B Cell Maturation Antigen Expression Research Articles

Antigen escape as a shared mechanism of resistance to BCMA-directed therapies in multiple myeloma

B-cell maturation antigen (BCMA)-targeting therapeutics have dramatically improved outcomes in relapsed/refractory multiple myeloma (RRMM). However, whether the mechanisms of resistance between these therapies are shared and how the identification of such mechanisms before therapy initiation could refine clinical decision-making remains undefined. We analyzed outcomes for 72 RRMM patients treated with teclistamab, a CD3 x BCMA bispecific antibody (BsAb), 42% (30/72) of whom had prior BCMA-directed therapy exposure. Malignant plasma cell BCMA expression was present in all BCMA therapy-naïve patients. Prior therapy-mediated loss of plasma cell BCMA expression before teclistamab treatment, measured by immunohistochemistry, was observed in 3 patients, none of whom responded to teclistamab, and one of whom also did not respond to ciltacabtagene autoleucel. Whole exome sequencing of tumor DNA from one patient revealed biallelic loss of TNFRSF17 following treatment with belantamab mafodotin. Low-to-undetectable peripheral blood soluble BCMA levels correlated with the absence of BCMA expression by bone marrow plasma cells. Thus, although rare, loss of BCMA expression following TNFRSF17 gene deletions can occur following any BCMA-directed therapy and prevents response to subsequent anti-BCMA-directed treatments, underscoring the importance of verifying the presence of a target antigen.

Correlation of immune fitness with response to teclistamab in relapsed/refractory multiple myeloma in the MajesTEC-1 study

AbstractTeclistamab, an off-the-shelf B-cell maturation antigen (BCMA) × CD3 bispecific antibody that mediates T-cell activation and subsequent lysis of BCMA-expressing myeloma cells, is approved for the treatment of patients with relapsed/refractory multiple myeloma (R/RMM). As a T-cell redirection therapy, clinical outcomes with teclistamab may be influenced by patient immune fitness and tumor antigen expression. We correlated tumor characteristics and baseline immune profiles with clinical response and disease burden in patients with R/RMM from the pivotal phase 1/2 MajesTEC-1 study, focusing on patients treated with 1.5 mg/kg of teclistamab (N = 165). Peripheral blood samples were collected at screening, and bone marrow samples were collected at screening and cycle 3. Better clinical outcomes to teclistamab correlated with higher baseline total T-cell counts in the periphery. In addition, responders (partial response or better) had a lower proportion of immunosuppressive regulatory T cells (Tregs), T cells expressing coinhibitory receptors (CD38, PD-1, and PD-1/TIM-3), and soluble BCMA and a T-cell profile suggestive of a more cytolytic potential, compared with nonresponders. Neither frequency of baseline bone marrow BCMA expression nor BCMA-receptor density was associated with clinical response to teclistamab. Improved progression-free survival was observed in patients with a lower frequency of T cells expressing exhaustion markers and immunosuppressive Tregs. Overall, response to teclistamab was associated with baseline immune fitness; nonresponders had immune profiles suggestive of immune suppression and T-cell dysfunction. These findings illustrate the importance of the contribution of the immune landscape to T-cell redirection therapy response. This trial was registered at www.ClinicalTrials.gov as #NCT03145181/NCT04557098.

IL-18-secreting multiantigen targeting CAR T cells eliminate antigen-low myeloma in an immunocompetent mouse model

AbstractMultiple myeloma is a plasma cell malignancy that is currently incurable with conventional therapies. Following the success of CD19-targeted chimeric antigen receptor (CAR) T cells in leukemia and lymphoma, CAR T cells targeting B-cell maturation antigen (BCMA) more recently demonstrated impressive activity in relapsed and refractory myeloma patients. However, BCMA-directed therapy can fail due to low expression of BCMA on myeloma cells, suggesting that novel approaches to better address antigen-low disease may improve patient outcomes. We hypothesized that engineered secretion of the proinflammatory cytokine interleukin-18 (IL-18) and multiantigen targeting could improve CAR T-cell activity against BCMA-low myeloma. In a syngeneic murine model of myeloma, CAR T cells targeting the myeloma-associated antigens BCMA and B-cell activating factor (BAFF-R) failed to eliminate myeloma when these antigens were weakly expressed, whereas IL-18–secreting CAR T cells targeting these antigens promoted myeloma clearance. IL-18-secreting CAR T cells developed an effector-like T-cell phenotype, promoted interferon-gamma production, reprogrammed the myeloma bone marrow microenvironment through type-I/II interferon signaling, and activated macrophages to mediate antimyeloma activity. Simultaneous targeting of weakly-expressed BCMA and BAFF-R with dual-CAR T cells enhanced T-cell:target-cell avidity, increased overall CAR signal strength, and stimulated antimyeloma activity. Dual-antigen targeting augmented CAR T-cell secretion of engineered IL-18 and facilitated elimination of larger myeloma burdens in vivo. Our results demonstrate that combination of engineered IL-18 secretion and multiantigen targeting can eliminate myeloma with weak antigen expression through distinct mechanisms.

CD4+ CAR T-cell exhaustion associated with early relapse of multiple myeloma after BCMA CAR T-cell therapy

AbstractMultiple myeloma is characterized by frequent clinical relapses after conventional therapy. Recently, chimeric antigen receptor (CAR) T cells targeting B-cell maturation antigen (BCMA) has been established as a treatment option for patients with relapsed or refractory disease. However, although >70% of patients initially respond to this treatment, clinical relapse and disease progression occur in most cases. Recent studies showed persistent expression of BCMA at the time of relapse, indicating that immune-intrinsic mechanisms may contribute to this resistance. Although there were no preexisting T-cell features associated with clinical outcomes, we found that patients with a durable response to CAR T-cell treatment had greater persistence of their CAR T cells than patients with transient clinical responses. They also possessed a significantly higher proportion of CD8+ T effector memory cells. In contrast, patients with short-lived responses to treatment have increased frequencies of cytotoxic CD4+ CAR T cells. These cells expand in vivo early after infusion but express exhaustion markers (Hepatitis A virus cellular receptor 2 [HAVCR2] and T cell immunoglobulin and mucin domain-containing-3 [TIGIT]) and remain polyclonal. Finally, we demonstrate that nonclassical monocytes are enriched in the myeloma niche and may induce CAR T-cell dysfunction through mechanisms that include transforming growth factor β. These findings shed new light on the role of cytotoxic CD4+ T cells in disease progression after CAR T-cell therapy.

Options at the time of relapse after anti-BCMA therapy.

