Introduction
Multiple myeloma (MM) is the second most prevalent form of blood cancer in adults, accounting for 10% of all hematologic diseases, and the incidence is expected to increase in an aging population. 1
The approval of 2 B-cell maturation antigen (BCMA)–targeting chimeric antigen receptor (CAR) therapies in patients with relapsed and refractory MM (RRMM) is considered to be a paradigm shift in the therapeutic approach for MM. 2 , 3 Furthermore, recent data supporting the use of CAR T cells in earlier lines of therapy underline the transformative potential of T-cell-engaging therapies. 4 T-cell bispecific antibodies (TCBs) have demonstrated significant potential in patients with MM, based on high objective response rate (ORR), deep and long-lasting responses, favorable safety profiles, and off-the-shelf availability. 5 , 6 Three TCBs have recently been approved for RRMM. 7 , 8 , 9 , 10 , 11 Teclistamab, elranatamab, and talquetamab exhibit a 1+1 immunoglobulin G configuration with monovalent binding to the tumor target, BCMA or G protein–coupled receptor class C group 5 member D (GPRC5D), and to CD3e on human T cells. 12 , 13 , 14 We hypothesized that bivalent targeting of a myeloma cell target would translate into an avidity-based gain in tumor cell lysis, as observed with glofitamab and alnuctamab. 15 , 16
GPRC5D represents the most differentiated target in MM with minimal normal tissue expression in the skin and little to no expression on healthy B and plasma cells. 17 , 18 , 19 , 20 Emerging RNA data sets confirm GPRC5D to be highly expressed across different stages of MM and to be elevated even further in patients with t(4;14) translocations and 1q amplifications. 18 , 21 , 22 , 23 The therapeutic potential of targeting GPRC5D was illustrated by strong clinical responses observed for talquetamab and GPRC5D CAR T cells, suggesting that TCB molecules with higher potency could achieve even deeper and more durable responses. 24 , 25
In the current study, we introduce forimtamig, a novel and more potent GPRC5D-TCB with 2+1 format. We delineated the mode of action of forimtamig in an orthotopic humanized mouse model and provided evidence for improved response in combination with standard-of-care (SoC) agents. Furthermore, we identified novel combination partners in BCMA TCB and cereblon E3 ligase modulatory drugs (CELMoDs) that allow to further increase progression-free survival (PFS) rates by preventing occurrence of tumor relapse.
Materials and methods
Protein target prevalence was assessed in baseline bone marrow (BM) aspirate samples from 64 evaluable patients with RRMM enrolled in phase 1 study NCT04557150 . GPRC5D and BCMA expressions were measured by flow cytometry and are shown as frequency of positive MM plasma cells (MMPCs) or molecules of equivalent soluble fluorophore. Chromosomal aberrations for determination of cytogenetic risk were detected by fluorescence in situ hybridization on immunomagnetically sorted plasma cells before patients entered the study. All patients provided a written informed consent.
All animal studies reported have been reviewed and approved by the local government under the animal licenses ROB-55.2-2532.Vet_03-16-10 or ROB-55.2-2532.Vet_03-20-170. Patients with cancer from Institut Universitaire du Cancer de Toulouse-Oncopole (Toulouse) gave a written informed consent, and collection was approved by the French Committee for the Protection of Persons (DC-2012-1654) and by local Institut Universitaire du Cancer de Toulouse-Oncopole review boards. Patients with cancer from Clínica Universidad de Navarra were collected according to the local ethics committee and the Declaration of Helsinki. All BM samples were collected after an informed consent was given by each patient, according to the local ethics committee and the Declaration of Helsinki.
A complete description of methods is presented in the supplemental Data , available on the Blood website.
Results
Forimtamig binds with high avidity to the N terminus of human GPRC5D inducing formation of stable synapses
Forimtamig has a 2+1 head-to-tail configuration designed to exhibit bivalent binding to GPRC5D on myeloma cells and monovalent binding to CD3ε on human T cells bearing a silent Fc with P329G LALA mutations to extend half-life and prevent Fc gamma receptor (FcγR) engagement ( Figure 1 A). 26 Forimtamig was confirmed to selectively bind to human GPRC5D but not GPRC5A, the closest homologue, 27 and exhibited no significant binding to murine GPRC5D nor cynomolgus monkey GPRC5D ( Figure 1 B). Forimtamig was found to bind to the unstructured N-terminal part of GPRC5D, furthest away from the GPRC5D dimerization interface. It is likely that a single GPRC5D homodimer can accept 2 forimtamig anti-GPRC5D Fabs, leading to a 2:2 complex such as scFv150-18, whereas binding of talquetamab anti-GPRC5D near the dimerization interface results in a 2:1 GPRC5D-Fab complex ( Figure 1 C). This is consistent with the observation that, although monovalent affinities of forimtamig to GPRC5D (11 nM) and to CD3ε (1.9 nM) were in the same range as talquetamab, 28 bivalent binding of forimtamig resulted in a more than 50-fold higher avidity of 0.2 nM ( Figure 1 D; supplemental Figure 1 A). We next compared the stability of synapses induced by forimtamig and 1+1 GPRC5D-TCB in vitro using live confocal imaging of labeled CD8 T cells and NCI-H929 or KMS-12BM tumor cells with high and low GPRC5D expression levels, respectively ( supplemental Figure 1 B). Immunological synapses formed by forimtamig were more stable, as indicated by stronger reduction in T-cell motility, combined with an increase in contact duration between T and tumor cells irrespective of target expression level ( Figure 1 E; supplemental Figure 1 C-D). We next used fluorescently labeled forimtamig to confirm its localization at the tumor cell–T-cell interface and observed synapse formation to correlate with increased caspase 3/7 expression in cancer cells ( supplemental Figure 1 E). Figure 1. Biochemical characterization of forimtamig and synapse formation. (A) The molecular model of forimtamig was generated and visualized with discovery studio 2021; residues involved in the P329G LALA silent Fc mutations are shown as spheres colored in cyan. (B) Binding kinetics was measured using horseradish peroxidase (HRP)-based colorimetric analysis in Chinese hamster ovary (CHO) cells transfected with mouse, cynomolgus, or human GPRC5D or human GPRC5A and incubated with increasing concentrations of forimtamig. (C) The likely epitope of forimtamig could be mapped to the unstructured N terminus of GPRC5D and is shown in the context of 2 recently published cryo-electro magnetic (EM) structures of GPRC5D homodimer, in complex with 2 scFvs (protein data bank (PDB) ID 8yzk) and in complex with a single talquetamab Fab anti-GPRC5D (PDB ID 9ima ); the core epitope (yellow ball and stick representation) consists of GPRC5D residues 5 to 10, but an additional involvement of residues 11 to 16 (gray stick representation) cannot be ruled out; as the unstructured N terminus is not or only partially resolved in the published structures, the missing N-terminal segments (purple ribbon representation) were remodeled from the AlphaFold model and the shown conformations represent only a placeholder for a whole ensemble of possible conformations; the actual conformation of GPRC5D’s N-terminal segment in complex with forimtamig anti-GPRC5D has yet to be determined. (D) Grating-coupled interferometry (GCI) was used to determine affinities (equilibrium dissociation constant, K D ) of forimtamig and forimtamig monovalent Fab to human GPRC5D and of forimtamig to human CD3ε; black lines indicate fit according to a 1:1 Langmuir model. (E) Correlation of contact duration and speed of fluorescently labeled T cells as measured by confocal live cell imaging to determine the stability of immunological synapses; all treatments are at 200 ng/mL. scFv, single-chain variable fragment.
