T cell characteristics impact response and resistance to T cell-redirecting bispecific antibodies in multiple myeloma.

PURPOSE: Bispecific antibodies (BsAbs) directed against B-cell maturation antigen (BCMA; teclistamab) or the orphan G protein-coupled receptor GPRC5D (talquetamab) induce deep and durable responses in heavily pretreated MM patients. However, mechanisms underlying primary and acquired resistance remain poorly understood. EXPERIMENTAL DESIGN: The anti-MM activity of teclistamab and talquetamab was evaluated in bone marrow (BM) samples from MM patients. T-cell phenotype and function were assessed in BM/peripheral blood samples obtained from MM patients who were treated with these BsAbs. RESULTS: In ex vivo killing assays with 41 BM samples from BsAb-naïve MM patients, teclistamab- and talquetamab-mediated MM lysis were strongly correlated (r=0.73, P<0.0001). Both BsAbs exhibited poor activity in samples with high regulatory T-cell (Treg) numbers and a low T-cell/MM cell-ratio. Furthermore, comprehensive phenotyping of BM samples derived from patients treated with teclistamab or talquetamab, revealed that high frequencies of PD-1+ CD4+ T-cells, CTLA4+ CD4+ T-cells, and CD38+ CD4+ T-cells were associated with primary resistance. Although this lack of response was linked to modest increase in expression of inhibitory receptors, increasing T-cell/MM cell-ratios by adding extra T-cells enhanced sensitivity to BsAbs. Further, treatment with BsAbs resulted in an increased proportion of T-cells expressing exhaustion markers (PD-1, TIGIT, and TIM-3), which was accompanied by reduced T-cell proliferative potential and cytokine secretion, as well as impaired anti-tumor efficacy in ex vivo experiments. CONCLUSIONS: Primary resistance is characterized by a low T-cell/MM cell-ratio and Treg-driven immunosuppression, while reduced T-cell fitness due to continuous BsAb-mediated T-cell activation may contribute to development of acquired resistance.

Teclistamab impairs humoral immunity in patients with heavily pretreated myeloma: importance of immunoglobulin supplementation.

ABSTRACT: Teclistamab and other B-cell maturation antigen (BCMA)-targeting bispecific antibodies (BsAbs) have substantial activity in patients with heavily pretreated multiple myeloma (MM) but are associated with a high rate of infections. BCMA is also expressed on normal plasma cells and mature B cells, which are essential for the generation of a humoral immune response. The aim of this study was to improve the understanding of the impact of BCMA-targeting BsAbs on humoral immunity. The impact of teclistamab on polyclonal immunoglobulins and B cell counts was evaluated in patients with MM who received once-weekly teclistamab 1.5 mg/kg subcutaneously. Vaccination responses were assessed in a subset of patients. Teclistamabinduced rapid depletion of peripheral blood B cells in patients with MM and eliminated normal plasma cells in ex vivo assays. In addition, teclistamab reduced the levels of polyclonal immunoglobulins (immunoglobulin G [IgG], IgA, IgE, and IgM), without recovery over time while receiving teclistamab therapy. Furthermore, response to vaccines against Streptococcus pneumoniae, Haemophilus influenzae type B, and severe acute respiratory syndrome coronavirus 2 was severely impaired in patients treated with teclistamab compared with vaccination responses observed in patients with newly diagnosed MM or relapsed/refractory MM. Intravenous immunoglobulin (IVIG) use was associated with a significantly lower risk of serious infections among patients treated with teclistamab (cumulative incidence of infections at 6 months: 5.3% with IVIG vs 54.8% with observation only [P < .001]). In conclusion, our data show severe defects in humoral immunity induced by teclistamab, the impact of which can be mitigated by the use of immunoglobulin supplementation. This trial was registered at www.ClinicalTrials.gov as #NCT04557098.

T-cell redirecting bispecific and trispecific antibodies in multiple myeloma beyond BCMA.