B-cell maturation antigen (BCMA)-directed therapies, including antibody-drug conjugates, bispecific antibodies (BsAbs), and chimeric antigen receptor T cells (CARTs), have shown remarkable efficacy in patients with late-line myeloma with prior exposure to immunomodulatory agents, proteasome inhibitors, and anti-CD38 antibodies. However, optimal sequencing of these agents remains to be determined, and management of these patients once they relapse has become a new unmet need. Fortunately, there are multiple options with demonstrated activity after anti-BCMA therapy, including a different BCMA-directed therapy, non-BCMA-directed CARTs and BsAbs, novel non-T-cell-engaging drugs, and standard triplet/quadruplet regimens or salvage stem cell transplant. Factors to consider when choosing a next therapy after anti-BCMA therapy include patient characteristics and preferences, prior therapies and toxicities, disease biology, timing from last anti-BCMA therapy, and, in the future, BCMA expression and immune profiling. While current data are limited to retrospective studies and small prospective cohorts, the serial use of T-cell-engaging therapies looks particularly promising, especially as BCMA-directed therapies move up earlier in the myeloma treatment course and additional CARTs and BsAbs against alternative targets (eg, G protein-coupled receptor, family C, group 5, member D and Fc receptor-homolog 5) become available. Going forward, ongoing prospective studies, large real-world data sets, and better tools to interrogate antigen expression and immune cell fitness hopefully will provide further insight into how to best individualize therapy for this difficult-to-treat population.

Identification of HLA‐restricted Cytotoxic T Lymphocyte Epitopes of B Cell Maturation Antigen as Promising Peptide Vaccine Candidates for Preventive and Therapeutic Strategies in Multiple Myeloma

AbstractMultiple myeloma (MM) is an incurable hematologic malignancy for which curative treatment remains elusive. Consequently, immunotherapeutic strategies that selectively induce MM‐specific T cell responses by targeting MM‐associated antigens have emerged. This study aims to identify HLA‐restricted cytotoxic T lymphocyte (CTL) epitopes derived from B cell maturation antigen (BCMA) as potential peptide vaccine candidates to advance preventive and therapeutic interventions for MM. BCMA expression in bone marrow tissue samples from 96.8% of MM patients is observed, and the positive membrane BCMA expression is associated with inferior overall survival outcomes. Through an in silico model, four BCMA epitope peptides that exhibit in vitro concentration‐dependent binding to the HLA*A11:01 molecule are identified. Comparative analyses reveal that BCMA‐specific CTLs stimulated by BCMA‐2, BCMA‐3, and BCMA‐4 peptides exhibit significantly greater cytolysis of cancer cells, including BCMA‐overexpressing KMS‐20 and KKU‐055 cells, in comparison to an unpulsed condition or an irrelevant control peptide derived from human immunodeficiency virus‐1 (HIV‐1). Moreover, the observed cytolytic activities of these CTLs are significantly distinct when compared to corresponding BCMA‐negative cells, thereby indicating antigen‐specificity. This study highlights the promising potential of utilizing immunogenic HLA‐A*11:01‐restricted BCMA peptides as a viable approach for cancer vaccination and T cell‐based therapy in the context of MM.

B-Cell Maturation Antigen (BCMA) Expression and Clinical Features of Plasmablastic Lymphoma (PBL)

Introduction: Plasmablastic lymphoma (PBL) is a rare and aggressive subtype of large B-cell lymphoma with plasmacytic differentiation. Although limited-stage (LS) PBL has a favorable outcome, extensive-stage (ES) PBL has a poor prognosis, with a median survival of 12-18 months. Patients with relapsed/refractory (r/r) PBL have limited treatment options and most die from the disease. B cell maturation antigen (BCMA) - directed cellular and immunotherapies are highly effective in MM. The data on BCMA expression in PBL is limited. Literature review identified one report including 7 PBL cases all of which expressed BCMA. We studied all patients with a diagnosis of PBL who presented to Moffitt Cancer Center in Florida, USA from Jan. 2006 to Apr. 2023, and retrospectively performed BCMA immunohistochemistry (IHC) staining on the available archived FFPE biopsy samples. Methods: Progression-free survival (PFS) was calculated as time from disease progression or last follow up to treatment start date. Overall survival (OS) was calculated as time from death or last follow up to date of PBL diagnosis. Association of BCMA expression with PFS or OS was evaluated using logrank test, with continuous variables converted to categorical by classifying as above or below median. All analyses were done using SAS 9.2. Results: A total of 32 PBL patients were identified. The median age was 59 years (range 31-83 years); 21 (65.6%) had stage III/IV (ES); 9 patients (28.1%) were HIV positive; and 57.1% of evaluated cases were EBV positive by EBER. Treatment data was available for 30 patients. The most common frontline therapies were EPOCH and CHOP with or without rituximab; four patients received bortezomib combined with EPOCH; 7 patients (23.3%) received autoSCT in 1 st remission (Table 1). Median follow-up was 77.6 months calculated using the reverse Kaplan-Meier method. Consistent with previous reports, patients with ES PBL had worse outcome than those with LS PBL, with 3-year OS rates of 24% and 100%, respectively, p=0.01 (Table 1). Among the 20 patients with ES PBL and known treatment responses, 11 did not achieve CR after frontline therapy. Ten of these 11 patients died due to disease progression, 9 within 2 years and 1 within 3 years. Two patients received CD19 CAR-T after multiple lines of therapy. Both died within 30 days, one from disease progression and the other from multiple treatment complications. Out of the 18 cases with tissue available for IHC study, 17 (94.4%) had positive BCMA expression (Figure 1). The median percentage of BCMA+ cells was 75% (Table 1). Eleven (61.1%) cases had intermediate to strong BCMA staining. Neither BCMA+ cell percentage nor BCMA expression level was associated with disease stage, response rate, PFS, or OS (p>0.10 for all tests). Discussion: In our study the majority of our PBL cases tested positive for BCMA, and the percentage of BCMA+ cells seemed comparable to MM cases.To our knowledge, this is the largest and the first report that systemically evaluated BCMA expression in PBL. Our results provide evidence to explore BCMA as a potential target in PBL. Currently two anti-BCMA CAR-T products and an anti-BCMA/CD3 bispecific T-cell engager have been approved for r/r MM. A BCMA-targeting antibody-drug conjugate (ADC) Belantamab mafodotin-blmf was initially granted accelerated approval by FDA for r/r MM but was later voluntarily withdrawn from the market. Given the expression of BCMA in PBL and the lack of effective treatment of r/r PBL, these patients could be considered in trials evaluating the BCMA-targeting therapies. In fact, in a case report recently published, one patient with refractory PBL achieved CR after one infusion of BCMA CAR-T and remained in CR after 14 months. Ten of our 11 ES patients who did not achieve CR after frontline chemotherapy died from the disease, emphasizing the importance of clinical trials for this condition. We did not find an association of BCMA expression with clinical outcomes as oppose of MM, in which higher BCMA expression was associated with shorter PFS and OS. The lack of association in our study may be due to the small sample size, or, it may reflect a difference in biological role of BCMA in PBL from MM. Our results will need to be corroborated with other series to confirm our findings. Translational studies need to evaluate the dynamics of BCMA protein expression on PBL cell membrane and the activity of BCMA-targeting agents in vitro and in animal models of PBL.