Forimtamig exhibits high potency against tumor cells with clinically relevant target expression level
To identify preclinical models with patient relevant target expression, we compared GPRC5D protein level in BM aspirates of 64 patients with RRMM enrolled in phase 1 study NCT04557150 with a set of MM cell lines using flow cytometry ( Figure 2 A; supplemental Figure 2 B). GPRC5D protein expression was detected in 97% of patients and MMPCs from target positive patients expressed on average 29 600 GPRC5D molecules ( Figure 2 A; supplemental Figure 2 A). GPRC5D level did not significantly differ between patient groups defined by cytogenetic risk factors, except for patients with t(11;14) translocation that exhibits less than 10 000 GPRC5D copy numbers ( Figure 2 B). Furthermore, GPRC5D protein level was not affected by the Internal Staging System score, number of previous lines of therapy, previous exposure to BCMA- or CD38-targeted therapies, or extramedullary disease (EMD) status ( supplemental Figure 2 C-D). Of note, most MMPCs expressed GPRC5D or BCMA or both markers, whereas double-negative cells were rare ( supplemental Figure 2 A). We next tested the in vitro activity of different TCBs against 4 MM cell lines covering a wide range of target expression as observed in patients with MM, using peripheral blood mononuclear cells (PBMCs) as effector cells. Compared with a 1+1 GPRC5D-TCB, forimtamig was 202-, 54-, 87-, or 156-fold more potent in killing of MMPCs with high (OPM-2), intermediate (NCI-H929), low (KMS-12BM), or ultra-low (U266B1) GPRC5D expression. Forimtamig exhibited higher killing potency than a 2+1 BCMA TCB against NCI-H929 and achieved comparable EC50 with KMS-12BM and U266B1, despite more than 4-, 20-, or even 50-fold higher BCMA than GPRC5D expression, respectively ( Figure 2 C-D; supplemental Figure 2 B). As with tumor cell killing, forimtamig was highly potent in activating CD8 + and CD4 + T cells, demonstrated by upregulation of CD69 and CD25 and enhanced secretion of granzyme-B (GZB), interleukin-2 (IL-2), and interferon gamma (IFN-γ) ( Figure 2 E-G; supplemental Figure 2 E-H). Figure 2. GPRC5D protein expression in patients with RRMM and evaluation of TCB potency in vitro. (A) GPRC5D receptor density on MMPCs in bone marrow aspirates from 61 patients with RRMM enrolled in study NCT04557150 (3 samples from 64 could not be reported due to technical issues); these values are depicted on the y-axis of the plot and each dot represents 1 sample; dashed lines on the graph represent mean values for GPRC5D receptor density for 7 cell lines with names of the cell lines specified respectively; GPRC5D receptor density was determined by bead quantification using MESF beads for patient samples and Quantum Simply Cellular kit for cell lines (technical details are described in supplemental Materials and methods ). (B) Association between overall cytogenetic risk or individual chromosomal aberrations and GPRC5D binding sites on MM cells in BM; high risk was defined as having ≥1 of the following chromosomal aberrations (regardless of 1q21gain/ampl): del(17p), t(4;14), t(14;16); the 1q21gain/ampl group was defined as having only 1q21gain/ampl reported; N indicates the number of samples that could be evaluated for the target expression. (C-G) MM cell lines with different GPRC5D and BCMA receptor densities (RDs) were cocultured with human peripheral blood mononuclear cells (PBMCs) from 4 to 5 individual healthy donors at an effector-to-target (E:T) ratio of 5:1 and treated with increasing concentrations of forimtamig, 2+1 BCMA TCB, 1+1 GPRC5D TCB, or a untargeted TCB control; LDH release was measured as a surrogate for tumor cell lysis after 40 hours; Triton X-100–treated samples were used to determined maximum tumor lysis and untreated samples to determine spontaneous tumor lysis; expression levels of CD69 and CD25 were measured by flow cytometry as a surrogate for CD8 T-cell activation after 40 hours; EC50 values were calculated from dose-response curves from individual donors; in case curve fits did not allow valid EC50 calculations (if curves either did not reach the top or bottom plateau or had a very wide confidence interval), data were not plotted; and statistical differences between treatment groups were determined per cell line using 1-way analysis of variance (ANOVA) with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005. EC50, median effective concentration; LDH, lactate dehydrogenase; MESF, molecules of equivalent soluble fluorophore; ns, not significant.
Forimtamig exhibits superior activity in BM aspirates and in vivo models of MM compared with 1+1 GPRC5D TCB
We next validated our results in fresh tumor material from patients with newly diagnosed MM (NDMM). We first exposed CD138 − BM mononuclear cells from at least 6 patients with NDMM to a different set of myeloma cell lines with a wide range of GPRC5D expression. Compared with other TCBs, forimtamig exhibited superior potency in activating tumor-infiltrating lymphocytes (TILs) as indicated by increased CD25 expression. Activation of TILs was independent of GPRC5D expression level and was associated with enhanced secretion of cytokines ( Figure 3 A-B). We then treated total BM aspirates from at least 6 patients with NDMM per cell line accordingly. Forimtamig was most potent in MMPC killing and TIL activation as confirmed by depletion of up to 60% MMPCs and increased expression of CD69 and CD25 ( Figure 3 C-E; supplemental Figure 3 A-C). To validate our findings in vivo, we treated NCI-H929 engrafted humanized mice with 4 weekly TCB doses at 0.1, 1, and 10 mg/kg ( supplemental Figure 3 D). In contrast to what we observed in vitro, NCI-H929 tumor exhibited patchy and low GPRC5D expression in mice ( supplemental Figure 3 F-G). Forimtamig induced tumor regression in all animals and at all doses tested whereas the maximum response observed for 1+1 GPRC5D TCB was tumor growth delay ( Figure 3 F; supplemental Figure 3 E). The 2+1 BCMA TCB was less efficacious especially at lower doses of 1 mg/kg and 0.1 mg/kg although tumors expressed a higher level of BCMA treatment start ( Figure 3 F; supplemental Figure 3 F-G). Figure 3. Benchmarking of forimtamig against other TCBs in preclinical models of MM. (A-B) Fresh bone marrow aspirates from patients with NDMM were depleted of CD138 + plasma cells and cocultured with RPMI-8226 (6 samples enrolled), MOLP-8 (9 samples enrolled), or AMO-1 (12 samples enrolled) tumor cells with a wide range of GPRC5D and BCMA RDs at a final E:T ratio of 5:1 in the presence of increasing concentrations of forimtamig, 1+1 GPRC5D TCB, 2+1 BCMA TCB, or an untargeted TCB control antibody at 0.007 nM; expression of CD25 on CD8 TILs was measured by flow cytometry after 48 hours; baseline CD25 expression, as detected by an untargeted TCB control, is represented by the horizontal black dotted line; cytokine secretion was analyzed in the supernatant of T-cell–myeloma cell line cocultures after 36 hours; graphs summarizing the mean + standard error of the mean (SEM) expression and statistics between groups were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005; ∗∗∗∗ P < .0001. (C-D) Total BM cells from 9 patients with NDMM were cultured in the presence of increasing concentrations of forimtamig, 1+1 GPRC5D TCB, 2+1 BCMA TCB, or an untargeted TCB control at 0.007 nM; expression of CD25 on CD4 and CD8 TILs was measured by flow cytometry after 36 hours; baseline CD25 expression, as detected by an untargeted TCB control, is represented by the horizontal black dotted line; and graphs summarizing the mean + SEM expression and statistics between groups were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005. (E) Total BM cells from 13 patients with NDMM were cultured in the presence of the indicated concentration of TCBs for 36 hours; graphs summarizing the mean + SEM depletion of MMPCs as calculated by the frequency of CD38 + CD138 + double-positive cells in relation to untreated TCB control; graphs summarizing the mean + SEM expression and statistics between groups were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005. (F) Humanized NSG mice (huNSG) were subcutaneously implanted with 2.5 × 10 6 NCI-H929 tumor cells and when tumors reached 200 to 250 mm 3 injected once a week IV with indicated TCB doses; tumor growth was measured over time using caliper and was plotted as individual spider plots for each of the 9 animals per group; animals treated with 2+1 BCMA TCB at 10 mg/kg were taken down after 2 cycles as they reached predefined termination criteria (see “Materials and methods”). TNFα, tumor necrosis factor α.
