B-cell Maturation Antigen Levels Research Articles

Soluble B-cell maturation antigen levels for disease monitoring in oligo- and non-secretory relapsed multiple myeloma

Soluble B-cell maturation antigen (sBCMA) is overexpressed on multiple myeloma (MM) cells. We investigated whether sBCMA levels correlated with other myeloma tumor volume indicators and its utility in monitoring oligo-secretory/non-secretory (O-S/Non-S) MM. In 115 patients with newly diagnosed MM, sBCMA was compared with M-protein levels, bone marrow plasma cells (BMPCs), circulating tumor cells (CTCs), and total diffusion volume (tDV; estimated by whole-body diffusion-weighted magnetic resonance imaging) at diagnosis. sBCMA levels increased significantly with International Staging System stage, chromosome 1q21 gain/amplification and CTC levels. sBCMA also correlated strongly with %BMPC (r = 0.65), moderately with tDV (r = 0.55) and paraprotein levels (involved immunoglobulin in IgG and IgA subtypes, r = 0.44 and 0.4; involved free light-chain levels in light-chain-only MM, r = 0.61, all P < 0.05). Longitudinal changes in sBCMA were consistent with disease status in both 17 O-S/Non-S and other secretory MM cases. Furthermore, sBCMA levels increased as early as 6 months pre-relapse in almost all O-S/Non-S relapsed patients. Thus, sBCMA correlates strongly with total tumor volume in MM, as assessed using different modalities. We suggest that sBCMA is useful, not only for monitoring responses in patients with O-S/Non-S MM but also for early relapse detection and prediction.

Deciphering the genetics and mechanisms of predisposition to multiple myeloma

Multiple myeloma (MM) is an incurable malignancy of plasma cells. Epidemiological studies indicate a substantial heritable component, but the underlying mechanisms remain unclear. Here, in a genome-wide association study totaling 10,906 cases and 366,221 controls, we identify 35 MM risk loci, 12 of which are novel. Through functional fine-mapping and Mendelian randomization, we uncover two causal mechanisms for inherited MM risk: longer telomeres; and elevated levels of B-cell maturation antigen (BCMA) and interleukin-5 receptor alpha (IL5RA) in plasma. The largest increase in BCMA and IL5RA levels is mediated by the risk variant rs34562254-A at TNFRSF13B. While individuals with loss-of-function variants in TNFRSF13B develop B-cell immunodeficiency, rs34562254-A exerts a gain-of-function effect, increasing MM risk through amplified B-cell responses. Our results represent an analysis of genetic MM predisposition, highlighting causal mechanisms contributing to MM development.

Expression of alternatively spliced BCMA RNA with skipping of transmembrane domain.

7514 Background: The TNFRSF17 gene, which encodes for the B cell maturation antigen (BCMA), is a small gene expressed mainly on the surface of plasma cells and some DLBCL cells. This gene is composed of three exons. Exon 1 is responsible for the extracellular domain, exon 2 for the transmembrane domain, and exon 3 for the TRAF binding domain. The BCMA is currently targeted by various types of immunotherapies as a major therapeutic approach for the treatment of multiple myeloma (MM). This includes CAR-T cells, bi-specific antibodies, and antibody-drug conjugates (ADC). However, emerging data indicates that alternative splicing in genes is an important mechanism for expression of different isoforms that may influence antibody-based therapies. We explored the potential of the presence of alternative splicing in BCMA transcripts via sequencing of BCMA RNA in patients with lymphoma or multiple myeloma. Methods: RNA was extracted from 587 fresh bone marrow samples with lymphoid/plasma cell neoplasms or from FFPE samples with lymphoma. In addition, cfRNA was extracted from 260 peripheral blood plasma samples from patients with lymphoma or multiple myeloma. RNA was sequenced using a hybrid capture-targeted RNA panel with analysis focused on TNFRSF17 (BCMA) gene transcript. Quantification of RNA transcript was done using Salmon algorithm. Results: Of the 587 lymphoma/plasma cell samples, 161 (27%) samples showed alternative splicing involving deletion of exon 2 (BCMA∆Ex2). Of the 260 cfRNA samples, 14 (6%) showed BCMA∆Ex2. The median percentage of BCMA∆Ex2 transcripts was 0.7% of total BCMA transcripts in cellular samples as compared with 9% in cfRNA samples. In cellular samples, there was a correlation between levels of BCMA and BCMA∆Ex2 (R=0.63). Cases with higher levels of BCMA had significantly higher levels of BCMA∆Ex2 (P&lt;0.0001, Kruskal-Wallis). In contrast, cfRNA showed no correlation between levels of BCMA and levels of BCMA∆Ex2 (R=021, Spearman) and mildly higher level of BCMA∆Ex2 in cases with higher levels of BCMA (P=0.002, Kruskal-Wallis). Conclusions: BCMA exon 2 skipping is detected in a significant number of patients with multiple myeloma and lymphoma. The percentage of skipping transcripts is low in cells and relatively higher in cfRNA. Since exon 2 skipping deletes the transmembrane domain, this BCMA protein may remain in the cytoplasm or perhaps is secreted as cell-free BCMA protein. The demonstration of relatively higher levels of BCMA∆Ex2 in cfRNA is likely reflecting higher turnover of cells and raises the possibility that cells with this isoform of BCMA are more aggressive than cells without the expression of this isoform. Further studies are needed to explore the clinical relevance of the expression of such abnormal BCMA protein on treatment with the various forms of anti-body-based therapy or CAR-T.

Association of patient (pt) factors and pharmacodynamic biomarkers with progression-free survival (PFS) after idecabtagene vicleucel (ide-cel) in pts from KarMMa-3.