Regulation of BCMA By the Ubiquitin Proteasome System Enables Optimization of BCMA-Targeting Therapies in Multiple Myeloma

Background: The exclusive expression of the B-cell maturation antigen (BCMA), in normal and malignant plasma cells, currently makes it a prominent target for immune cell therapies in multiple myeloma (MM) including chimeric antigen receptor T-cell (CART) and bi-specific T cell engaging antibody (BiTE) therapies. Despite initial high response rates, most patients eventually relapse highlighting the need to enhance therapy efficacy. Although loss of BCMA-antigen expression marks one reason for therapy failure, little is known about the mechanistic means of antigen escape. The ubiquitin proteasome system (UPS) controls abundance and protein degradation of about 80-90% of all cellular proteins. Protein degradation is initiated by substrate ubiquitination, which is orchestrated by the sequential action of enzymes whereby only E3-ligases possesses substrate specificity. Deubiquitinating enzymes (DUBs) can reverse substrate ubiquitination. Against this background, we raised the question whether BCMA antigen presentation, expression, and activity is regulated by the UPS and whether the UPS can be utilized to enhance BCMA directed CART therapies in MM. Methods: Protein turnover and regulation by the UPS was assayed using cycloheximide and proteasomal inhibitors (PI) followed by immunoblotting. Direct substrate ubiquitylation was confirmed by ubiquitylation assays based on immunoprecipitation and image-based analysis. Changes in antigen surface presentation were determined using FACS analysis. Changes in cellular signaling and downstream effects were assessed by immunoblotting and quantitative PCR (qPCR). Primary CD138 + cells were obtained from bone marrow aspirations followed by MACS ® cell separation. To assess the effect of PI on CART killing capacity, a classic co-culture was set up in vitro. Using a proximity-based protein purification method (TurboID), UPS-related interactors of BCMA were identified. Results: For the first time, we show that BCMA is a short-lived protein with a half-life of approximately 30 min in MM cells. Degradation of BCMA is abrogated upon PI treatment. Inhibition of γ-secretase, an established protease of the extracellular domain of BCMA does not impair BCMA turnover. Furthermore, we show that BCMA is indeed ubiquitylated in MM cells and that ubiquitylation is induced by the ligand APRIL. These results support the hypothesis that the UPS regulates BCMA abundance. In both MM cell line models and primary CD138 + MM cells of PI refractory/ relapsed MM (RRMM) patients, inhibition of the proteasome leads to a significant increase of BCMA on the cell surface without promoting oncogenic NF-κB signaling in MM cells. Subsequently, we show that the increase in BCMA surface expression upon PI treatment substantially increases the efficacy of BCMA directed CART cells in co-cultures with MM cells and primary CD138 + MM patient cells. Specific members of the UPS (E3 ligases and DUBs) identified from our proteome wide BCMA interactome analysis are currently being validated. Conclusion: Here we show, for the first time, that BCMA is ubiquitylated and degraded by the ubiquitin proteasome system in multiple myeloma. The increase in BCMA expression upon proteasomal inhibition enhanced efficacy of BCMA-targeting CART cells. These insights provide a rationale for combining BCMA CART cells with PI in patients with BCMA CART relapsed/refractory disease and persistence. The identification of BCMA-specific E3-ligases and DUBs may represent novel therapeutic targets to specifically prevent BCMA degradation and antigen escape.

Longitudinal Correlative Profiles of Responders, Nonresponders, and Those with Relapse on Treatment with Teclistamab in the Phase 1/2 MajesTEC-1 Study of Patients with Relapsed/Refractory Multiple Myeloma

Introduction: Teclistamab (tec) is the only approved B-cell maturation antigen (BCMA) × CD3 bispecific antibody with a personalized, weight-based dosing schedule for the treatment of triple-class exposed relapsed/refractory multiple myeloma (RRMM). In the phase 1/2 MajesTEC-1 study (NCT03145181/NCT04557098), tec demonstrated deep and durable responses. To better understand mechanisms of resistance and relapse in MajesTEC-1, we assessed longitudinal BCMA expression and immune profiles. Methods: Patients (N=165) with RRMM received subcutaneous tec 1.5 mg/kg weekly after step-up doses of 0.06 and 0.3 mg/kg, with the option to switch to every-other-week dosing if they achieved at least a partial response after ≥4 cycles of therapy in phase 1 or a complete response or better for ≥6 months in phase 2. Bone marrow and peripheral blood samples at baseline, on treatment, and at disease progression (PD) were analyzed by flow cytometry for BCMA expression and immune cell populations. Bone marrow aspirates were analyzed by cytometry by time of flight (CyTOF) for immune cell populations. Soluble BCMA (sBCMA) was analyzed in serum samples by electrochemiluminescence ligand binding. Results: Patients who responded to tec had a greater recovery of CD3+ T cells in the periphery and bone marrow during the first treatment cycles, which was sustained over time compared with nonresponders. Greater activation during the first cycle was observed in responders, indicated by a transient increase in CD38 on CD8+ T cells. In contrast, an exhausted T-cell phenotype in the blood and bone marrow was observed longitudinally in nonresponders vs responders, indicated by increasing and sustained levels of activation and exhaustion markers including persistence of CD38+ T cells and expression of LAG-3, PD-1 (Figure 1A), PD-1/LAG-3, and PD-1/TIM-3 on CD4+ or CD8+ T cells. Higher levels of immunosuppressive regulatory T cells (Tregs), including CD38+ Tregs, were also sustained in the periphery in nonresponders compared with responders. BCMA expression and sBCMA levels were assessed in a subset of patients who initially responded then relapsed on tec, who had matched baseline and end-of-treatment/PD samples. There were no significant changes in the frequency of BCMA+ plasma cells, BCMA receptor density, or sBCMA levels at PD relative to baseline, suggesting BCMA loss was not a mechanism of relapse in these patients. In contrast, higher proportions of peripheral CD4+ and/or CD8+ T cells expressing CD38, TIM-3, PD-1, and PD-1/TIM-3 were observed at relapse vs baseline. Analysis of bone marrow progression samples using CyTOF in patients who relapsed showed significantly lower CD28 expression on CD8+ T cells, significantly higher expression of exhaustion markers (CD38, PD-1, TIM-3, EOMES, and TOX) and perforin on CD4+ and/or CD8+ T cells, a higher frequency of CD4+ and CD8+ T cells expressing TIM-3 and co-expressing CD38/TOX, PD-1/TIM-3, and TOX/TIM-3 (Figure 1B), and significantly higher CD38 expression on T-cell receptor gamma delta T cells, at PD than at baseline. PD-1 and TIGIT expression was also significantly higher at PD than at baseline across CD25hiCD127dim and/or CD25hiCD127dimFoxP3+ Tregs in patients who relapsed. Conclusions: Patients responding to tec exhibited a differential immune profile in early cycles compared with nonresponders, with greater transient T-cell activation. In contrast, an exhausted T-cell phenotype was sustained in nonresponders during the first 2 treatment cycles. While mutations in BCMA cannot be excluded, we did not detect BCMA loss as a mechanism of relapse, but observed a dysfunctional immune phenotype at PD, with increased T-cell exhaustion and higher frequency of gamma delta T cells and immunosuppressive Tregs. These results suggest the importance of immune fitness and T-cell function in achieving and maintaining a response to tec. Ongoing studies will evaluate these correlatives in earlier treatment lines, where patients may have more favorable immune profiles.