Forimtamig induces killing of MM cells in the BM microenvironment
We next generated an orthotopic MM model and tracked growth of luciferase-labeled NCI-H929 cells in the femur using bioluminescence imaging or soluble BCMA (sBCMA), a correlative biomarker for tumor load in mice ( supplemental Figure 4 A). Tumor load in BM was reduced at all forimtamig doses tested, with most animals being tumor-free before the third administration ( Figure 4 A; supplemental Figure 4 C). A longitudinal biomarker analysis ( supplemental Figure 4 B) confirmed rapid killing of MMPCs in the BM as early as 72 hours after cycle 1 (C1) dosing as indicated by a drop in sBCMA levels by almost 100% ( Figure 4 B). Forimtamig induced a 10-fold reduction of circulating CD8 T cells 48 hours after C1 injection and an almost fivefold CD8 T-cell expansion 24 hours after C2 dosing ( Figure 4 C). A transient decline of CD8 T cells observed in the tumor tissue at C1 was followed by an up to fivefold expansion of intratumoral T cells 72 hours after C1 and C2 dosing ( Figure 4 C). Timing of T-cell expansion in tumors and drop of sBCMA correlated with a shift of T-cell phenotype from naïve CD62L + CD45RA + toward CD4RA − CD62L − effector memory CD8 + T cells in the blood and in the tumor ( Figure 4 D). Forimtamig induced systemic cytokine release at C1 with peak concentrations at 4 hours (IL-2, macrophage inflammatory protein-1 alpha, and granulocyte-macrophage colony-stimulating factor), 24 hours (tumor necrosis factor α [TNFα], CXCL-10, granulocyte colony-stimulating factor, and IL-10), and 48 to 72 hours (IFN-γ, IL-6, IL-8, or sCD25). In the absence of detectable tumor mass, no cytokine release was observed when forimtamig was dosed at C2 ( Figure 4 E). We next split dosing in C1 and explored the impact of step-up dosing (SUD) on cytokine release. We treated mice with a SUD regimen of 0.005, 0.05, and 0.5 mg/kg at C1 day 1 (C1D1), day 8 (C1D8), and day 15 (C1D15) and compared cytokines 10 minutes, 4 hours, and 72 hours after each administration to flat dosing at 0.5 mg/kg ( Figure 4 F). SUD decreased the peak serum level of CXCL-10, IL-8, and IFN-γ by 2- up to 16-fold and delayed the cytokine peak from C1D1 to C1D8 ( Figure 4 F). To evaluate the impact of SUD on efficacy, we compared the SUD and flat dosing regimen for a fixed duration in vivo. The use of SUD induced a comparable efficacy compared with flat dosing ( Figure 4 G; supplemental Figure 4 C). Figure 4. Evaluation of forimtamig’s mode of action in an orthotopic mouse model of MM. (A) Luciferase-labeled NCI-H929 cells were injected into the femur of humanized NSG (huNSG) mice and randomized into 4 different treatment arms based on bioluminescent signals measured as photons per second (p/s); animals were treated once weekly with indicated doses of forimtamig and tumor growth was monitored over time; background signals were determined in nontumor-bearing mice; each group included 10 animals and is presented as mean + SEM. (B) Orthotopically engrafted huNSG mice (n = 5) were treated once weekly with 0.1 mg/kg forimtamig and sBCMA levels were detected in serum of animals before, 4, 24, 48, 72, and 168 hours after dosing using enzyme-linked immunosorbent assay (ELISA). (C-E) Orthotopically engrafted humanized mice (n = 40) were treated once weekly with 0.1 mg/kg forimtamig; BM, blood and serum were harvested from n = 5 animals at 4, 24, 48, 72, and 168 hours, after C1 and C2 dosing, and subjected to fluorescence-activated cell sorting (FACS) or cytokine analysis; CD8 T-cell counts in the tumor tissue (right femur) were normalized against CD8 counts from healthy BM from the same animal (left femur); naïve CD8 T cells were defined as CD62L + CD45RA + and effector memory T cells as CD4RA − CD62L − . (F) Orthotopically engrafted huNSG mice (n = 15 animals per group) were treated once weekly with 0.5 mg/kg forimtamig flat or 0.005, 0.05, and 0.5 mg/kg SUD at C1D1, C1D8, and C1D15; serum was harvested from n = 5 animals at 10 minutes, 4 hours, and 72 hours after dosing and cytokine levels were measured by bioplex; and fold change in cytokine release was calculated by normalization to vehicle animals at the same time points. (G) Orthotopically engrafted huNSG mice (n = 15 animals per group) were treated once weekly with 0.5 mg/kg forimtamig flat or 0.005, 0.05, and 0.5 mg/kg SUD at C1D1, C1D8, and C1D15, and tumor monitored over time by bioluminescence imaging; and background signals were determined in nontumor-bearing mice. TNF-α, tumor necrosis factor α.
Combination of forimtamig with SoC agents improves antimyeloma responses
We next evaluated the combination of forimtamig with SoC agents using autologous BM aspirate samples from treatment-naïve patients with NDMM. Forimtamig induced lysis of 34% MMPCs, which was significantly increased in combination with daratumumab (dara) or with dara plus pomalidomide (pom) but not pom alone ( Figure 5 A). Forimtamig induced upregulation of activation markers on CD8 and CD4 TILs that was not significantly changed in combinations ( Figure 5 B; supplemental Figure 5 A). Forimtamig enhanced activation of NK cells as measured by CD25 and CD107a expression, which was increased in triple combination with dara and pom ( supplemental Figure 5 C). We next combined forimtamig with high-dose carfilzomib (carfi) and observed comparable tumor cell killing or T-cell activation ( Figure 5 C-D; supplemental Figure 5 B). To explore depth and durability of response of SoC combinations in vivo, we increased the number of treatment cycles from 4 to 6 followed by a treatment-free interval of 2 weeks ( supplemental Figure 5 D). Although forimtamig induced regressions at C3, tumor relapse occurred in 50% to 70% of mice ( Figure 5 E-G; supplemental Figure 5 G-I). Although SoC monotherapies did not achieve significant tumor growth inhibition ( supplemental Figure 5 E), we next tested their potential to improve response to forimtamig. Combination with pom resulted in a 20% enhanced response at C1D4, an increased median PFS (mPFS) and 20% higher probability of PFS at termination ( Figure 5 E; supplemental Figure 5 F-G). Although dara did not improve early response to forimtamig or mPFS, first relapse occurred only 10 days after therapy was stopped ( Figure 5 F; supplemental Figure 5 F,H). In combination with carfi we observed 30% enhanced efficacy at C1D4, an increased mPFS and a 30% high PFS rate at termination ( Figure 5 G; supplemental Figure 5 F,I). We next measured cytokines in serum of mice 48 hours after treatment initiation. Although pom combination increased serum levels of IFN-γ, IL-2, and TNF-α compared with forimtamig alone, dara induced only a moderate increase of TNF-α and there was a slight decrease in systemic cytokine release for the carfi combination ( Figure 5 H-J). Figure 5. Combination of forimtamig with SoC agents. (A-D) Total BM cells from 4 (carfi) or 11 (dara and pom) patients with NDMM were cultured in the presence of 1 nM forimtamig, 10 nM daratumumab, 1 μM pomalidomide (48 hours before incubation), 3 nM carfilzomib, or combination of forimtamig with either of the SoC agents for 48 hours, and tumor cell lysis as well as expression of CD69, CD25, CD137, and PD-1 by CD8 + lymphocytes was measured by flow cytometry; tumor cell lysis was calculated by the percentage reduction of CD38 + CD138 + double-positive cells in relation to untreated TCB control; and graphs summarizing the mean ± SEM expression and statistical differences against forimtamig monotherapy were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005; ∗∗∗∗ P < .0001. (E-G)Humanized NSG (huNSG) mice were subcutaneously implanted with NCI-H929 tumor cells and when tumors reached 200 mm 3 injected once weekly with 0.1 mg/kg forimtamig or combination of forimtamig with 8 mg/kg dara (once weekly, IV), 10 mg/kg pomalidomide (every day, by mouth [po]), or 3 mg/kg carfi (twice weekly, IV). Tumor volume was measured over time using a caliper; treatment was stopped after 6 cycles and mice were monitored for at least 2 additional weeks to check for tumor relapse; each group includes 10 animals and is presented as mean + SEM. (H-J) Serum was harvested from n = 5 animals 48 hours after first forimtamig and 24 hours after first SoC dosing and cytokine levels were measured by bioplex analysis; graphs summarizing the mean ± SEM expression and statistical differences between multiple groups were determined by1e-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005; ∗∗∗∗ P < .0001. hu, human; TNF-α, tumor necrosis factor α.