7527 Background: Ide-cel, a B-cell maturation antigen (BCMA) CAR T cell therapy (tx), significantly improved PFS vs standard regimens in pts with triple-class–exposed relapsed or refractory MM (13.8 vs 4.4 mo, P&lt;0.001) in KarMMa-3. Median PFS in the ide-cel treated population (pop; n=225) was 15.7 (95% CI, 12.5–18.9) mo (Rodríguez-Otero ASH 2023). PFS benefit of ide-cel was consistent across high-risk pt subgroups. Previous analyses indicated that lower levels of baseline soluble BCMA (sBCMA; measure of tumor burden) and lower basal inflammatory markers were positively associated with response to ide-cel. This analysis identified pre- and post-tx pt factors and pharmacodynamic biomarkers associated with longer PFS with ide-cel in the treated pop. Methods: Median PFS (15.7 mo) of the treated KarMMa-3 pop was used to divide pts into 2 groups: longer PFS (&gt;15.7 mo; n=106) or shorter PFS (≤15.7 mo; n=110). Censored pts (n=9) who had a PFS &lt;15.7 mo were excluded. Pre- and post-tx values for laboratory parameters, MM assessments, sBCMA, inflammatory cytokines, ide-cel transgene persistence, and minimal residual disease (MRD; 10−5 sensitivity) were evaluated. Serum free light chains (sFLCs) were evaluated as biomarkers of plasma cell clearance; clearance was defined as level below limit of detection of both involved and heterotypic sFLC. Analyses were post hoc and exploratory; statistical tests were used to identify relationships of interest. Results: In total,94% of pts with longer PFS achieved a best overall response of ≥very good partial response and had lower levels of β-2 microglobulin ( P&lt;0.01), lactate dehydrogenase ( P&lt;0.01), and sBCMA ( P&lt;0.01) at baseline. Inflammatory markers (eg, tumor necrosis factor, ferritin, C-reactive protein) were significantly higher ( P&lt;0.05) in pts with shorter vs longer PFS at baseline. All pts with longer PFS (100%) achieved sFLC clearance after ide-cel infusion. Among pts who achieved initial clearance, pts with longer PFS had a longer duration of sFLC clearance than shorter PFS ( P&lt;0.001), suggesting more durable engraftment of ide-cel and tumor control. Within MRD-evaluable pts, those with longer PFS achieved higher MRD negativity vs shorter PFS (87% vs 41%) at 6 mo post infusion with sustained MRD negativity (53% vs 4%) lasting 8–12+ mo. No apparent relationship was observed between PFS and ide-cel persistence at 6-, 9- and 12-mo post-infusion. Conclusions: Pts with longer PFS after ide-cel infusion had relatively lower tumor burden and lower levels of peripheral inflammatory markers before tx, highlighting the importance of managing baseline tumor burden before CAR T cell infusion to achieve optimal response. Complete and sustained clearance of sFLCs within the first few mo post infusion was associated with longer PFS. MRD negativity at 6 mo and sustained MRD negativity were enriched in pts with longer PFS. Clinical trial information: NCT03651128 .

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

AbstractB-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 × BCMA bispecific antibody, 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 1 of whom also did not respond to ciltacabtagene autoleucel. Whole exome sequencing of tumor DNA from 1 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.

Increased Bone Turnover and Decreased BCMA Levels in Patients with Response to Anti-BCMA CAR T Cell Therapy in Relapsed/Refractory Multiple Myeloma

Introduction For most patients (pts), multiple myeloma (MM) remains an incurable disease. Osteolytic lesions (OL) are present in 80% of pts at diagnosis and skeletal-related events are the most common cause for symptomatic relapse. Hence, myeloma bone disease (MBD) is a major cause of morbidity and mortality especially among pts with relapsed/refractory MM (RRMM). B cell maturation antigen (BCMA) targeted chimeric antigen receptor (CAR) T cells have revolutionized treatment of patients with RRMM. While remission rates are high and pts experience long-standing treatment free intervals after CAR T cell treatment, little is known about its effects on MBD and bone turnover. The latter can be determined in plasma by measuring markers of bone formation and resorption. We aimed to analyze the longitudinal development of bone turnover during and after anti-BCMA CAR T cell treatment in RRMM pts. Methods We analyzed 28 consecutive pts with RRMM who underwent treatment with either Idecabtagene-Vicleucel (n=27) or Ciltacabtagen-Autoleucel (n=1) at our center between December 2021 and February 2023. Median age at infusion was 63 (range 39-75) years. 57% of pts were male. All pts were triple-class exposed, 21% had penta-class refractory disease. Status prior to CAR T cell infusion was complete remission (CR, 11%), partial remission (PR, 21%), very good PR (14%), stable disease (4%), or progressive disease (PD, 50%). The impact of CAR T cell treatment on bone turnover was assessed by collecting plasma samples on the day of T cell apheresis, as well as days (d) 30 and 100 after CAR T cell infusion. Samples were analyzed by ELISA to evaluate plasma levels of bone formation marker Osteocalcin (OCN), bone resorption markers Tartrate-resistant acid phosphatase 5b (TRAcP5b) and fibroblast growth factor 23 (FGF23) as well as BCMA as a marker for the targeted myeloma cells. Median levels were compared between subgroups by Wilcoxon tests. Remission status after CAR T cell therapy was first evaluated at d 30 according to International Myeloma Working Group consensus criteria. Median follow-up was 6 months. Results Of the 28 pts treated with anti-BCMA CAR T cells, 64% achieved at least a PR (responders), while 36% had PD shortly (30 d) after. Within 14 d after infusion, 82% developed a cytokine release syndrome (CRS, °I or II), in 30% of which Tocilizumab was applied. Only 1 pt developed immune effector cell-associated neurotoxicity syndrome. In responders, BCMA levels in plasma decreased significantly between apheresis and d 30 ( P&amp;lt;.01) and between apheresis and d 100 ( P&amp;lt;.01), while pts with PD showed no significant change in BCMA levels at the same timepoints ( P=.6 and P=.8). Furthermore, BCMA levels were significantly higher in pts with PD at d 30 ( P&amp;lt;.01) and d 100 ( P=.02), compared to responders (Figure 1). Regarding bone resorption markers, TRAcP5b levels were significantly increased in responders ( P=.01) but not pts with PD ( P=.56) on d 30 after apheresis. Furthermore, we determined a significant increase of intact FGF23 between apheresis and d 30 ( P&amp;lt;.01) and a subsequent significant decrease of intact FGF23 ( P=.02) between d 30 and d 100 in responders. Since FGF23 and interleukin-6 (IL-6) are physiologically dependent, the observed FGF23 dynamics are likely to be an effect of increasing IL-6 in the context of CRS. Aptly, pts who received Tocilizumab because of full-blown CRS had higher intact FGF23 levels at d 30 ( P=.02) compared to pts without Tocilizumab treatment. The bone formation marker OCN showed a significant increase in responders between apheresis and d 100 ( P=.01), as well as between d30 and d100 ( P=.02), while there were no significant changes in OCN levels over the time in pts with PD (Figure 2). Conclusion Pts responding to treatment with anti-BCMA CAR T cells, showed an increased bone turnover shortly after infusion. The increase of bone resorption marker levels appeared to be a short-term trend and could possibly be related to inflammation and high interleukin levels during the first days after infusion. However, we also saw a steady increase of bone formation marker OCN, which remained significant till d 100 in responders. Our data implies, that response to anti-BCMA CAR T cells is accompanied by increasing bone formation. Further analyses are needed to evaluate, whether this treatment can have positive long-term effects on MBD.