BCMA Re-Emergence As a Prognostic Indicator of Clinical Relapse after Chimeric Antigen Receptor T-Cell Therapy (CART) in Multiple Myeloma

Background: The outcomes of patients with triple and penta-class relapsed refractory multiple myeloma (RRMM) are poor. B-cell maturation antigen (BCMA)-directed chimeric antigen receptor T-Cell (CAR-T) therapies in RRMM have been shown to improve outcomes since approval in 2021; however, relapses are common. We previously reported that patients lost BCMA expression after BCMA-directed therapy. Early re-emergence of surface BCMA on plasma cells (PC) in bone marrow (BM) may be a prognostic indicator for clinical relapse. Furthermore, there is limited published literature about clinical factors that increase risk of early BCMA re-emergence as well as clinical relapse post-CAR-T therapy. Methods: The study is a prospective cohort study that includes RRMM patients who underwent BCMA CAR-T at the University of Kansas Health System between May 2021 and May 2023 and were followed at regular intervals for the first-year post-infusion. All patients had standard procedure of BM evaluation at 1, 3, and 6 months, and 1 year. Eight-color MFC was performed on the BM specimens. BCMA antibody was purchased from R & D Systems (Minneapolis, MN). Data analysis was performed using FCS Express 5 software (De Novo Software, Los Angeles, CA). 500,000 to up to 2,000,000 events were acquired in all the cases. Responses were evaluated using the International Myeloma Working Group criteria. BCMA re-emergence was defined as BCMA greater than nadir on either polyclonal or monoclonal BM plasma cells. Results: 57 patients were evaluated and 44 patients (77 %) who lost their PC BCMA expression by multicolor flow cytometry (MFC) on bone marrow biopsy (BMBX) at Day 30 from baseline were included in the analysis. Demographic data of all 44 RRMM BCMA CAR-T recipients is shown Table 1. Median follow-up was 7 (4-14) months. Of the 44 patients, BCMA re-emergence was as follows: at 3-months (n=14, 32% of total sample), 6-months (n=5, 11%), 9-months (n=0, 0% of total sample), and 12-months (n=5, 11% of total sample). 14 (32%) of the 44 total patients had clinical relapse within 12 months. Of the 14 patients, 11 (79%) had BCMA re-emergence at any time point. 10 out of 11 (91%) of these patients had BCMA re-emergence before or at the same time as clinical relapse. The median time to BCMA re-emergence was 3 months and the median time to clinical relapse was 6 months. Only 1 patient had clinical relapse prior to BCMA re-emergence.Furthermore, at the 6-month mark, there were no significant clinical or demographic factors between those who relapsed vs. those who had a response to CAR-T therapy other than BCMA re-emergence (p-value 0.0445, Table 1). Conclusion: BCMA re-emergence on BM detected by MFC may be a prognostic indicator for clinical relapse in RRMM recipients of BCMA CAR-T therapies. It may be valuable to carry out close monitoring that includes evaluation of BCMA expression on BMBx at regular intervals post CAR-T therapy. Additional prospective studies with larger cohorts are needed to determine the risk factors for early BCMA re-emergence and clinical relapse in RRMM patients who receive BCMA CAR-T therapy.

The Impact of Soluble BCMA and BCMA Gain on Anti-BCMA Immunotherapies in Multiple Myeloma