Forimtamig in combination with a BCMA-directed TCB overcomes tumor escape
To identify more efficient strategies for relapse prevention, a better understanding of underlying mechanisms is essential. Given that target expression is a determinant of TCB response, 16 we quantified GPRC5D in NCI-H929 tumors at baseline and after relapse to forimtamig by immunohistochemistry. All relapsed tumors stained negative for GPRC5D whereas expression of BCMA was not affected ( Figure 6 A-B). To explore whether simultaneous targeting of BCMA and GPRC5D could improve duration of response, we combined forimtamig with a 1+1 BCMA TCB using concomitant, alternating, or consecutive treatment schedules ( supplemental Figure 6 A). Forimtamig induced deep responses in all mice at C4 and tumor escape observed in 47% of animals ( Figure 6 C; supplemental Figure 6 B). Notably, 1+1 BCMA TCB treatment induced tumor growth delay during C1-2 but tumors rapidly progressed and mPFS was only slightly increased compared with the control arm. Concomitant injection of forimtamig and 1+1 BCMA TCB at C1, C2, or C3 induced strong and sustained tumor regression resulting with all animals being progression free at study termination. Similar results were obtained when both TCBs were administered in an alternating schedule or when therapy was switched from forimtamig to 1+1 BCMA TCB at C4. Switching from 1+1 BCMA TCB to forimtamig at C3 did not significantly increase mPFS but enhanced PFS rate by 40% ( Figure 6 C; supplemental Figure 6 C). Interestingly, tumor load of BCMA TCB-exposed animals at C3 correlated with durability of response to forimtamig ( supplemental Figure 6 D). Combination of forimtamig and 1+1 BCMA TCB did not significantly exacerbate cytokine release compared with respective monotherapies ( Figure 6 D). We observed increased levels of CXCL-10 when 1+1 BCMA TCB was added to forimtamig at C2 and C3 or when the treatment was switched at C2 or C4 ( Figure 6 D). Concomitant injection at C1 resulted in IFN-γ levels equivalent to 1+1 BCMA TCB but much lower than forimtamig ( Figure 6 D). We observed signs of tolerance as indicated by reduced cytokine release upon first dosing in TCB pre-exposed animals ( Figure 6 D). We observed clear trends toward increased GZB expression by CD8 T cells when combination was started at C2 and C3 and higher levels of TIGIT and PD-1 across different time points when a second TCB was added ( Figure 6 E). Forimtamig induced a shift from naïve to effector memory T cells in the blood and we observed the same kinetics in combination with BCMA TCB ( supplemental Figure 6 E). The expansion of circulating T cells induced by forimtamig at C2, C3 and C4 was hampered by addition of 1+1 BCMA TCB especially at C1 but less pronounced when TCBs were alternated. There was a persistence of GZB-positive circulating CD8 T cells during C1 to C2 upon concomitant injection at C2 and C3 ( Figure 6 E). Figure 6. Combination of forimtamig with BCMA TCB. (A-B) BCMA and GPRC5D expression was analyzed in ∼1000 mm 3 NCI-H929 tumors that relapsed upon forimtamig treatment at 0.1 mg/kg in experiment 5F by immunohistochemistry and quantified using visiopharm (n = 5 animals per group); control tumors were matched in tumor size but were removed from the study at earlier time points; data are shown as mean ± SEM expression and statistical differences between multiple groups were determined using Student t tests. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005. (C) huNSG mice were subcutaneously implanted with NCI-H929 tumor cells and when tumors reached 200 to 250 mm 3 injected once weekly with forimtamig at 0.1 mg/kg or 1+1 BCMA TCB at 1 mg/kg or the combination of both TCBs using a concomitant schedule starting at C1, C2, or C3; an alternating schedule; or different switch schedules in which 1 TCB was dosed after stopping the second TCB; treatment was stopped after 8 injections and tumor escape was monitored for 10 additional days; tumors growth was monitored twice weekly using caliper measurements. (D) Serum was harvested from n = 5 animals 48 hours after C1, C2, C3, and C4 dosing and cytokine levels were measured by bioplex analysis; and graphs summarizing the mean ± SEM expression and statistical differences between multiple groups were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005. (E) Blood was collected 48 hours after C1, C2, C3, and C4 TCB dosing and human CD3 + T cells were quantified and characterized using spectral flow cytometry; and graphs summarizing the mean ± SEM expression and statistical differences between multiple groups were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005; ∗∗∗∗ P < .0001. TNF-α, tumor necrosis factor α.