Rationale for and Design of Papilio-1: A Phase 1/2, Multicenter, Open-Label Study to Evaluate the Feasibility, Safety, and Efficacy of Point-of-Care-Manufactured Anti-B-Cell Maturation Antigen Chimeric Antigen Receptor T Cells (GLPG5301) in Relapsed/Refractory Multiple Myeloma

Background and Significance Despite improvements in treatment, patients with multiple myeloma (MM) ultimately relapse or become refractory to available regimens. Triple-refractory (refractory to CD38 monoclonal antibodies [mAbs], 1 proteasome inhibitor [PI] and 1 immunomodulatory drug [IMiD]) or penta-refractory (refractory to CD38 mAbs, 2 PIs and 2 IMiDs) patients have a poor prognosis and are in urgent need of novel treatment options. Chimeric antigen receptor (CAR) T-cell therapies have been used to target B-cell maturation antigen (BCMA) and have shown good clinical activity in patients with relapsed/refractory MM (RRMM). Faster manufacturing procedures may improve clinical outcomes by lowering discontinuation rates and reducing the need for bridging therapy. GLPG5301, a BCMA-directed CAR T-cell therapy, is produced using a novel decentralized and automated point-of-care manufacturing model. This allows administration of a fresh product within 7 days of apheresis. The objective of Papilio-1 is to evaluate the feasibility, safety and efficacy of GLPG5301, manufactured at the point-of-care, in adult patients with RRMM. Study Design and Methods Papilio-1 (CTIS: 2022-500782-27-00) is a Phase 1/2, multicenter, open-label study of GLPG5301 in patients with RRMM after ≥2 prior lines of therapy. Key eligibility criteria are presented ( Table). The study will be conducted in centers across Europe. The Phase 1 primary objective is to evaluate the safety of GLPG5301 and determine the recommended Phase 2 dose (RP2D). The Phase 1 primary endpoint is incidence of serious adverse events, including dose-limiting toxicities (DLTs), until 14 days post-infusion. The Phase 2 primary objective is to evaluate GLPG5301 efficacy. The Phase 2 primary endpoint is best objective response until 2 years post-infusion. Secondary endpoints include: type, frequency and severity of adverse events; duration of response; progression-free survival (PFS); overall survival (OS); minimal residual disease; soluble BCMA levels; time to response; time to progression; GLPG5301 levels in blood and bone marrow; serum levels of chemokines and cytokines; and number of successfully manufactured GLPG5301 products. Exploratory assessments include additional GLPG5301 pharmacokinetics (PK) and pharmacodynamics (PD). Patients will undergo leukapheresis of mononuclear cells for point-of-care manufacturing of GLPG5301 after screening ( Figure). Prior to infusion of GLPG5301, patients will receive fludarabine/cyclophosphamide lymphodepleting chemotherapy, followed by a resting period of at least 2 days. GLPG5301 will be administered as a fresh product, followed by in-hospital monitoring for 7 days. Safety, efficacy, PK and PD will be assessed until end of treatment (100 days post-infusion), and monthly until study completion (2 years). Patients will be enrolled in a long-term follow-up for up to 15 years. In Phase 1, dose escalation will be based on the occurrence of DLTs occurring within 14 days of infusion, according to predefined criteria. Three dose levels (DLs) will be assessed (DL1, DL2 and DL3). Six patients will receive the lowest dose level (DL1) and be evaluated for DLTs and efficacy. The appropriate dose for the following cohort will be then determined by the Safety Review Committee. This will be repeated until RP2D is determined. Approximately 30 patients will be treated with the RP2D in Phase 2. Safety and efficacy data will be analyzed using descriptive statistics. Response status will be summarized as best response (number, percentage, 95% confidence interval) and graphically displayed using swimmer plots by dose. Other binary endpoints will be described using a similar method. PFS and OS will be assessed using the Kaplan‒Meier method and calculated from the median survival time. Summary In Papilio-1, the feasibility, safety and efficacy of GLPG5301, a point-of-care-manufactured BCMA-directed CAR T-cell therapy, will be evaluated in adult patients with RRMM.