B cell maturation antigen (BCMA) targeting chimeric antigen receptor T cells (CAR T) and bispecific T cell engagers (TCE) have remarkable efficacy in multiple myeloma (MM). While BCMA antigen escape, driven by TNFRSF17 deletions or mutations, is a mediator of anti-BCMA CAR T/ TCE resistance, the majority of patients retain BCMA surface expression at progression. Furthermore, patients with high disease burden and extramedullary disease associated with poorer response to anti-BCMA therapies, have high levels of soluble BCMA (sBCMA). The mechanisms mediating this resistance, and in particular the contribution of sBCMA, have not been fully elucidated. In order to assess the effect of sBCMA on anti-BCMA CAR T function, we measured CAR T cells activation with an in-house manufactured anti-BCMA CAR transduced with an NFAT-GFP reporter gene after co-culture with increasing concentrations of sBCMA (0-2500 ng/mL). sBCMA bound to CAR scFv in a dose dependent manner, effectively competing with the fluorophore tagged BCMA peptide binding. Despite their stable engagement, sBCMA failed to induce the expression of GFP, CD69, CD25, or 41BB on CAR T cells. To further examine the functional effect of sBCMA on CAR T cytotoxicity, we generated isogenic OPM2 cell lines with distinct BCMA expression profiles: OPM2 over-expressing BCMA (OPM2_BCMA high) generated by lentiviral transduction of TNFRSF17, and Cas9 BCMA knockout OPM2 (OPM2_BCMA -/-). OPM2_BCMA high cells exhibited one-log-fold increase in surface BCMA expression and 10-fold increase in sBCMA and were more resistant to CAR T mediated lysis. In addition, in co-culture assays, elevated sBCMA impaired OPM2 target cell lysis by anti-BCMA CAR T in a dose dependent-manner. To gain insight into the effect of chronic sBCMA exposure on CAR T, we exposed anti-BCMA CAR T to OPM2 cells secretome in the absence of direct cell to cell contact in a trans-well co-culture system. On day 24, CAR T in co-culture with OPM2_BCMA high (sBCMA levels of 2343 ng/mL) significantly contracted with no viable cells, while CAR T in culture with parental OPM2 (sBCMA level of 267 ng/mL) did persist with up-regulated TIM3 and TIGIT. In contrast CAR T cultured with OPM2_BCMA -/- remained viable with no expression of exhaustion markers. In examining the impact of sBCMA on TCE activity, sBCMA level as low as 50 ng/mL in vitro impaired anti-BCMA TCE binding to K562 cells transduced to stably express BCMA (K562_wtBCMA) . Consistent results were found in cytotoxicity assays wherein the addition of 2 ng/mL of sBCMA was sufficient to induce a reduction in anti-BCMA TCE mediated cytotoxicity of K562_wtBCMA cells by healthy donor peripheral blood mononuclear cells. To identify genomic events that lead to elevated sBCMA expression and subsequent resistance to CAR T/TCE, we conducted bulk whole genome sequencing (100X) on CD138+ bone marrow MM cells obtained from 40 MM patients treated with anti-BCMA CAR T/ TCE. Structural variants (SVs) at or adjacent to TNFRSF17 locus were identified in four patients. These included i) focal copy number gain of TNFRSF17 (3 copies) in a patient who progressed within 3 months post anti-BCMA TCE, ii) subclonal duplication of TNFRSF17 locus in another patient (n=1), as well as iii) focal copy number losses 5‘ or 3‘ to the TNFRSF17 gene locus (n=2). In addition to SVs affecting TNFRSF17 locus, we also identified mutations involving TNFRSF17 transcriptional regulators. Notably, MM clones at progression 4 months post anti-BCMA TCE (n=1) harboured copy number gains (3 copies) of POU2AF1 gene on chr11q23. POU2AF1 encodes a key transcriptional regulator of TNFRSF17 expression. Indeed, sBCMA level from this patient was significantly elevated at relapse at a level (638 ng/mL) correlated with resistance to anti-BCMA TCE. In addition, we identified translocation involving MAP3K14 encoding the NF-kB regulator NIK in a patient with short remission post anti-BCMA CAR T. Of interest, analysis of the CoMMpass dataset revealed significantly higher BCMA transcripts in patients with translocations involving MAP3K14 locus. Our findings highlight the impact of sBCMA levels and its chronicity of exposure on anti-BCMA TCE and CAR T activities in MM. We further identified structural genomic events driving TNFRSF17 over-expression and their correlation to sBCMA levels facilitating tumoral immunotherapeutic escape.

Biomarker Correlates of Response to Ciltacabtagene Autoleucel in Patients with Relapsed or Refractory Multiple Myeloma from CARTITUDE-1, a Phase 1b/2 Open-Label Study, at the ~3 Year Follow-up

Ciltacabtagene autoleucel (cilta-cel) consists of autologous T cells genetically modified to express a two-binding domain chimeric antigen receptor (CAR). The target antigen of the CAR is B-cell maturation antigen (BCMA), which is highly expressed on malignant plasma cells (PC) from multiple myeloma subjects. In the Phase 1b/2 CARTITUDE-1 study, cilta-cel led to early, deep, and durable responses and was approved for the treatment of patients with relapsed or refractory multiple myeloma (RRMM) after four or more prior lines of therapy by FDA. Despite cilta-cel's outstanding overall response rate (97.9%) and duration of response (DOR) in RRMM patients, relapse still occurs. Biomarkers associated with this durable response and acquired resistance remain to be elucidated. We present updated correlative data from the CARTITUDE-1 study. At the ~3 year follow-up (October 2022 data cutoff), 97 patients had received a single infusion of cilta-cel (median 0.71 x10 6 cells/kg [range 0.52 x10 6-0.94 x10 6]); the median follow up was 33.4 months (range 1.5-45.2), median DOR was 33.9 months (95% CI, 25.5-not estimable [NE]) and median PFS was 34.9 months (95% CI, 25.2-NE) (Lin Y et al, ASCO 2023). Drug product (DP), baseline and on-treatment whole blood and bone marrow samples from enrolled patients were analyzed by methods including flow cytometry, MSD immunoassays, Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq) and TCR sequencing. In CARTITUDE-1, the DP contained a mixture of transduced and non-transduced T cells; the median transduction efficiency was 16% (range 5-32%), with a balanced distribution of CAR+CD4+ and CAR+CD8+ cells, median frequency 12% (range 2-28%) and 6% (range 2-20%), respectively. Further, the DP T cell subset composition was variable but balanced between central (Tcm) and effector memory (Tem) phenotypes. CAR+ T cells expanded reaching a median peak concentration (C max of 730 cells/µL; range 3-13805 cells/µL) in blood between days 12-14 post-infusion and persisted in circulation for a median of 100 days (range 20-912 days). High efficacy (best response) and DOR were achieved despite variable CAR-T cell expansion and lack of detectable CAR-T cell persistence over time. At C max, the CD4:CD8 ratio (based on % of CAR+ cells) was 0.29, demonstrating a CAR+CD8+ T cell preferential expansion, and both CAR+CD4+ and CAR+CD8+ T cells were predominantly of a Tcm phenotype, median 95% (range 62-99.5%) and 96% (range 33-99.7%), respectively. Higher frequency of early-memory T cell phenotypes has been associated with clinical responses to CAR-T therapies in MM (Dancy et al., Blood 2018). CITE-seq analyses showed that DP characteristics such as a high CD8+ stem-like phenotype and a low CD4+ Treg-like phenotype in the CAR+ T cell compartment associated with longer DOR. Some of the most abundant clones in the DP single cell TCR data were CAR+CD8+ T cells expressing granzyme B, suggesting high cytotoxic potential. Additional bulk TCR sequencing data suggest both diversity and individual clonal expansion longitudinally after cilta-cel infusion. We also evaluated tumor and patient intrinsic characteristics at baseline. BCMA expression on bone marrow PC was prevalent and highly variable among RRMM patients; however, expression did not associate with best response or DOR. Further, high ORR was observed in patients with high-risk cytogenetics, high tumor burden (>= 60% bone marrow PC), or baseline plasmacytomas, although with a non-statistically significant shorter PFS. Sustained MRD negativity strongly associated with longer DOR. Conversely, patient-intrinsic characteristics associated with inflammation (CRP, ferritin, neutrophils, pro-inflammatory cytokines) correlated with shorter DOR. Treatment benefit of the magnitude seen with cilta-cel had not been reported with any other approved therapy in this setting. Correlative analyses suggest an emerging profile to help understand such outstanding clinical activity: a balanced CD4/CD8 ratio and Tcm/Tem phenotype of the manufactured DP and preferential expansion of CAR+CD8+ Tcm cells. Further, certain baseline patient-intrinsic and tumor characteristics may explain the longer DOR in the RRMM setting. These investigations help identify markers of response to cilta-cel and may lead to CAR-T cell design or manufacturing strategies that enhance drug product characteristics and thus clinical efficacy.