Novel combinations maximize efficacy and allow lowering forimtamig target dose
Antigen escape reported in TCB-exposed patients with MM potentially limits the durability of TCB-TCB combinations. 7 , 16 , 29 We next tested whether enhanced antitumor and immunostimulatory activity of CELMoDs could represent an antigen-independent and novel combination strategy. 30 , 31 We first tested forimtamig in combination with iberdomide in a fixed duration in vivo experiment starting to combine at C1. All mice treated with the combination of forimtamig and iberdomide were considered progression free at the end of the observation period whereas monotherapies failed to control tumor growth over time ( Figure 7 A; supplemental Figure 7 B-D). Increased efficacy was correlated with a significant increase in cytokine production 48 hours after forimtamig dosing ( Figure 7 B). Later administration of iberdomide (C3) resulted in shorter mPFS and 40% lower PFS at termination but was associated with decreased levels of cytokines in response to combination ( Figure 7 C-D; supplemental Figure 7 C-D). Figure 7. Combination of forimtamig with CELMoDs. (A,C) Humanized NSG (huNSG) mice were subcutaneously implanted with NCI-H929 tumor cells and when tumors reached 200 mm 3 injected once weekly IV with forimtamig at 0.1 mg/kg or combination with oral dosing of iberdomide at 10 mg/kg 5 times weekly (5q7d) starting at C1D2 or C3D2; tumor volume of individual mice was measured over time using a caliper; treatment was stopped after 7 forimtamig injections and mice were monitored for at least 2 additional weeks to check for tumor relapse; each group included 10 animals. (B,D) Serum was harvested from n = 5 animals 48 hours after C1 and C3 forimtamig dosing and 24 hours after iberdomide dosing and cytokine levels were measured by bioplex analysis; and graphs summarizing the mean ± SEM expression and statistical differences between multiple groups were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005. (E) huNSG mice were subcutaneously implanted with NCI-H929 tumor cells and, when tumors reached 200 mm 3 , injected once weekly with step-up doses of forimtamig at 0.0005, 0.002, and 0.04 mg/kg or combination with oral administration of 1 mg/kg or 3 mg/kg mezigdomide once weekly (1q7d), thrice weekly (3q7d), or 5 times weekly (5q7d). Tumor volume was measured over time using a caliper; treatment was stopped after 7 forimtamig injections and mice were monitored for at least 2 additional weeks to check for tumor relapse; each group included 10 animals. (F) Serum was harvested from n = 5 animals 48 hours after C1D1, C1D8, and C1D15 forimtamig dosing and 24 hours after mezigdomide dosing and cytokine levels were measured by bioplex analysis; graphs summarizing the mean ± SEM expression and statistical differences between multiple groups were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005. (G) NCI-H929 tumors were isolated from 3 animals of groups A, B, and F ( supplemental Figure 7 A) 48 hours after 0.002 mg/kg SUD and intratumoral T-cell number and phenotype was analyzed by flow cytometry; graphs summarizing the mean ± SEM expression and statistical differences between multiple groups were determined by 1-way ANOVA with Tukey posttest analysis. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005. (H-I) Total BM cells from 6 patients with NDMM were cultured in the presence of 0.1 nM forimtamig, 0.1 to 10 nM mezigdomide, or the combination for 96 hours and tumor cell lysis and expression of CD69, CD25, HLADR, CD107a, LAG-3, and PD-1 by CD8 lymphocytes was measured by flow cytometry; untreated and untargeted-TCB treated samples served as reference controls; graphs summarizing the mean ± SEM expression and statistical differences between multiple groups were determined by 2-way ANOVA with Tukey posttest analysis; and statistical analysis for activation markers was referenced against forimtamig monotherapy. ∗ P < .05; ∗∗ P < .005; ∗∗∗ P < .0005; ∗∗∗∗ P < .0001. MIG, monokine induced by gamma-interferon; MIP-1a, macrophage inflammatory protein-1 alpha; TNF-α, tumor necrosis factor α.
To maximize efficacy while limiting cytokine release, we introduced subcutaneous SUD at 0.0005, 0.002, and 0.04 mg/kg for forimtamig and combined with mezigdomide. Dose levels were calculated to match the clinically achieved relevant exposure of forimtamig in phase 1 (average plasma concentration (C avg ), data not shown). Mezigdomide dosing was started at C1D2 at 1 or 3 mg/kg using a once weekly, thrice weekly, or 5 times weekly schedule and a treatment-free interval after C2 ( supplemental Figure 7 A). Tumor growth delay was the best overall response observed for mezigdomide (3 mg/kg, 5 times weekly) ( supplemental Figure 7 B). Forimtamig induced transient tumor regressions after C1D1 and C1D15 with 80% of mice showed progressive disease at the end of C1. In contrast, potent tumor growth inhibition with increased mPFS and 90% (5 times weekly) and 60% (thrice weekly) progression-free animals at termination were observed when forimtamig was combined with 1 mg/kg mezigdomide. A single administration of mezigdomide at 1 mg/kg increased mPFS by only 9 days and failed to significantly improve response ( Figure 7 E; supplemental Figure 7 F-G). Combination with high-dose mezigdomide led to rapid onset of tumor regression during C1 correlating with 80% or even 100% PFS rate when mezigdomide was administered thrice weekly or 5 times weekly, respectively. Even when mezigdomide was administered once per week, mPFS was increased by 37 days and terminal PFS by 20% ( Figure 7 E; supplemental Figure 7 E,G).
We next measured systemic cytokine release 48 hours after each forimtamig SUD and detected a maximal 2.5-fold increase in combination with 1 and 3 mg/kg mezigdomide compared with forimtamig. Release of IL-2, CXCL-10, and monokine induced by gamma-interferon (MIG) induced by target dose administration of forimtamig was boosted most significantly by once weekly dosing of mezigdomide. Mezigdomide combination induced significantly increased levels of TNF-α at C1D1 and C1D15 irrespective of the dose and schedule. Serum levels of monocyte-associated cytokines macrophage inflammatory protein-1 alpha and IL-8 tend to decrease with higher and more frequent dosing of mezigdomide ( Figure 7 F). In a next step, we analyzed circulating immune cells in the blood of mice before C4 administration of forimtamig (48-120 hours after last mezigdomide dosing). We observed correlations of more frequent dosing of mezigdomide with reduced number of circulating B cells and increased frequency of PD-1, LAG-3, and TIGIT-positive CD4 and CD8 T cells ( supplemental Figure 7 H). To improve mechanistic understanding of synergistic antitumor effects, we analyzed the number and phenotype of human T cells in the tumor tissue of 3 mice treated with forimtamig or the combination with 1 mg/kg mezigdomide (5 times weekly schedule) at C1D10. We detected a significantly higher number of human T cells and a higher proportion of GZB-positive CD8 and CD4 T cells in tumors of mice treated with the combination of both drugs ( Figure 7 G).
We validated our findings in fresh BM aspirates from patients with NDMM using a fixed dose of forimtamig and a dose range of mezigdomide. Despite using 10-fold lower concentration than SoC combinations, forimtamig as a single agent induced lysis of 40% tumor cells after 96 hours. We also observed dose-dependent efficacy of mezigdomide achieving maximum tumor cell lysis of ∼25% at 10 nM. Efficacy was further increased for the combination with mezigdomide at all dose levels tested, achieving maximum tumor lysis of ∼60% at 1 nM ( Figure 7 H). T-cell activation induced by forimtamig was further increased by the combination with mezigdomide as indicated by an increase of HLA-DR-, CD25-, and CD107a-positive cells and an increase of PD-1-, TIGIT-, and LAG-3-positive TILs. Although there was no significant impact of mezigdomide monotherapy on TIL activation, we observed a dose-dependent decrease of TIGIT on both CD4 and CD8 T-cell subsets and an increase of LAG-3 on both T-cell subsets ( Figure 7 I; supplemental Figure 7 I).
Discussion
Although recent advances have been made in the treatment of MM with the introduction of proteasome inhibitors (PIs), immunomodulatory drugs, and CD38-targeting compounds, prognosis for patients with extensive treatment history, high-risk cytogenetics, and EMD is poor, and therapies benefiting a broader patient population are urgently needed. 32 , 33 , 34 , 35
Forimtamig exhibits higher potency against MMPCs even in the presence of low GPRC5D expression and clears MM cells in the BM after a single injection suggesting the potential to achieve deeper responses than approved TCBs early at treatment and at lower target doses. 36 , 37 , 38 , 39 , 40 , 41 In an ongoing phase 1 trial, forimtamig achieves ORR and complete response rates similar to talquetamab but at threefold to 180-fold lower target dose with 30% of patients achieving early minimal residual disease (MRD) negativity of 10 −5 at C2D1 and independent of target expression level. 13 , 42 Further dose optimization will potentially allow to maintain efficacy while reducing the risk of skin and mucosal toxicities associated with GPRC5D targeted TCBs in MM. Our data support targeting GPRC5D in MM given that protein expression was high across patient subsets with high medical need and that we observed superior killing when targeting GPRC5D compared with BCMA using the same 2+1 antibody format.