CD19/BCMA Dual-Targeting Fastcar-T GC012F for Patients with Relapsed/Refractory B-Cell Non-Hodgkin's Lymphoma: An Update

Background CD19-directed CAR-T cell therapy is a valuable treatment option for relapsed/refractory B-cell non-Hodgkin's lymphoma (r/r B-NHL). It has been shown that a significant percentage of B-NHL clinical samples also express B cell maturation antigen (BCMA). To further improve safety and efficacy, we have been exploring utility of a CD19 and BCMA dual-targeting CAR-T, GC012F, manufactured through a novel next-day FasTCAR-T process, for treatment of r/r B-NHL. Here, we present updated results of this ongoing IIT trial with more patients and longer follow-up. Methods From October 2021 to June 2023, nine eligible refractory/ relapsed defuse large B cell lymphoma (r/r DLBCL) patients (pts) were enrolled in this study and treated with a single infusion of GC012F in escalating dosing cohorts after a standard lymphodepletion regimen over 3 days (20 - 30 mg/m 2/day Flu, 250 - 300 mg/m 2/day Cy). The primary objectives of this study were safety and tolerability assessment; the secondary objectives were pharmacokinetics and efficacy. Cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS) were graded by ASBMT 2019, and adverse effects were evaluated according to CTCAE 5.0. Efficacy assessment of GC012F was based on the Lugano criteria. Expansion of CAR-T cells and CAR copy numbers were analyzed by flow cytometry and qPCR, respectively. Results As of data cut-off of June 30 th, 2023, with a median of 372 days (range 131 - 625) of follow-up, all nine patients received single GC012F infusions at dose levels from 3.7×10 4 to 3×10 5 CAR-T/kg, and completed 3 months or longer time follow-up evaluations. The median age was 52 years (range 18 - 60). Among the enrolled patients, eight pts were Ann Arbor stage III/IV disease, one patient with double hit, and four pts had IPI score of 3. The lymphoma samples of all patient expressed CD19, and 7 out of 8 expressed various levels of BCMA. The median SPD was 2690.81 mm 2 (408.3 - 13325.96 mm 2). Patients received a median of 2 prior lines (range 2 - 3) of therapy including rituximab and anthracyclines. Two patients received prior auto-HSCT. All patients responded to GC012F treatments, 100% (9/9) pts achieved ORR at month 3 post GC012F infusion; 77.8% (7/9) CR at month 3, 66.7% (6/9) CR at month 6 and 62.5% (5/8) CR at month 9, respectively. Up to date, the longest duration of remission was 20.8 months. No dose-limiting toxicities were observed. Five patients experienced grade 1 CRS, and one patient in the highest dose group (3×10 5 CAR-T/kg) had grade 3 CRS within 2 days. No ICANS were observed. Of the nine treated patients, grade ≥ 3 hematologic toxicities were neutropenia (9/9), leukopenia (8/9), lymphocytopenia (8/9), and thrombocytopenia (7/9). All TEAEs were resolved after treatment with standard of care and supportive care. The median peak copy number of CAR-T cells in the peripheral blood was 71,000 copies/μg DNA (range 9263 - 185,039), and the median peak time was 14 days (range 9 - 21d). CAR-T geometric mean AUC 0-28 for patients was 658,538 copies/μg DNA × day (319,104 - 1,359,031; 95% CI). CAR-T cells were also detected in tumor biopsies from all five patients tested. Conclusion This first-in-human trial of GC012F, a CD19-BCMA dual targeting CAR-T product, for the treatment of r/r B-NHL showed a manageable safety profile and promising clinical responses. The ORR was 100% at month 3 with 77.8% (7/9) achieving CR. GC012F CAR-T cells were detected in the tumor biopsies, indicating the infiltration of CAR-T cells into the tumor lesions. Overall, based on the safety and efficacy results, CD19-BCMA dual targeting CAR-T product GC012F is a promising therapy for patients with refractory/ relapsed DLBCL.

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.

High-Throughput Plasma Proteomics to Define the Precursor Multiple Myeloma Proteome and Identify Candidate High-Risk Disease Biomarkers of Progression