CD8 effector T cells enhance teclistamab response in BCMA-exposed and -naïve multiple myeloma

AbstractTeclistamab, a B-cell maturation antigen (BCMA)- and CD3–targeting bispecific antibody, is an effective novel treatment for relapsed/refractory multiple myeloma (R/RMM), but efficacy in patients exposed to BCMA-directed therapies and mechanisms of resistance have yet to be fully delineated. We conducted a real-world retrospective study of commercial teclistamab, capturing both clinical outcomes and immune correlates of treatment response in a cohort of patients (n = 52) with advanced R/RMM. Teclistamab was highly effective with an overall response rate (ORR) of 64%, including an ORR of 50% for patients with prior anti-BCMA therapy. Pretreatment plasma cell BCMA expression levels had no bearing on response. However, comprehensive pretreatment immune profiling identified that effector CD8+ T-cell populations were associated with response to therapy and a regulatory T-cell population associated with nonresponse, indicating a contribution of immune status in outcomes with potential utility as a biomarker signature to guide patient management.

Abstract 5081: Anti-tumor activity of HPN217, a BCMA-targeting tri-specific T cell engager, is enhanced by γ-secretase inhibitors in preclinical models

Abstract HPN217 is a Tri-specific T Cell-Activating Construct (TriTAC) currently being evaluated in a phase 1 clinical trial for relapsed or refractory multiple myeloma (MM) (NCT04184050). It consists of a single domain antibody (sdAb) specific for B-cell maturation antigen (BCMA), a serum albumin-specific sdAb for half-life extension, and a single chain Fv (scFv) specific for the CD3ε subunit of the T cell receptor (TCR) complex for T cell engagement. BCMA is a cell surface receptor highly expressed on malignant plasma cells in MM patients and plays an important role in B-cell proliferation and survival. Soluble BCMA (sBCMA) is produced when the extracellular domain of BCMA is cleaved by γ-secretase and enters circulation. sBCMA may act as a sink for BCMA targeted therapies, potentially decreasing anti-tumor efficacy. Previous studies have shown γ-secretase inhibitors (GSIs) can increase membrane bound BCMA expression on MM cells and decrease sBCMA concentrations, providing a rationale for combining GSIs with HPN217. Here we describe preclinical studies evaluating that combination in vitro and in vivo. To evaluate the effects of GSIs on BCMA expression, MM cell lines MOLP8, KMS-12-BM, and MM.1S were incubated with GSIs LY3039478, PF03084014, or RO4929097overnight. GSI treatment led to a 3-7-fold increase in BCMA expression on MM cell surface. To determine the effects of increased BCMA expression mediated by GSIs on the activity of HPN217, an in vitro T cell dependent cell cytotoxicity assay (TDCC) was conducted. GSIs enhanced the activity of HPN217, in some cases leading to a 3.5-fold increase in potency compared to HPN217 alone. Next, the impact of the combination was tested in vivo using a disseminated MOLP-8 xenograft model engrafted with T cells from a healthy human donor. Vehicle treated mice succumbed to disease burden with a median survival of 23 days as assessed by body weight loss and clinical observations. A suboptimal dose of 4 ug/kg HPN217 protected mice from disease burden for an additional 3 days. By contrast, the combination of HPN217 and LY3039478 led to a significant increase in survival compared to vehicle or either monotherapy, with a median survival of 43 days. Taken together these results demonstrate the potential utility of GSIs to enhance the anti-tumor activity of HPN217 and supports further investigation of this combinatorial approach in patients. Citation Format: Payton C. Laurie, Mary Ellen Molloy, Patrick P. Ng, Banmeet S. Anand. Anti-tumor activity of HPN217, a BCMA-targeting tri-specific T cell engager, is enhanced by γ-secretase inhibitors in preclinical models. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5081.

Abstract 2960: The novel trifunctional anti-BCMA NK cell engager SAR’514 has potent in-vitro, in-vivo and ex-vivo anti-myeloma effect through dual NK cell engagement

Abstract Multiple Myeloma (MM) is the second most prevalent hematopoietic malignancy, representing 10% of total blood cancers. Despite the emergence of new therapies, it is still an incurable disease. Safe, potent and innovative approaches with long lasting beneficial effects are required. BCMA (B Cell Maturation Antigen) is a cell surface receptor selectively expressed on normal and malignant plasma cells and promote cell proliferation and survival upon binding of its ligands APRIL (A PRoliferation Inducing Ligand) and BAFF (B cell Activating Factor). BCMA expression is highly prevalent on myeloma tumor cells and is maintained after standard of care treatments such as anti-CD38 therapies, or even BCMA-targeting agents (Cohen AD et al. J Clin Invest 2019). The ability of NK cells to intrinsically kill tumor cells, leaving healthy cells unharmed, with minimal pro-inflammatory cytokine release induction as compared to T cell-based therapies makes NK cells ideal immune cells for a safe and efficacious therapeutic approach. We developed SAR’514, a trifunctional NK Cell Engager (NKCE) that activates NK cells through a dual engagement of NKp46 and CD16a, two major NK cell activating receptors highly expressed on NK cells in MM patients, and which redirects the activated NK cells to engage and kill BCMA+ tumor cells. We demonstrated that SAR’514 NK dual engagement was more potent than the single NK engagement with NKp46 or CD16a as well as the combination of NKp46 and CD16a engagement. SAR’514 leads to NK cell activation, degranulation and release of effector cytokines only in the presence of BCMA+ tumor cells. This antitumor activity is associated with very low IL-1β, IL-6, TNFα and IFNγ cytokine release as compared to a T cell engager targeting the same BCMA antigen, in PBMC and in whole blood settings in the presence of BCMA+ tumor target cells. In addition, the in vivo anti-tumor activity of an anti-murine NKp46 surrogate NKCE molecule was investigated in huFcgR transgenic mice engrafted with the EL4-huBCMA murine thymoma model. SAR’514 induced a significant mouse survival at 0.5 to 5 mg/kg with an overall survival of 90% as compared to the control group in which only 20% of mice survived. Importantly, SAR’514 exhibits ex vivo efficacy using bone marrow mononuclear cells (BMMC) from MM patients in an autologous setting showing an active and efficient primary MM cell killing against MM cells from patients that have failed diverse therapies, including standard of care treatments. In summary, these results demonstrate the efficacy of SAR’514 for controlling MM tumors in vivo and ex vivo, and provide consistent support for its clinical development. Citation Format: Alexandre Tang, Laurent Gauthier, Jochen Beninga, Benjamin Rossi, Nicolas Gourdin, Audrey Blanchard-Alvarez, Céline Amara, Jacqueline Courta, Alexandra Basset, Dorothée Bourges, Alexandre Desjonqueres, Emmanuelle Menoret, Catherine Pellat-Deceunynck, Philippe Moreau, Yannis Morel, Marielle Chiron, Eric Vivier, Angela Virone-Oddos. The novel trifunctional anti-BCMA NK cell engager SAR’514 has potent in-vitro, in-vivo and ex-vivo anti-myeloma effect through dual NK cell engagement [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2960.