We observed a clear correlation between tumor load and pharmacodynamic effects induced by forimtamig supporting extended treatment intervals and suggesting 72 hours to represent an ideal time point for biomarker assessment in the blood and the tumor tissue. Almost 60% of patients with MM treated with TCBs experience cytokine release syndrome (CRS) especially during early treatment cycles. 6 We provide important evidence that introduction of SUD provides an efficient CRS mitigation strategy even for a highly potent molecule such as forimtamig. 6 , 43 Our data underline the potential of forimtamig to provide benefit to patients with MM as a monotherapy and underpin the value of using stem cell engrafted humanized NSG (huNSG) mouse models to test clinical lead molecules for safety and efficacy assessment. 15 , 44
Moreover, forimtamig was shown to represent a high-potential combination partner for SoC agents in MM. Synergistic antitumoral and natural killer stimulatory effects suggest a benefit of dara combinations in patients with high tumor burden and a high risk of developing CRS. Initial results from phase 1/2 TRIMM-2 and MajesTEC-2 trials confirm a favorable safety and efficacy profile of dara in combination with talquetamab or teclistamab. 45 , 46 The broad immunomodulatory effects of pom could help sustain T-cell responses in patients that passed the critical CRS window. In this context, promising efficacy and manageable safety have been reported from the MonumenTAL-2 trial combining talquetamab with pom. 47 PIs have been considered suboptimal combination partners for T-cell therapies given that detrimental effects on T-cell responses have been reported for 1+1 GPRC5D-TCBs exhibiting lower potency. 21 Interestingly, carfi achieved a favorable efficacy and safety profile compared with anti-CD38 and immunomodulatory drugs. Considering the potential of PIs to induce immunogenic cells enhancing major histocompatibility complex class I–dependent T-cell responses, we identified PIs as promising combination partners for highly potent TCBs such as forimtamig. 48 , 49 Although our results provide only limited mechanistic insights, we provide preclinical evidence that combining with SoC agents will potentially allow forimtamig to move to earlier lines of therapy to induce deeper responses early on and increase the time in remission.
Target modulation promotes relapse to CAR T-cell and TCB monotherapies underscoring the importance of optimized combination strategies. 10 , 11 , 25 Biomarker data from phase 1 study NCT04557150 support targeting more than 1 MM surface marker to promote rapid and deeper tumor clearance. 50 Here we provide preclinical evidence that a combination of forimtamig with a 1+1 BCMA TCB can prevent tumor relapse and induce long-lasting responses although we acknowledge the need for even longer follow-ups. Additional experiments with a longer follow-up are needed. Lower ORR rates have been reported in patient subsets exposed to TCBs, 51 and strong antitumoral responses we observed upon consecutive dosing of TCBs support the findings of T-cell fitness correlating with response to TCBs in MM. 49 Our TCB combination data collectively suggest that coadministration of 2 TCBs provides no benefit over alternating or consecutive dosing as long as the higher potent molecule is used at early cycles. Cytokine release data suggest that combination of 2 TCBs does not induce a higher risk of CRS, which potentially allows combining 2 potent TCB molecules in consecutive order. These preclinical findings are supported by initial results from BCMA TCB and GPRC5D-TCB combination trials reporting high ORR rates even in patients with EMD but also highlight the need for optimal balance between efficacy and safety. 34 Although BCMA TCB combination data look encouraging, the observation of tumor relapse driven by BCMA target modulation and increased risk of high-grade infections highlight the need for identifying target independent combination partners. 52
Our results highlight that combination of forimtamig with CELMoDs during early treatment cycles can prevent tumor escape and substantiate recent preclinical data reporting enhanced efficacy in combination with alnuctamab. 31 , 53 , 54 Our data confirm a dual mode of action of mezigdomide as we observed tumor cell killing of autologous MMPCs in the absence of TIL activation and boosted T-cell response in combination with forimtamig. Mezigdomide, when dosed at least 3 times a week, was identified as an alternative combination partner for prevention of relapse to forimtamig. Considering the reported on-target off-tumor toxicities related to GPRC5D-TCB treatment, the significant increase in PFS observed for low-dose forimtamig and intermittent dosing of mezigdomide could enable a broad therapeutic window for this novel combination although mechanistic understanding of tumor escape will be critical. This assumption is underlined by only moderate increase in T-cell cytokines and a decrease of monocyte-derived cytokines observed for the most efficacious regimens. 30 , 54 The observed trend toward higher expression of exhaustion markers such as PD-1 and LAG-3 suggests a potential benefit of testing triple combinations of forimtamig, CELMoDs, and checkpoint inhibitors. 55 Although more frequent dosing of mezigdomide was correlated with better PFS, bystander effects such as reduced B-cell numbers are to be monitored. Although our data confirm forimtamig to represent an attractive combination partner to boost efficacy against MMPCs, more advanced preclinical models will be required explore impact of combinations on on-target off-tumor toxicities as well as infections.
In summary, we provide evidence for the best-in-class potential of the 2+1 TCB forimtamig for the treatment of MM, both as a monotherapy and in combination with other agents supporting the currently undergoing phase 1 clinical testing in patients with RRMM.
Conflict-of-interest disclosure: J.E., T.F., R.R., C.H., T.P., G.H., R.F., S.G., R.A., C. Kirstenpfad, L.K., S. Lechner, A.B., I.D., A.-M.B., J.A., I.L., F.C., S. Leclair, T.C., G.F., C.D., L.M., C.B., Q.D., F.O., F.B., C.G., S.D., T.K., M.K., M.B., S. Lechner, and P.U. are employees of Roche. C.K., A.C., M.M., L. Bernasconi, and W.X. are former employees of Roche. S.D., F.B., C. Klein, I.D., T.F., J.A., L.K., C.H., F.C., M.B., and M.M. hold stock options of F. Hoffmann-La Roche. J.E., T.F., M.B., S.L., T.C., A.C., A.B., A.-M.B., I.D., F.B., L.K., F.C., G.F., C.G., S.D., C. Kirstenpfad, T.K., C.H., M.M., L.B., S. Leclair, W.X., C. Klein, and P.U. declare patents with Roche. The remaining authors declare no competing financial interests.