Introduction Multiple Myeloma (MM) develops from well-defined precursors Monoclonal Gammopathy of Undetermined Significance (MGUS) and Smoldering Multiple Myeloma (SMM), where patients remain stable or may unknowingly rapidly progress. Bone marrow (BM) biopsies are not routine for precursor disease management, and precursor patients are limited to monitoring few proteins within peripheral blood (PB) for signs of progressive disease. Deep proteome profiling of PB circulating proteins may help track disease; however, the dynamic range of the plasma proteome has limited the depth of detection for MS-based proteomics without first depleting abundant proteins and fractionating the samples after digestion to peptides. Technological advancements in multiplex immunoassays with low cross-reactivity and off-target events have enabled plasma profiling for disease stage classification, defining high-risk disease features, and novel therapeutic target discovery. Here, we perform the first comprehensive plasma proteomic profiling study on patients across the MM disease continuum and longitudinal sequential samples from progressive and stable disease. Methods We carried out high-throughput plasma proteomic profiling for approximately 3000 proteins simultaneously using the Olink® Explore 3072 library and Proximity Extension Assay (PEA) technology. Targeted proteins are recognized by multiplexed, matched pairs of antibodies labelled with unique DNA oligonucleotides that upon binding come into proximity, hybridize, and are extended to generate a unique sequence for protein identification with NextGen DNA sequencing. We profiled 423 PB plasma samples from 348 individuals, including MGUS (n=67), SMM (n=179), MM (n=44), and healthy controls (n=58). Sequential samples from patients with progressive disease (n=27) and patients with stable disease with matched clinical follow-up time were also profiled. Precursor defined samples from progressors ranged 1.03-5.88 yrs prior to diagnosis and were untreated in the precursor setting, while MM disease samples were also collected prior to any active disease therapy. Patients had a median clinical follow-up time of 7.05 years. T-tests, ANOVAs, and a linear mixed effect (LME) model were used to identify proteins that change across disease stages, progression status, and time. Results were adjusted for multiple testing using the Benjamini-Hochberg Method. Results We identified 751 significantly dysregulated proteins with the majority upregulated in progressive disease. We captured circulating levels of proteins highly expressed on the surface of plasma cells, including CD38, SDC1, BCMA and SLAMF7, highlighting the utility of PEA technology to monitor clinically-relevant targets for which monoclonal antibodies, antibody-drug conjugates, CAR-T and BiTE therapies are being developed. We identified proteins that significantly distinguished MGUS, SMM, and NDMM from healthy donors (n=222, 423 and 494), where increasing B-cell maturation antigen (BCMA) was a significant classifier and positive control. Consistent with previous findings, baseline BCMA levels were also significantly elevated in SMM-MM progressors vs. SMM non-progressors, further supporting the potential utility of BCMA measurements during routine blood tests of precursor MM patients. Proinflammatory cytokines were also identified, including IL1, IL5, IL6, IL16, and IL18, known to create a BM environment that promotes malignant cell development by suppressing the microenvironment, promoting cellular adhesion, or increasing angiogenesis. Four novel proteins that are vital for calcium homeostasis and integrin-mediated cell adhesion significantly increased from healthy to MM and were also significantly elevated in SMM progressors vs. SMM non-progressors, nominating these proteins as candidate biomarkers of high-risk disease. Conclusion We performed the most comprehensive plasma proteomics study to date, which characterized disease stage proteomes and identified candidate high-risk disease biomarkers in longitudinal progressor samples. Further advancements are underway to validate the accuracy levels of the novel candidates, test the performance of a classification model that recognizes disease stage-specific proteins, and determine how best to integrate proteins into current MM risk stratification models.

Early Safety Results of P-BCMA-ALLO1, a Fully Allogeneic Chimeric Antigen Receptor T-Cell (CAR-T), in Patients with Relapsed / Refractory Multiple Myeloma (RRMM)

P-BCMA-ALLO1 is an allogeneic CAR-T that targets B-cell Maturation Antigen (BCMA) and is currently being investigated for the treatment of RRMM. P-BCMA-ALLO1 cells are manufactured from healthy donor T-cells using non-viral transposon-based integration (piggyBac® DNA Delivery System) that introduces a human anti-BCMA V H-based CAR and an iCas9 safety switch. The piggyBac® DNA delivery system produces a highly enriched T stem cell memory product. The Cas-CLOVER™ Site-Specific Gene Editing System eliminates endogenous T cell receptor (TCR) expression via knockout of the TCR beta chain 1 gene to prevent graft-vs-host disease (GvHD), and the beta-2 microglobulin gene to reduce Major Histocompatibility Complex (MHC) class I expression to eliminate host-vs-graft responses. P-BCMA-ALLO1 demonstrated compelling activity in MM xenografts, providing rationale for this first-in-human phase I study. The primary objective is to assess the safety and maximum tolerated dose (MTD) of P-BCMA-ALLO1 based on dose limiting toxicity (DLT) in RRMM patients with measurable disease who have received a proteasome inhibitor (PI), immunomodulatory agent (IMiD) and anti-CD38 monoclonal antibody therapy. Key secondary objective will assess the anti-myeloma effect of P-BCMA-ALLO1. Exploratory objectives will assess CAR-T related cytokines, serum BCMA levels and cellular kinetics. The trial utilizes a standard 3 + 3 dose escalation design to test seven planned dose levels of intravenous P-BCMA-ALLO1 ranging from 0.0625 × 10 6 to 15 × 10 6 cells/kg (NCT04960579). Multiple lymphodepletion (LD) strategies and cyclic administration of P-BCMA-ALLO1 are being investigated. Repeat P-BCMA-ALLO1 administration is allowed for patients who do not achieve at least a partial response with the first dose after 4 weeks of follow up. We are currently dosing patients in arm S (Cyclophosphamide (Cy) 300 mg/m 2 + Fludarabine (Flu) 30 mg/m 2 X 3 days), arm P1 (Cy 500 mg/m 2 + Flu 30 mg/m 2 X 3 days), arm P2 (Cy 1,000 mg/m 2 + Flu 30 mg/m 2 X 3 days) and arm C (multiple P-BCMA-ALLO1 doses following Cy 300 mg/m 2 + Flu 30 mg/m 2 X 3 days). The median patient age was 73 (33, 85) years, 61% were female, and median time since MM diagnosis was 6.27 (1.48,18.95) years. Eight (35%) patients had high risk disease by cytogenetic criteria. The enrolled patients were heavily pretreated, having received median 7 (2,16) prior lines of therapy, with 30% having undergone prior BCMA targeting therapy. As of 10 Jul 2023, 24 patients received P-BCMA-ALLO1 at 4 dose levels. Twenty-two were treated in arm S, 1 each in arms P1 and P2. One patient received 0.25 X 10 6 cells/kg (cohort -1, arm S), 7 received 0.75 X 10 6 cells/kg (cohort 1, arm S), 12 received 2 X 10 6 cells/kg (cohort 2, 10 in arm S, 1 in arm P1 and 1 in arm P2), and 4 received 6 X 10 6 cells/kg (cohort 3). One of the cohort 2 patients received a second P-BCMA-ALLO1 infusion at the cohort 2 dose level following disease progression. The median time from enrollment to start of LD chemotherapy was 2 (1-8) days and from enrollment to CAR-T infusion was 7 (6-13) days. No patient needed bridging therapy between enrollment and the start of LD. Twenty-two (all in arm S) of the 24 treated patients completed DLT evaluation by data cutoff. None of the patients treated thus far had DLTs. Most common treatment emergent adverse events (TEAEs) were anemia (36%), neutropenia (36%), constipation (36%) and leukopenia (32%). Most common ≥ grade (G) 3 TEAEs were neutropenia (36%), leukopenia (32%) and anemia (23%). Most of the AEs were considered unrelated to P-BCMA-ALLO1. Three (14%) patients developed grade 1 Cytokine Release Syndrome (CRS). One (4%) patient experienced grade 1 Immune Effector Cell Associated Neurotoxicity Syndrome (ICANS), which resolved with one dose of corticosteroids. None of the patients experienced GvHD by the last follow up. P-BCMA-ALLO1 is a non-viral transposon-generated allogeneic “off-the-shelf” CAR-T that is rapidly available for administration and well tolerated in a heavily pretreated RRMM patient population with minimal CRS/ICANS risk. Updated results including safety, preliminary efficacy, and selected biomarkers will be presented at the American Society of Hematology 2023 meeting.