PURPOSE OF REVIEW: B-cell maturation antigen (BCMA)-directed T-cell immunotherapies, such as chimeric antigen receptor T-cells (CAR T-cells) and bispecific antibodies (BsAbs) have markedly improved the survival of triple-class refractory multiple myeloma (MM). However, the majority of patients still develops disease progression, underlining the need for new agents for these patients. RECENT FINDINGS: Novel T-cell redirecting BsAbs targeting alternative tumor-associated antigens have shown great promise in heavily pretreated MM, including patients previously exposed to BCMA-directed therapies. This includes the G-protein-coupled receptor class 5 member D (GPRC5D)-targeting BsAbs talquetamab and forimtamig, as well as the Fc receptor-homolog 5 (FcRH5)-targeting BsAb cevostamab. Toxicity associated with these BsAbs includes cytokine-release syndrome, cytopenias, and infections. In addition, GPRC5D-targeting BsAbs are associated with specific 'on target/off tumor' toxicities including rash, nail disorders, and dysgeusia. Trispecifc antibodies targeting two different MM-associated antigens to prevent antigen escape are in early clinical development, as well as trispecific antibodies (TsAbs) that provide an additional co-stimulatory signal to T-cells to prevent their exhaustion. SUMMARY: Various T-cell redirecting BsAbs are in advanced stages of clinical development with promising activity and a manageable toxicity profile. Ongoing studies are evaluating combination strategies, fixed-duration treatment, and use of BsAbs in earlier lines of therapy. TsAbs hold great promise for the future.

NK Cell Phenotype Is Associated With Response and Resistance to Daratumumab in Relapsed/Refractory Multiple Myeloma.

The CD38-targeting antibody daratumumab has marked activity in multiple myeloma (MM). Natural killer (NK) cells play an important role during daratumumab therapy by mediating antibody-dependent cellular cytotoxicity via their FcγRIII receptor (CD16), but they are also rapidly decreased following initiation of daratumumab treatment. We characterized the NK cell phenotype at baseline and during daratumumab monotherapy by flow cytometry and cytometry by time of flight to assess its impact on response and development of resistance (DARA-ATRA study; NCT02751255). At baseline, nonresponding patients had a significantly lower proportion of CD16+ and granzyme B+ NK cells, and higher frequency of TIM-3+ and HLA-DR+ NK cells, consistent with a more activated/exhausted phenotype. These NK cell characteristics were also predictive of inferior progression-free survival and overall survival. Upon initiation of daratumumab treatment, NK cells were rapidly depleted. Persisting NK cells exhibited an activated and exhausted phenotype with reduced expression of CD16 and granzyme B, and increased expression of TIM-3 and HLA-DR. We observed that addition of healthy donor-derived purified NK cells to BM samples from patients with either primary or acquired daratumumab-resistance improved daratumumab-mediated MM cell killing. In conclusion, NK cell dysfunction plays a role in primary and acquired daratumumab resistance. This study supports the clinical evaluation of daratumumab combined with adoptive transfer of NK cells.

Redirecting T-cell Activity with Anti-BCMA/Anti-CD3 Bispecific Antibodies in Chronic Lymphocytic Leukemia and Other B-cell Lymphomas.

T-cell redirecting bispecific antibodies hold high promise for treatment of B-cell malignancies. B-cell maturation antigen (BCMA) exhibits high expression on normal and malignant mature B cells including plasma cells, which can be enhanced by inhibition of γ-secretase. BCMA is considered a validated target in multiple myeloma but whether mature B-cell lymphomas can be targeted by the BCMAxCD3 T-cell redirector teclistamab is currently unknown. BCMA expression on B-cell non-Hodgkin lymphoma and primary chronic lymphocytic leukemia (CLL) cells was assessed by flow cytometry and/or IHC. To assess teclistamab efficacy, cells were treated with teclistamab in presence of effector cells with/without γ-secretase inhibition. BCMA could be detected on all tested mature B-cell malignancy cell lines, while expression levels varied per tumor type. γ-secretase inhibition universally increased BCMA surface expression. These data were corroborated in primary samples from patients with Waldenstrom's macroglobulinemia, CLL, and diffuse large B-cell lymphoma. Functional studies with the B-cell lymphoma cell lines revealed teclistamab-mediated T-cell activation, proliferation, and cytotoxicity. This was independent of the level of BCMA expression, but generally lower in mature B-cell malignancies compared with multiple myeloma. Despite low BCMA levels, healthy donor T cells and CLL-derived T cells induced lysis of (autologous) CLL cells upon addition of teclistamab. These data show that BCMA is expressed on various B-cell malignancies and that lymphoma cell lines and primary CLL can be targeted using teclistamab. Further studies to understand the determinants of response to teclistamab are required to identify which other diseases might be suitable for teclistamab targeting. Significance: Besides reported BCMA expression on multiple myeloma, we demonstrate BCMA can be detected and enhanced using γ-secretase inhibition on cell lines and primary material of various B-cell malignancies. Furthermore, using CLL we demonstrate that low BCMA-expressing tumors can be targeted efficiently using the BCMAxCD3 DuoBody teclistamab.