References
1 Ludwig H Durie SN Meckl A Hinke A Durie B Multiple myeloma incidence and mortality around the globe; interrelations between health access and quality, economic resources, and patient empowerment Oncol 25 9 2020 e1406 e1413 10.1634/theoncologist.2020-0141 PMC7485361 32335971
2 Berdeja JG Madduri D Usmani SZ Ciltacabtagene autoleucel, a B-cell maturation antigen-directed chimeric antigen receptor T-cell therapy in patients with relapsed or refractory multiple myeloma (CARTITUDE-1): a phase 1b/2 open-label study Lancet 398 10297 2021 314 324 34175021 10.1016/S0140-6736(21)00933-8
3 Munshi NC Jr. Shah N Anderson LD Jr. Idecabtagene vicleucel in relapsed and refractory multiple myeloma N Engl J Med 384 8 2021 705 716 33626253 10.1056/NEJMoa2024850
4 San-Miguel J Dhakal B Yong K Cilta-cel or standard care in lenalidomide-refractory multiple myeloma N Engl J Med 389 4 2023 335 347 37272512 10.1056/NEJMoa2303379
5 O’Neill C van de Donk NWCJ T-cell redirecting bispecific antibodies in multiple myeloma: current landscape and future directions EJHaem 4 3 2023 811 822 37601851 10.1002/jha2.729 PMC10435697
6 Noori M Yazdanpanah N Rezaei N Safety and efficacy of T-cell-redirecting bispecific antibodies for patients with multiple myeloma: a systematic review and meta-analysis Cancer Cell Int 23 1 2023 193 37670301 10.1186/s12935-023-03045-y PMC10478206
7 Lee H Neri P Bahlis NJ BCMA- or GPRC5D-targeting bispecific antibodies in multiple myeloma: efficacy, safety and resistance mechanisms Blood J 143 13 2024 1211 1217 10.1182/blood.2023022499 38194680
8 Lee H Neri P Bahlis NJ Current use of bispecific antibodies to treat multiple myeloma Hematology 2023 1 2023 332 339 38066842 10.1182/hematology.2023000433 PMC10727080
9 Lee H Ahn S Maity R Immune and genome profiling of myeloma patients treated with sequential immunotherapies reveal differential non-overlapping mechanisms of resistance [abstract] Blood 142 suppl 1 2023 1945
10 Samur MK Fulciniti M Aktas Samur A Biallelic loss of BCMA as a resistance mechanism to CAR T cell therapy in a patient with multiple myeloma Nat Commun 12 1 2021 868 33558511 10.1038/s41467-021-21177-5 PMC7870932
11 Letouzé E Moreau P Munshi NC Samur M Minvielle S Touzeau C Mechanisms of resistance to bispecific T cell engagers in multiple myeloma and their clinical implications Blood Adv 8 11 2024 2952 2959 38513088 10.1182/bloodadvances.2023012354 PMC11302375
12 Lesokhin AM Tomasson MH Arnulf B Elranatamab in relapsed or refractory multiple myeloma: phase 2 MagnetisMM-3 trial results Nat Med 29 9 2023 2259 2267 37582952 10.1038/s41591-023-02528-9 PMC10504075
13 Chari A Minnema MC Berdeja JG Talquetamab, a T-cell–redirecting GPRC5D bispecific antibody for multiple myeloma N Engl J Med 387 24 2022 2232 2244 36507686 10.1056/NEJMoa2204591
14 Cliff ERS Mian H Mohyuddin GR Teclistamab in relapsed or refractory multiple myeloma N Engl J Med 387 18 2022 1721 1722 10.1056/NEJMc2211969 36322860
15 Bacac M Umaña P Herter S CD20 Tcb (RG6026), a novel “2:1” T cell bispecific antibody for the treatment of B cell malignancies Blood 128 22 2016 1836
16 Lee H Ahn S Maity R Mechanisms of antigen escape from BCMA- or GPRC5D-targeted immunotherapies in multiple myeloma Nat Med 29 9 2023 2295 2306 37653344 10.1038/s41591-023-02491-5 PMC10504087
17 Rodriguez-Otero P van de Donk NWCJ Pillarisetti K GPRC5D as a novel target for the treatment of multiple myeloma: a narrative review Blood Cancer J 14 1 2024 24 38307865 10.1038/s41408-023-00966-9 PMC10837198
18 Verkleij CPM Broekmans MEC van Duin M Preclinical activity and determinants of response of the GPRC5DxCD3 bispecific antibody talquetamab in multiple myeloma Blood Adv 5 8 2021 2196 2215 33890981 10.1182/bloodadvances.2020003805 PMC8095149
19 Inoue S Nambu T Shimomura T The RAIG family member, GPRC5D, is associated with hard-keratinized structures J Invest Dermatol 122 3 2004 565 573 15086536 10.1046/j.0022-202X.2004.12628.x
20 Hasselbalch Riley C Hutchings M Yoon S-S S180: RG6234, a novel GPRC5D T-cell engaging bispecific antibody, induces rapid responses in patients with relapsed/refractory multiple myeloma: preliminary results from a first-in-human trial HemaSphere 6 2022 81 82
21 Tomita U Ishimoto Y Ri M A novel T cell-redirecting anti-GPRC5D × CD3 bispecific antibody with potent antitumor activity in multiple myeloma preclinical models Sci Rep 14 1 2024 5135 38429446 10.1038/s41598-024-55143-0 PMC10907593
22 Smith EL Harrington K Staehr M GPRC5D is a target for the immunotherapy of multiple myeloma with rationally designed CAR T cells Sci Transl Med 11 485 2019 eaau7746 10.1126/scitranslmed.aau7746 PMC7508042 30918115
23 Pillarisetti K Edavettal S Mendonça M A T-cell–redirecting bispecific G-protein–coupled receptor class 5 member D x CD3 antibody to treat multiple myeloma Blood 135 15 2020 1232 1243 32040549 10.1182/blood.2019003342 PMC7146017
24 Mailankody S Brentjens RJ Smith EL GPRC5D-targeted CAR T cells for myeloma N. Engl. J. Med 387 24 2022 2295 2296 36516098 10.1056/NEJMc2213985
25 Zhang M Wei G Zhou L GPRC5D CAR T cells (OriCAR-017) in patients with relapsed or refractory multiple myeloma (POLARIS): a first-in-human, single-centre, single-arm, phase 1 trial Lancet Haematol 10 2 2023 e107 e116 36725117 10.1016/S2352-3026(22)00372-6
26 Schlothauer T Herter S Koller CF Novel human IgG1 and IgG4 Fc-engineered antibodies with completely abolished immune effector functions Protein Eng Des Sel 29 10 2016 457 466 27578889 10.1093/protein/gzw040
27 Kurtenbach S Mayer C Pelz T Hatt H Leese F Neuhaus EM Molecular evolution of a chordate specific family of G protein-coupled receptors BMC Evol Biol 11 1 2011 234 21827690 10.1186/1471-2148-11-234 PMC3238225
28 Li F Wu G Peng H A novel T-cell-engaging therapeutical antibody against GPRC5D&BCMA for multiple myeloma treatment [abstract] Cancer Res 83 7 suppl 2023 Abstract 1875
29 Mi X Penson A Abdel-Wahab O Mailankody S Genetic basis of relapse after GPRC5D-targeted CAR T cells [abstract] Blood 142 suppl 1 2023 336 37819961 10.1056/NEJMc2308544 PMC11660085
30 Wong L Narla RK Leisten J CC-92480, a novel cereblon E3 ligase modulator, is synergistic with dexamethasone, bortezomib, and daratumumab in multiple myeloma [abstract] Blood 134 suppl 1 2019 1815
31 Thakurta A Pierceall WE Amatangelo MD Flynt E Agarwal A Developing next generation immunomodulatory drugs and their combinations in multiple myeloma Oncotarget 12 15 2021 1555 1563 34316334 10.18632/oncotarget.27973 PMC8310669
32 Walker BA Mavrommatis K Wardell CP A high-risk, double-Hit, group of newly diagnosed myeloma identified by genomic analysis Leukemia 33 1 2019 159 170 29967379 10.1038/s41375-018-0196-8 PMC6326953
33 Goldsmith SR Vij R Evolving paradigms of therapy for multiple myeloma: state of the art and future directions JCO Oncol. Pr 17 7 2021 415 418 10.1200/OP.21.00210 34110937