Clinical Outcomes and Serum B-Cell Maturation Antigen Levels in a Real-World Unselected Population of Newly Diagnosed Multiple Myeloma Patients.

Progression-free survival (PFS) and overall survival (OS) of newly diagnosed multiple myeloma (MM) patients have been widely published in the clinical trials setting, but data published from real-world settings are limited. We determined the survival and factors that predict outcomes among 161 unselected, newly diagnosed MM patients whose frontline therapy was started at a single clinic specializing in the treatment of this B-cell malignancy. None of these patients underwent an autologous stem cell transplantation as part of their initial therapy and the population had a high proportion (35%) of cytogenetic high-risk patients. With a median follow-up of 42.7 months, the cohort had a median PFS of 22.8 months and a median OS of 136.2 months. The 1-, 3-, and 5-year survival rates were 97.5%, 85.3%, and 76.2%, respectively. These results are considerably better than those reported from patients enrolled in clinical trials and those from countries with national registries. Age <65years predicted for a longer OS (p=0.0004). Baseline serum B-cell maturation antigen (sBCMA) levels were also assessed and showed median and mean levels of 320.3 ng/mL and 551.1 ng/mL, respectively. Furthermore, patients with baseline sBCMA levels in the lowest quartile (≤136.2 ng/mL) showed a longer PFS (p=0.0262). These results provide clinicians with a real-world understanding of the survival of unselected, newly diagnosed patients initiating therapy in a clinic specializing in the care of MM patients.

Population pharmacokinetic-pharmacodynamic model of nirogacestat effects on B-cell maturation antigen in healthy subjects.

8027 Background: B-cell maturation antigen (BCMA) is expressed on the membrane of normal plasma cells (PCs) and multiple myeloma (MM) cells and is the target of several investigational agents and approved products for MM treatment. Mechanistically, BCMA is cleaved from the cell surface by the enzyme gamma secretase, resulting in reduced levels of membrane-bound BCMA (mbBCMA) and release of soluble BCMA. Gamma secretase inhibitors (GSIs) have been shown to increase BCMA receptor density and potentiate the activity of several BCMA-targeted therapies in both clinical trials and in vitro studies. Although the effect of GSIs on BCMA receptor density has been well characterized in vitro, their effect on BCMA dynamics is incompletely understood in humans. This study sought to evaluate the pharmacodynamics (PD) of the GSI nirogacestat on mbBCMA in healthy subjects. Methods: A total of 23 healthy subjects were administered single doses of 50, 150, or 300 mg nirogacestat or multiple doses of 100 mg every 12 hours. Density of mbBCMA on PCs isolated from whole blood and bone marrow was measured by flow cytometry at baseline and 1, 2, 4, 8, 24, and 48 hours after single doses and 24 and 48 hours after the first dose in subjects receiving multiple doses. Nirogacestat serum concentrations were also measured at multiple time points to fully characterize the pharmacokinetics (PK). A population PK-PD model of the effect of nirogacestat on BCMA dynamics in humans was developed and used to simulate various administration dose levels and schedules to find an optimal nirogacestat regimen that continually increases mbBCMA over the dosing interval. Results: A 2-compartment model with linear absorption and elimination described serum nirogacestat in healthy subjects. A sequential PK-PD model that included an effect compartment and drug effect on mbBCMA cleavage rate described mbBCMA values. Simulations indicated that a 100-mg dose administered twice daily sufficiently maintained elevated and clinically relevant mbBCMA levels. Exposure and efficacy benchmarks from in vitro experiments and published clinical studies were used to contextualize simulations and hypothesize various dose and schedule scenarios to minimize dose of the BCMA-targeting therapy while maximizing BCMA potentiation. Conclusions: These results suggest that nirogacestat dose and schedule may have direct implications on the potentiation of BCMA-targeting therapies. Simulations suggest that the 100-mg, twice-daily nirogacestat dose results in elevated mbBCMA levels that could even allow for use of lower doses of BCMA-targeted agents while maintaining or improving efficacy. Study results along with this PK-PD model will be used to support nirogacestat dose and schedule in ongoing and planned studies in patients with MM.

Effects of teclistamab and talquetamab on soluble BCMA levels inpatients with relapsed/refractory multiple myeloma.