The Effect of Belantamab Mafodotin on Primary Myeloma–Stroma Co-Cultures: Asymmetrical Mitochondrial Transfer between Myeloma Cells and Autologous Bone Marrow Stromal Cells

Belantamab mafodotin (belamaf) is an afucosylated monoclonal antibody conjugated to the microtubule disrupter monomethyl auristatin-F (MMAF) that targets B cell maturation antigen (BCMA) on the surface of malignant plasma cells. Belamaf can eliminate myeloma cells (MMs) through several mechanisms. On the one hand, in addition to inhibiting BCMA-receptor signaling and cell survival, intracellularly released MMAF disrupts tubulin polymerization and causes cell cycle arrest. On the other hand, belamaf induces effector cell-mediated tumor cell lysis via antibody-dependent cellular cytotoxicity and antibody-dependent cellular phagocytosis. In our in vitro co-culture model, the consequences of the first mentioned mechanism can be investigated: belamaf binds to BCMA, reduces the proliferation and survival of MMs, and then enters the lysosomes of malignant cells, where MMAF is released. The MMAF payload causes a cell cycle arrest at the DNA damage checkpoint between the G2 and M phases, resulting in caspase-3-dependent apoptosis. Here, we show that primary MMs isolated from different patients can vary widely in terms of BCMA expression level, and inadequate expression is associated with extremely high resistance to belamaf according to our cytotoxicity assay. We also reveal that primary MMs respond to increasing concentrations of belamaf by enhancing the incorporation of mitochondria from autologous bone marrow stromal cells (BM-MSCs), and as a consequence, MMs become more resistant to belamaf in this way, which is similar to other medications we have analyzed previously in this regard, such as proteasome inhibitor carfilzomib or the BCL-2 inhibitor venetoclax. The remarkable resistance against belamaf observed in the case of certain primary myeloma cell cultures is a cause for concern and points towards the use of combination therapies to overcome the risk of antigen escape.

Hypogammaglobulinemia and Risk of Infectious Complications in Patients Receiving Idecabtagene vicleucel

Idecabtagene vicleucel (ide-cel) is a B-cell maturation antigen (BCMA)-directed chimeric antigen receptor T-cell therapy approved for treatment of relapsed/refractory multiple myeloma. Given BCMA expression on plasma cells, ide-cel has the potential for significant hypogammaglobulinemia and infections, however there is limited data to date describing this association.

A Novel BCMA Immunohistochemistry Assay Reveals a Heterogenous and Dynamic BCMA Expression Profile in Multiple Myeloma

B-cell maturation antigen (BCMA) is a promising target for the treatment of multiple myeloma (MM) because the expression of this protein is largely limited to B-cell sets, plasma cells, MM, and other B-cell malignancies. Early studies assessing BCMA protein expression and localization have used insufficiently qualified immunohistochemistry assays, which have reported broad ranges of BCMA expression. As a result, our understanding of BCMA tissue expression derived from these data is limited, specifically the prevalence of BCMA expression on the cell surface/membrane, which has mechanistic relevance to the antimyeloma activity of several novel biotherapeutics. Here, we report on the qualification and application of a novel anti-BCMA immunohistochemistry antibody, 805G12. This antibody shows robust detection of BCMA in formalin-fixed, decalcified bone marrow tissue and provides key insights into membrane BCMA expression. The clone 805G12, which was raised against an intracellular C-terminal domain peptide of membrane BCMA, exhibited increased sensitivity and superior specificity across healthy and diseased tissue compared with the frequently referenced commercial reagent AF193. The new clone also demonstrated a broad range of expression of BCMA in MM and diffuse large B-cell lymphoma specimens. Additionally, cross-reactivity with closely related tumor necrosis factor receptor family members was observed with AF193 but not with 805G12. Furthermore, via established 805G12 and other independent BCMA assays, it was concluded that proteolytic processing by γ-secretase contributes to the levels of BCMA localized to the plasma membrane. As BCMA-directed therapeutics emerge to address the need for more effective treatment in the relapsed or refractory MM disease setting, the implementation of a qualified assay would ensure that reliable and consistent data on BCMA surface expression are used to inform clinical trial decisions and patient responses.

Elranatamab, a BCMA Targeted T-Cell Engaging Bispecific Antibody, Induces Durable Clinical and Molecular Responses for Patients with Relapsed or Refractory Multiple Myeloma