Current State of the Art and Prospects of T Cell-Redirecting Bispecific Antibodies in Multiple Myeloma.

Multiple myeloma (MM) patients eventually develop multi-drug-resistant disease with poor survival. Hence, the development of novel treatment strategies is of great importance. Recently, different classes of immunotherapeutic agents have shown great promise in heavily pre-treated MM, including T cell-redirecting bispecific antibodies (BsAbs). These BsAbs simultaneously interact with CD3 on effector T cells and a tumor-associated antigen on MM cells, resulting in redirection of T cells to MM cells. This leads to the formation of an immunologic synapse, the release of granzymes/perforins, and subsequent tumor cell lysis. Several ongoing phase 1 studies show substantial activity and a favorable toxicity profile with BCMA-, GPRC5D-, or FcRH5-targeting BsAbs in heavily pre-treated MM patients. Resistance mechanisms against BsAbs include tumor-related features, T cell characteristics, and impact of components of the immunosuppressive tumor microenvironment. Various clinical trials are currently evaluating combination therapy with a BsAb and another agent, such as a CD38-targeting antibody or an immunomodulatory drug (e.g., pomalidomide), to further improve response depth and duration. Additionally, the combination of two BsAbs, simultaneously targeting two different antigens to prevent antigen escape, is being explored in clinical studies. The evaluation of BsAbs in earlier lines of therapy, including newly diagnosed MM, is warranted, based on the efficacy of BsAbs in advanced MM.

Efficacy and safety of daratumumab combined with all-trans retinoic acid in relapsed/refractory multiple myeloma.

The efficacy of daratumumab depends partially on CD38 expression on multiple myeloma (MM) cells. We have previously shown that all-trans retinoic acid (ATRA) upregulates CD38 expression and reverts daratumumab-resistance ex vivo. We therefore evaluated the optimal dose, efficacy, and safety of daratumumab combined with ATRA in patients with daratumumab-refractory MM in a phase 1/2 study (NCT02751255). In part A of the study, 63 patients were treated with daratumumab monotherapy. Fifty patients with daratumumab-refractory MM were subsequently enrolled in part B and treated with daratumumab (reintensified schedule) combined with ATRA until disease progression. The recommended phase 2 dose of ATRA in combination with daratumumab was defined as 45 mg/m2. At this dose, the overall response rate (ORR) was 5%, indicating that the primary endpoint (ORR ≥15%) was not met. However, most patients (66%) achieved at least stable disease. After a median follow-up of 43 months, the median progression-free survival (PFS) for all patients was 2.8 months. Patients who previously achieved at least a partial response or minimal response/stable disease with prior daratumumab monotherapy had a significantly longer PFS compared with patients who immediately progressed during daratumumab as single agent (median PFS 3.4 and 2.8 vs 1.3 months). The median overall survival was 19.1 months. The addition of ATRA did not increase the incidence of adverse events. Flow cytometric analysis revealed that ATRA temporarily increased CD38 expression on immune cell subsets. In conclusion, the addition of ATRA and reintensification of daratumumab had limited activity in patients with daratumumab-refractory MM, which may be explained by the transient upregulation of CD38 expression. This trial was registered at www.clinicaltrials.gov as #NCT02751255.