B-cell maturation antigen (BCMA) is selectively expressed as a surface protein on plasma cells and is a validated target for multiple myeloma (MM). A soluble form of BCMA (sBCMA) is elevated in the sera of patients with MM. The bispecific antibodies teclistamab (BCMA×CD3) and talquetamab (G protein-coupled receptor family C group 5 member D [GPRC5D]×CD3) are in clinical development as therapies for MM. Using data from patients with relapsed/refractory MM (N=300) who participated in phase 1 studies of teclistamab (NCT03145181) or talquetamab (NCT03399799), baseline serum sBCMA levels were quantitatively analyzed relative to response to treatment, percentage of bone marrow plasma cells (BMPCs), and cytogenetic risk. By cycle 3 day 1 of treatment, sBCMA levels declined from baseline in 50 (88%) of 57 responders and 49 (98%) of 50 responders to teclistamab and talquetamab, respectively. Depth of response correlated with the magnitude of sBCMA reduction. In contrast, sBCMA increased in 33 (80%) of 41 patients and 24 (49%) of 49 patients who did not respond to teclistamab or talquetamab, respectively. Baseline sBCMA levels correlated with the percentage of BMPC; patients with extramedullary plasmacytomas who had low levels of BMPC (≤10%) tended to have high baseline sBCMA levels (≥400 ng/mL), based on limited data. Baseline sBCMA levels and changes in sBCMA levels at cycle 3 day 1 were similar in patients with high- and standard-risk cytogenetics treated with teclistamab or talquetamab. These results support the use of sBCMA as a potential surrogate marker of tumor burden and treatment response in MM.

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.

Both APRIL and antibody-fragment-based CAR T cells for myeloma induce BCMA downmodulation by trogocytosis and internalization

BackgroundChimeric antigen receptor (CAR) T cell therapy targeting B cell maturation antigen (BCMA) on multiple myeloma (MM) produces fast but not long-lasting responses. Reasons for treatment failure are poorly understood....

Prognostic value of B-cell maturation antigen, CD56 expression and neutrophil lymphocyte ratio in patients with multiple myeloma

Introduction: B-cell maturation antigen (BCMA) plays a critical role in regulating B-cell proliferation and differentiation into plasma cells. CD56 is involved in the adhesion of myeloma cells to the bone marrow matrix. Thus, lack of CD56 expression is associated with a higher incidence of extramedullary disease. An elevated neutrophil-lymphocyte ratio (NLR) has been recognized as a poor prognostic factor in various hematological malignancies. The aim of this study was to evaluate the role of BCMA, CD56 and NLR as novel prognostic markers in multiple myeloma (MM). Material and methods: The study included 80 subjects, 40 MM patients, and 40 normal healthy age- and sex-matched controls. BCMA was analyzed by enzyme-linked immune sorbent assay. Flow cytometry was used for the determination of CD56 expression. NLR was calculated from the complete blood count differential count. All patients received six cycles of bortezomib, cyclophosphamide and dexamethasone (VCD). Treatment outcome was assessed and progression-free survival (PFS) was estimated using Kaplan-Meier survival analysis. Results: Patients who achieved complete remission showed lower BCMA levels, positive CD56 expression, and lower NLR. Moreover, higher BCMA levels and CD56 negative expression were significantly associated with shorter PFS. Conclusions: Our study emphasizes the importance of BCMA, CD56 and NLR to predict the clinical outcome in MM patients. This could help in better risk stratification and tailored clinical management of MM patients.

Serum B-cell maturation antigen level in multiple myeloma

Background Multiple myeloma (MM) is a malignant neoplasm of plasma cells that accumulate in the bone marrow producing excessive amounts of monoclonal immunoglobulins (Ig) and suppression of other normal polyclonal Igs, leading to bone destruction and marrow failure. B-cell maturation antigen (BCMA) is known to bind to B-cell-activating factor (BAFF) or a proliferation-inducing ligand (APRIL), leading to the activation of cell proliferation pathways and upregulation of antiapoptotic proteins, which is considered the main step in the pathogenesis of MM. Aim This study was planned to evaluate serum BCMA levels in MM patients and its clinical significance. Patients and methods This study was carried out on 40 newly diagnosed MM patients and 40 apparently healthy individuals served as the control. The two studied groups were compared as regards serum BCMA levels using the enzyme-linked immunosorbent assay technique. Results Results showed that there was a statistically significant increase in MM patients when compared with the control group, and there was a significant positive correlation between BCMA level and plasma cells in bone marrow, M protein, number of bone lesions, and beta-2 microglobulin. However there was no correlation between BCMA level and age, creatinine, and hemoglobin. Conclusion Serum BCMA level was significantly elevated in MM patients compared with healthy controls that may suggest the role of BCMA in the pathogenesis of MM. The high sensitivity of BCMA suggests that it may be a novel diagnostic marker, and its level was positively correlated with other prognostic markers. It may be used as a prognostic marker in MM patients.

Abstract 6251: Evaluation of a dual-targeting BCMA-CS1 HLE BiTE® molecule for multiple myeloma