34 Cohen YC Morillo D Gatt ME First results from the RedirecTT-1 study with teclistamab (tec) + talquetamab (tal) simultaneously targeting BCMA and GPRC5D in patients (pts) with relapsed/refractory multiple myeloma (RRMM) J Clin Oncol 41 16 suppl 2023 8002
35 Bhatt P Kloock C Comenzo R Relapsed/refractory multiple myeloma: a review of available therapies and clinical scenarios encountered in myeloma relapse Curr Oncol 30 2 2023 2322 2347 36826140 10.3390/curroncol30020179 PMC9954856
36 Munshi NC Avet-Loiseau H Anderson KC A large meta-analysis establishes the role of MRD negativity in long-term survival outcomes in patients with multiple myeloma Blood Adv 4 23 2020 5988 5999 33284948 10.1182/bloodadvances.2020002827 PMC7724898
37 Landgren O Devlin S Boulad M Mailankody S Role of MRD status in relation to clinical outcomes in newly diagnosed multiple myeloma patients: a meta-analysis Bone Marrow Transplant 51 12 2016 1565 1568 27595280 10.1038/bmt.2016.222 PMC5571752
38 Mohty M Avet-Loiseau H Malard F Harousseau J-L Potential future direction of measurable residual disease evaluation in multiple myeloma Blood 142 18 2023 1509 1517 37471603 10.1182/blood.2023020284
39 Munshi NC Berdeja JG Lin Y Early MRD negativity to predict deepening myeloma response in relapsed/refractory multiple myeloma (RRMM) patients treated with bb2121 anti-BCMA CAR T cells J Clin Oncol 36 15 suppl 2018 8024
40 Girgis S Wang Lin SX Pillarisetti K Effects of teclistamab and talquetamab on soluble BCMA levels in patients with relapsed/refractory multiple myeloma Blood Adv 7 4 2023 644 648 36006441 10.1182/bloodadvances.2022007625 PMC9979748
41 Jelinek T Zihala D Sevcikova T Beyond the marrow: insights from comprehensive next-generation sequencing of extramedullary multiple myeloma tumors Leukemia 38 6 2024 1323 1333 38493239 10.1038/s41375-024-02206-w PMC11147761
42 Carlo-Stella C Mazza R Manier S RG6234, a GPRC5DxCD3 T-cell engaging bispecific antibody, is highly active in patients (pts) with relapsed/refractory multiple myeloma (RRMM): updated intravenous (IV) and first subcutaneous (SC) results from a phase I dose-escalation study [abstract] Blood 140 suppl 1 2022 397 399
43 Scott SA Marin EM Maples KT Prophylactic tocilizumab to prevent cytokine release syndrome (CRS) with teclistamab: a single-center experience Blood Cancer J 13 1 2023 191 38114481 10.1038/s41408-023-00963-y PMC10730907
44 Sam J Hofer T Kuettel C CD19-CD28: an affinity-optimized CD28 agonist for combination with glofitamab (CD20-TCB) as off-the-shelf immunotherapy Blood 143 21 2024 2152 2165 38437725 10.1182/blood.2023023381 PMC11143537
45 Searle E Quach H Wong SW Teclistamab in combination with subcutaneous daratumumab and lenalidomide in patients with multiple myeloma: results from one cohort of MajesTEC-2, a phase1b, multicohort study [abstract] Blood 140 suppl 1 2022 394 396
46 Dholaria BR Weisel K Mateos M-V Talquetamab (tal) + daratumumab (dara) in patients (pts) with relapsed/refractory multiple myeloma (RRMM): updated TRIMM-2 results J Clin Oncol 41 16 suppl 2023 8003
47 Matous J Biran N Perrot A Talquetamab + pomalidomide in patients with relapsed/refractory multiple myeloma: safety and preliminary efficacy results from the phase 1b MonumenTAL-2 Study [abstract] Blood 142 suppl 1 2023 1014
48 Matsushita M Kashiwazaki S Kamiko S Immunomodulatory effect of proteasome inhibitors via the induction of immunogenic cell death in myeloma cells Pharmaceuticals 16 10 2023 1367 37895838 10.3390/ph16101367 PMC10609901
49 Friedrich MJ Neri P Kehl N The pre-existing T cell landscape determines the response to bispecific T cell engagers in multiple myeloma patients Cancer Cell 41 4 2023 711 725.e6 36898378 10.1016/j.ccell.2023.02.008
50 Dekhtiarenko I Lelios I Jacob W Co-expression of GPRC5D, FcRH5 and BCMA suggests that targeting more than one cell surface marker may be a viable strategy in relapsed/refractory multiple myeloma (RRMM): biomarker results from the phase I study of forimtamig, a GPRC5DxCD3 bispecific antibody [abstract] Blood 142 suppl 1 2023 1948
51 Jakubowiak AJ Anguille S Karlin L Updated results of talquetamab, a GPRC5D×CD3 bispecific antibody, in patients with relapsed/refractory multiple myeloma with prior exposure to T-cell redirecting therapies: results of the phase 1/2 MonumenTAL-1 Study [abstract] Blood 142 suppl 1 2023 3377
52 Reynolds G Cliff ERS Mohyuddin GR Infections following bispecific antibodies in myeloma: a systematic review and meta-analysis Blood Adv 7 19 2023 5898 5903 37467036 10.1182/bloodadvances.2023010539 PMC10558589
53 Gaffney B Shi Y de Jong P Mezigdomide (CC-92480), a novel cereblon E3 ligase modulator, induces vulnerability of multiple myeloma cells to T-cell-mediated killing [abstract] Blood 140 suppl 1 2022 7108 7109
54 Paiva B Gaffney B Burnett K Synergistic antitumor activity of alnuctamab (ALNUC; BMS-986349; CC-93269), a BCMA 2 + 1 T cell engager (TCE), and CELMoD agents in multiple myeloma (MM) preclinical models [abstract] Blood 140 suppl 1 2022 7054 7055
55 Verkleij CPM O’Neill CA Broekmans MEC T-cell characteristics impact response and resistance to T-cell-redirecting bispecific antibodies in multiple myeloma Clin Cancer Res 30 14 2024 3006 3022 38687588 10.1158/1078-0432.CCR-23-3333
Supplementary Material
Supplemental Figures, Methods, and References
Acknowledgments
The authors thank Martin Weisser, Wolfgang Jacob, Natalie Dimier, Shannon Moore, Cedric Dos-Santos, and Vallari Shah for fruitful discussions and valuable advice; Petra Ulrich, Daniela Geiss, Jennifer Stechele, Petra Falkner, Naailah Roheemun, and Monika Friedrich for technical assistance; Sophie Huber from leadXproAG for the GPRC5D purification; and Yunje Cho for sharing the GPRC5D coordinates with us.
Authorship
Contribution: J.E. designed experiments, analyzed, and interpreted data and wrote the manuscript; A.Z., T.F., R.R., C.H., T.P., G.H., L.K., S. Lechner, A.C., A.-M.B., C. Kirstenpfad, I.D., T.K., I.L., F.C., M.M., T.C., G.F., C.D., S.D., R.F., R.A., L.M., C.G., S. Leclair, W.X., and M.B. designed experiments and analyzed and interpreted data; L. Blanco, S. Lorenz, C.B., Q.D., F.O., F.B., C.G., S.D., M.K., L. Bernasconi, N.C., S.K., S.G., and R.R. performed experiments and collected data; J.A. and C.B. collected and interpreted data; and F.P., B.P., C. Klein, and P.U. interpreted data and supervised the study.
C. Klein and P.U. are joint last authors.
Data are available on request from the corresponding authors, Jan Eckmann ( jan.eckmann@roche.com ) and Pablo Umaña ( pablo.umana@roche.com ).
The online version of this article contains a data supplement.
There is a Blood Commentary on this article in this issue.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked “advertisement” in accordance with 18 USC section 1734.