Monitoring the M-protein of multiple myeloma patients treated with a combination of monoclonal antibodies: the laboratory solution to eliminate interference.

OBJECTIVES: The therapeutic monoclonal antibody (t-mAb) daratumumab, used to treat multiple myeloma (MM) patients, interferes with routine, electrophoretic based M-protein diagnostics. Electrophoretic response assessment becomes increasingly difficult when multiple t-mAbs are combined for use in a single patient. This is the first study to address the analytical challenges of M-protein monitoring when multiple t-mAbs are combined. METHODS: In this proof-of-principle study we evaluate two different methods to monitor M-protein responses in three MM patients, who receive both daratumumab and nivolumab. The double hydrashift assay aims to resolve t-mAb interference on immunofixation. The MS-MRD (mass spectrometry minimal residual disease) assay measures clonotypic peptides to quantitate both M-protein and t-mAb concentrations. RESULTS: After exposure to daratumumab and nivolumab, both t-mAbs become visible on immunofixation electrophoresis (IFE) as two IgG-kappa bands that migrate close to each other at the cathodal end of the γ-region. In case the M-protein co-migrates with these t-mAbs, the observed interference was completely abolished with the double IFE hydrashift assay. In all three patients the MS-MRD assay was also able to distinguish the M-protein from the t-mAbs. Additional advantage of the MS-MRD assay is that this multiplex assay is more sensitive and allows quantitative M-protein-, daratumumab- and nivolumab-monitoring. CONCLUSIONS: Daratumumab and nivolumab interfere with electrophoretic M-protein diagnostics. However, the M-protein can be distinguished from both t-mAbs by use of a double hydrashift assay. The MS-MRD assay provides an alternative method that allows sensitive and simultaneous quantitative monitoring of both the M-protein and t-mAbs.

Efficacy and Safety of Durvalumab Combined with Daratumumab in Daratumumab-Refractory Multiple Myeloma Patients.

Daratumumab is active both as a single agent and in combination with other agents in multiple myeloma (MM) patients. However, the majority of patients will develop daratumumab-refractory disease, which carries a poor prognosis. Since daratumumab also has immunomodulatory effects, addition of the PD-L1 blocking antibody durvalumab at the time of progression may reverse daratumumab-resistance. The efficacy and safety of daratumumab and durvalumab in daratumumab-refractory relapsed/refractory MM patients was evaluated in this prospective, single-arm phase 2 study (NCT03000452). None of the 18 enrolled patients achieved PR or better. The frequency of serious adverse events was 38.9%, with one patient experiencing an immune related adverse event (grade 2 hyperthyroidism). No infusion-related reactions were observed. Analysis of tumor- and immune cell characteristics was performed on bone marrow samples obtained at baseline and during treatment. Daratumumab combined with durvalumab reduced the frequency of regulatory T-cells and decreased the proportion of T-cells expressing LAG3 and CD8+ T-cells expressing TIM-3, without altering T- and NK-cell frequencies. Durvalumab did not affect tumor cell characteristics associated with daratumumab resistance. In conclusion, the addition of durvalumab to daratumumab following development of daratumumab-resistance was associated with an acceptable toxicity profile, but was not effective. This indicates that inhibition of the PD-1/PD-L1 signaling pathway at the time of daratumumab-resistance is insufficient to reverse daratumumab-resistance.

Preclinical activity and determinants of response of the GPRC5DxCD3 bispecific antibody talquetamab in multiple myeloma.