Abstract Multiple myeloma (MM) is a cancer of the antibody-producing plasma cells (PC). MM is invariably fatal due to frequent disease relapse and/or treatment refractoriness, and therapies that provide deeper and more durable responses are needed. BiTE® (Bispecific T Cell Engager) molecules are immunotherapy agents that redirect a patient’s T cells to lyse tumor cells by simultaneously engaging a tumor associated antigen (TAA) on cancer cells and CD3ε on T cells. Clinical activity has been observed in MM using BiTE® molecules and other T cell engagers that target B-cell maturation antigen (BCMA). BCMA is an attractive MM TAA due to its broad prevalence and restricted normal tissue profile, mainly PCs and plasmablasts. However, heterogenous expression in MM cancer cells, potential antigen loss, and the presence of high levels of soluble BCMA in MM patient sera present challenges that may prevent BCMA-only-targeted therapies from achieving their full potential. To address these issues, we generated a BiTE® molecule capable of engaging BCMA and a second MM TAA, CS1 (also known as SLAMF7). This BiTE® molecule also contains an Fc-based domain to provide half-life extension (HLE). Here, we evaluated the in vitro and in vivo properties of this BCMA-CS1 HLE BiTE® molecule. The BCMA-CS1 HLE BiTE® molecule had nanomolar binding affinity for human BCMA, CS1, and CD3ε and the non-human primate (NHP) orthologues. In vitro, we observed picomolar activity against human MM cell lines with a range of BCMA and CS1 expression in a T cell dependent cellular cytotoxicity (TDCC) assay using human T cells or NHP peripheral blood mononuclear cells. The BCMA-CS1 HLE BiTE® molecule retained TDCC activity in the presence of soluble BCMA up to 2500 ng/mL as well as against human MM cells engineered to express only BCMA or CS1. In vivo, this BiTE® molecule inhibited tumor growth in a MM xenograft model. We also evaluated the BCMA-CS1 HLE BiTE® molecule in NHP over 15 days (IV dosing; intra-animal dose escalation from 60→240 μg/kg or 180→540 μg/kg on days 1 and 8). We observed hallmarks of BiTE® molecule activity in all groups, including transient decreases in circulating lymphocytes and moderate increases in cytokines like MCP-1. We measured the PC-specific transcripts BCMA and J-chain in NHP bone marrow and blood as surrogates for PC levels using ddPCR. These transcripts were reduced in both treatment groups (≥90%) with the strongest effects occurring in the 180→540 μg/kg group. Lastly, we showed that CS1 is highly and broadly expressed in MM patient samples and is restricted to a few normal hematopoietic cell types including PCs, NK cells, T cells, and some monocytes. These data suggest that the BCMA-CS1 HLE BiTE® molecule has potent in vitro and in vivo activity and may provide therapeutic benefit for MM patients by expanding the population of MM cancer cells that can be eliminated by a BiTE® molecule while overcoming common mechanisms that can impair BCMA-only-targeted MM therapies. Citation Format: Elizabeth T. Andrews, Stephanie C. Casey, Mohammad Farhad Amani, Grit Lorenczewski, Mozhgan Farshbaf, Lisa Winkel, Matthias Klinger, John M. Harrold, Famke Aeffner, Ana Goyos, Matthias Friedrich, Tara Arvedson, Matthew G. Chun. Evaluation of a dual-targeting BCMA-CS1 HLE BiTE® molecule for multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6251.

Abstract 6020: Preclinical activity of LM-305 targeting G-protein-coupled receptor class 5 member D (GPRC5D) antibody drug conjugate for the treatment of multiple myeloma

Abstract Emerging therapies including chimeric antigen receptor (CAR) T therapy targeting B cell maturation antigen (BCMA) have shown promising clinical outcomes in relapsed and refractory multiple myeloma (RRMM). However, majority of patients eventually develop progressive disease due to heterogeneity in BCMA expression or loss of antigen during treatment. Therefore, new targets are required to overcome the problem of resistance encountered with these agents. The expression of GPRC5D (G-protein-coupled receptor class 5 member D) is high on MM cells and low in normal tissues. Although high levels of GPRC5D and BCMA were found in similar proportions of MM patients, they are independently expressed by MM cells. GPRC5DxCD3 bispecific T-cell-redirecting antibody is currently under clinical investigation. Preliminary results have shown early evidence in efficacy with tolerable safety profile. These findings identified GPRC5D as an ideal therapeutic antibody drug conjugate (ADC) target for the treatment of patients with RRMM. Therefore, the present study was aimed at developing a novel anti-GPRC5D ADC drug LM-305 and evaluating its preliminary efficacy by performing preclinical activity using in vitro and in vivo mouse models. The binding affinity of LM-305 was evaluated by flow cytometry in engineered GPRC5D over-expressing MM tumor cells and endogenous GPRC5D-expressing MM tumor cells. The internalization of LM-305 was assessed with pH-dependent dye in NCI-H929 and MM.1R cell lines. The cytotoxicity of LM-305 was assessed by cell viability assay in MM tumor cell lines. In vivo anti-tumor activity of multiple doses of LM-305 (1mg/kg, 3mg/kg, 10mg/kg) was evaluated in NCI-H929 and MM.1R tumor cell line derived xenograft (CDX) models. In vitro studies revealed that LM-305 binds with high affinity to GPRC5D over-expressing cell lines and GPRC5D endogenously expressing MM cells in a dose-dependent manner. It can be efficiently internalized by GPRC5D expressing cells and further lysosomal lysis was detected by pHrodo dye. LM-305 displayed potent cytotoxicity when co-cultured with MM tumor cells (NCI-H929 and MM.1R) with IC50 values ranging from 0.1 to 0.3 nM. In vivo tumor xenograft models suggested that treatment with LM-305 resulted in dose-dependent inhibition of tumor growth in tumor bearing mice. Additionally, LM-305 exhibited complete response (CR) in the GPRC5D high-expressing MM CDX models at a dose of 3mg/kg. Moreover, LM-305 showed good safety profile in the animal studies. In conclusion, this preclinical data suggested that LM-305 is a novel GPRC5D targeting ADC with best-in-class potential, and therefore it can be a promising therapeutic candidate for the treatment of RRMM patients expressing GPRC5D. Citation Format: Wentao Huang, Jie Luo, Yifan Li, Da Fei, Xia Qin, Runsheng Li. Preclinical activity of LM-305 targeting G-protein-coupled receptor class 5 member D (GPRC5D) antibody drug conjugate for the treatment of multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6020.