Introduction: Multiple myeloma (MM) is a plasma cell dyscrasia characterized by expression of B-cell maturation antigen (BCMA). Elranatamab (PF-06863135), a bispecific antibody that targets BCMA and CD3 on T-cells, activates and redirects the T-cell mediated immune response against MM. This clinical trial, MagnetisMM-1 (NCT03269136), is the ongoing Phase 1 first-in-human study evaluating the safety, pharmacokinetics (PK), pharmacodynamics, and efficacy of elranatamab for patients (pts) with relapsed or refractory MM. Methods: Elranatamab was administered subcutaneously (SC) at doses from 80 to 1000µg/kg either weekly or every 2 weeks. Treatment-emergent adverse events (TEAEs) were graded by National Cancer Institute Common Terminology Criteria for Adverse Events (v4.03), whereas cytokine release syndrome (CRS) was assessed by criteria of the American Society for Transplantation and Cellular Therapy. PK, cytokines, lymphocyte subsets, and soluble BCMA were analyzed over time. Clinical response was assessed by criteria of the International Myeloma Working Group (IMWG). Minimal residual disease (MRD) was assessed by next generation sequencing at a sensitivity of 1×10-5 in accordance with IMWG criteria. Results: A total of 55 pts received single-agent elranatamab SC at a dose ≥215μg/kg as of 22-Jun-2022. Median age was 64 (range 42-80) years, and 27% of pts were Black/African American or Asian. Median number of prior regimens was 5 (range 2-14), 91% were triple-class refractory, 69% had prior stem cell transplantation, 29% had high cytogenetic risk, and 24% received prior BCMA-targeted therapy. The most common TEAEs regardless of causality included CRS, neutropenia, anemia, injection site reaction, and lymphopenia. With pre-medication and a single priming dose (600µg/kg or 44mg), the overall incidence of CRS at the recommended dose (1000µg/kg or 76mg) was 67% and limited to Grade 1 (33%) or Grade 2 (33%), with no events Grade 3 or higher. Elranatamab exposure was dose dependent. Cytokine increases occurred with the first dose and were reduced by pre-medication. Soluble BCMA decreased with disease response, elranatamab therapy was associated with increased peripheral T-cell proliferation, and median time to response was 36 days (range 7-262). With a median follow-up of 12.0 months (range 0.3-29.0) and including only IMWG confirmed responses, the objective response rate (ORR) was 64% (95% CI 50-75%) with 56% of pts (31/55) achieving very good partial response (VGPR) or better and 38% of pts (21/55) achieving complete response (CR) or better. Among 13 pts with prior BCMA-directed therapy (antibody drug conjugate, chimeric antigen receptor T-cell therapy, or both), 54% (7/13) achieved response including 46% (6/13) with VGPR or better. For responders (n=35), the probability of being event-free at 12 months was 59% (95% CI 39-74%) and the Kaplan-Meier estimate for median duration of response was 17.1 months (95% CI 10.6-NE). Single-agent elranatamab induced durable clinical and molecular responses, and 100% (12/12) of evaluable pts with confirmed CR or better achieved MRD negativity at a sensitivity of 1×10-5 including 2 pts with MRD negativity and ongoing stringent CR beyond 2 years. Conclusions: Elranatamab induced durable clinical and molecular responses for pts with relapsed or refractory MM. The ORR was 64% with more than half of these pts (38%) achieving CR or better, and 100% of evaluable pts achieved MRD negativity. Together with emerging data from pivotal studies MagnetisMM-3 (NCT0469359) and MagnetisMM-5 (NCT05020236), these results support further development of elranatamab for patients with MM.

Trial in Progress: REGN5458, a BCMAxCD3 Bispecific Antibody, in a Phase Ib Multi-Cohort Study of Combination Regimens for Patients with Relapsed/Refractory Multiple Myeloma

Background Despite the increased effectiveness of combination drug therapy, multiple myeloma (MM) remains incurable, and patients eventually succumb to relapsed disease. With each subsequent line of therapy, relapsed/refractory MM (RRMM) becomes more challenging to treat as high-quality responses become harder to achieve. Therefore, there remains a significant unmet need for new combination regimens that leverage drugs with novel mechanisms of action to improve outcomes in MM. REGN5458 is a bispecific antibody capable of binding to B-cell maturation antigen (BCMA) on MM cells and CD3 on T cells, inducing targeted T-cell-mediated cytotoxicity of MM cells. Results from a Phase I dose escalation study of REGN5458 monotherapy in patients with RRMM showed early, deep, and durable responses with a manageable safety and tolerability profile (NCT03761108; Zonder, et al. EHA 2022. Abstract S189). Among patients treated at the 200-800 mg dose levels, the response rate was 75%. Cytokine release syndrome was reported in 38% of patients, and the severity was mostly Grade 1 with no Grade ≥3 events. Given the encouraging early efficacy and limited overlapping toxicity of REGN5458, it is reasonable to explore the potential benefit of combining REGN5458 with other anti-myeloma agents, with the aim of improving the depth and duration of responses. Study Design and Methods This global, Phase Ib, open-label, multi-cohort study (NCT05137054) is designed to assess the safety, tolerability, and efficacy of REGN5458 in combination with other cancer treatments in patients with RRMM. There are 4 cohorts (Figure panel A), where REGN5458 will be combined with daratumumab plus dexamethasone (cohort 1); carfilzomib plus dexamethasone (cohort 2); lenalidomide plus dexamethasone (cohort 3); and bortezomib plus dexamethasone (cohort 4). Eligible patients cannot be refractory to the cohort-specific combination treatment, but prior exposure is permitted provided at least a 6-month washout period has elapsed. The study will include a dose-finding portion to select an appropriate REGN5458 dose, followed by a dose-expansion portion for each cohort. REGN5458 will be administered intravenously on a weekly basis starting with a step-up dosing schedule. After 15-16 weeks, the dosing frequency will be reduced to every 2-3 weeks depending on the cohort. The other cancer treatments will be administered at the approved doses. All regimens will be given until disease progression or any other reason for discontinuation (Figure panel B). Estimated total enrollment is up to 256 patients. The study will accrue patients with RRMM that have progressed after ≥3 lines of therapy, or ≥2 lines of therapy and either prior exposure to at least 1 anti-CD38 antibody, 1 immunomodulatory drug (IMiD), and 1 proteasome inhibitor (PI) or double refractory to 1 PI and 1 IMiD. Eligible patients must be aged ≥18 years, have an Eastern Cooperative Oncology Group performance status ≤1, and have adequate liver and kidney function. Key exclusion criteria are the presence of MM brain lesions or meningeal involvement, and previous treatment with a BCMA-directed bispecific antibody (BCMA-directed antibody-drug conjugate is permitted) or chimeric antigen receptor T cell therapy. The primary endpoint is safety and tolerability of the REGN5458 combinations, in terms of dose-limiting toxicities (dose-finding portion only) and treatment-emergent adverse events for each cohort. Secondary endpoints are objective response rate, duration of response, progression-free survival, minimal residual disease status, pharmacokinetics, incidence of anti-drug antibodies, and overall survival. There is no formal statistical hypothesis for this study. The results will be reported in a descriptive manner. Exploratory analyses will evaluate measures of immune cell function in the bone marrow and peripheral blood that may be modulated by the combination regimens. Biomarkers of particular interest in the bone marrow include plasma and immune cell phenotypes, BCMA expression, and soluble BCMA. This study is actively recruiting patients at sites across the United States, Spain, France, and Greece. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal