcutaneously to establish colon cancer-bearing mice. After tumor growth, lenalidomide (50 mg/kg/day) was injected intraperitoneally on 3 consecutive days in combination with tumor antigen-loaded DC vaccination on days 8, 12, 16, and 20. The tumor antigen-loaded DCs plus lenalidomide combination treatment exhibited a significant inhibition of tumor growth compared with the other groups. These effects were associated with a reduction in immune suppressor cells, such as myeloid-derived suppressor cells and regulatory T cells, with the induction of immune effector cells, such as natural killer cells, CD4+ T cells and CD8+ T cells in spleen, and with the activation of cytotoxic T lymphocytes and NK cells. This study suggests that a combination of tumor antigen-loaded DC vaccination and lenalidomide synergistically enhanced antitumor immune response in the murine colon cancer model, by inhibiting the generation of immune suppressive cells and recovery of effector cells, and demonstrated superior polarization of Th1/Th2 balance in favor of Th1 immune response. This combination approach with DCs and lenalidomide may provide a new therapeutic option to improve the treatment of colon cancer.

Chaetocin enhances dendritic cell function via the induction of heat shock protein and cancer testis antigens in myeloma cells.

Dendritic cells (DC)-based vaccines are considered useful in cancer immuno-therapy, and the interactions of DC and dying tumor cells are important and promising for cancer immunotherapy. We investigated whether chaetocin could be used to induce death of myeloma cells, for loading onto DCs can affect DCs function. In this study, we show that the dying myeloma cells treated with chaetocin resulted in the induction of heat shock protein (HSP) 90, which was inhibited by antioxidant N-acetyl cysteine, and showed an increase in the expression of MAGE-A3 and MAGE-C1/CT7. DCs loaded with chaetocin-treated dying myeloma cells produced low levels of IL-10 and enhanced the cross presentation of DCs. Additionally, these DCs most potently inhibited regulatory T cells, induced Th1 polarization and activated myeloma-specific cytotoxic T lymphocytes compared with DCs loaded with UVB-irradiated dying myeloma cells. These results suggest that the pretreatment of myeloma cells with chaetocin can enhance DC function through the up-regulation of HSP90 and cancer testis antigens in dying myeloma cells and can potently induce the Th1 polarization of DCs and myeloma-specific cytotoxic T lymphocytes.

Immunotherapy for the treatment of multiple myeloma.

Immunotherapy has recently emerged as a promising treatment for multiple myeloma (MM). There are now several monoclonal antibodies that target specific surface antigens on myeloma cells or the checkpoints of immune and myeloma cells. Elotuzumab (targeting SLAMF7), daratumumab (targeting CD38), and pembrolizumab (targeting PD-1) have shown clinical activity in clinical studies with relapsed/refractory MM. Dendritic cell vaccination is a safe strategy that has shown some efficacy in a subset of myeloma patients and may become a crucial part of MM treatment when combined with immunomodulatory drugs or immune check-point blockade. Genetically engineered T cells, such as chimeric antigen receptor T cells or T cell receptor-engineered T cells, have also shown encouraging results in recent clinical studies of patients with MM. In this paper, we discuss recent progress in immunotherapy for the treatment of MM.

Lenalidomide enhances the function of dendritic cells generated from patients with multiple myeloma.

Lenalidomide (LEN) has been used as an immunomodulatory drug with direct and indirect anti-tumor effects. In this study, we evaluated the effect of LEN on the differentiation, maturation, and function of dendritic cells (DCs) in patients with multiple myeloma in vitro. Various doses of LEN were added after the monocytes had differentiated into immature DCs and were activated into mature DCs. LEN (5 µg/mL) was the optimal concentration to promote differentiation and maturation of DCs. Immature DCs treated with LEN exhibited enhanced endocytic capacity. Mature DCs treated with LEN produced higher levels of interleukin-12p70, possessed stronger allogeneic T-cell stimulation capacity, reduced the number of suppressor cells, and generated antigen-specific cytotoxic T lymphocytes more potently compared with control DCs. These results suggest that LEN enhanced the function of DCs generated from patients with multiple myeloma by stimulating the capacity of allogeneic T cells, inhibiting the generation of immunosuppressive cells, inducing naïve T cells toward Th1 polarization, and generating potent myeloma-specific cytotoxic T lymphocytes.