Cell surface expression levels of GPRC5D, an orphan G protein-coupled receptor, are significantly higher on multiple myeloma (MM) cells, compared with normal plasma cells or other immune cells, which renders it a promising target for immunotherapeutic strategies. The novel GPRC5D-targeting T-cell redirecting bispecific antibody, talquetamab, effectively kills GPRC5D+ MM cell lines in the presence of T cells from both healthy donors or heavily pretreated MM patients. In addition, talquetamab has potent anti-MM activity in bone marrow (BM) samples from 45 patients, including those with high-risk cytogenetic aberrations. There was no difference in talquetamab-mediated killing of MM cells from newly diagnosed, daratumumab-naïve relapsed/refractory (median of 3 prior therapies), and daratumumab-refractory (median of 6 prior therapies) MM patients. Tumor cell lysis was accompanied by T-cell activation and degranulation, as well as production of pro-inflammatory cytokines. High levels of GPRC5D and high effector:target ratio were associated with improved talquetamab-mediated lysis of MM cells, whereas an increased proportion of T cells expressing PD-1 or HLA-DR, and elevated regulatory T-cell (Treg) counts were associated with suboptimal killing. In cell line experiments, addition of Tregs to effector cells decreased MM cell lysis. Direct contact with bone marrow stromal cells also impaired the efficacy of talquetamab. Combination therapy with daratumumab or pomalidomide enhanced talquetamab-mediated lysis of primary MM cells in an additive fashion. In conclusion, we show that the GPRC5D-targeting T-cell redirecting bispecific antibody talquetamab is a promising novel antimyeloma agent. These results provide the preclinical rationale for ongoing studies with talquetamab in relapsed/refractory MM.

Preclinical Rationale for Targeting the PD-1/PD-L1 Axis in Combination with a CD38 Antibody in Multiple Myeloma and Other CD38-Positive Malignancies.

The CD38-targeting antibody daratumumab mediates its anti-myeloma activities not only through Fc-receptor-dependent effector mechanisms, but also by its effects on T-cell immunity through depletion of CD38+ regulatory T-cells, regulatory B-cells, and myeloid-derived suppressor cells. Therefore, combining daratumumab with modulators of other potent immune inhibitory pathways, such as the PD-1/PD-L1 axis, may further improve its efficacy. We show that multiple myeloma (MM) cells from relapsed/refractory patients have increased expression of PD-L1, compared to newly diagnosed patients. Furthermore, PD-1 is upregulated on T-cells from both newly diagnosed and relapsed/refractory MM patients, compared to healthy controls. In short-term experiments with bone marrow samples from MM patients, daratumumab-mediated lysis was mainly associated with the MM cells' CD38 expression levels and the effector (NK-cells/monocytes/T-cells)-to-target ratio, but not with the PD-L1 expression levels or PD-1+ T-cell frequencies. Although PD-1 blockade with nivolumab did not affect MM cell viability or enhanced daratumumab-mediated lysis in short-term ex vivo experiments, nivolumab resulted in a mild but clear increase in T-cell numbers. Moreover, with a longer treatment duration, PD-1 blockade markedly improved anti-CD38 antibody-mediated cytotoxicity in vivo in murine CD38+ tumor models. In conclusion, dual targeting of CD38 and PD-1 may represent a promising strategy for treating MM and other CD38-positive malignancies.

T-cell redirecting bispecific antibodies targeting BCMA for the treatment of multiple myeloma.

B-cell maturation antigen (BCMA)-targeting bispecific antibodies and bispecific T-cell engagers (BiTEs) redirect T-cells to BCMA-expressing multiple myeloma (MM) cells. These MM cells are subsequently eliminated via various mechanisms of action including the release of granzymes and perforins. Several phase 1, dose-escalation studies show pronounced activity of BCMA-targeting bispecific antibodies, including teclistamab, AMG420 and CC-93269, in heavily pretreated MM patients. Cytokine release syndrome is the most common adverse event, which can be adequately managed with tocilizumab or steroids. Several clinical trials are currently evaluating combination therapy with a BCMA-specific bispecific antibody, based on preclinical findings showing that immunomodulatory drugs or CD38-targeting antibodies enhance the activity of bispecific antibodies. In addition, bispecific antibodies, targeting other MM cell surface antigens (i. e. GPRC5D, CD38 and FcRH5), are also evaluated in early phase clinical trials. Such bispecific antibodies, targeting other antigens, may be given to patients with low baseline BCMA expression, disease with substantial heterogeneity in BCMA expression, following prior BCMA-targeted therapy, or combined with BCMA bispecific antibodies to prevent development of antigen escape.

Immunotherapy in multiple myeloma: when, where, and for who?

PURPOSE OF REVIEW: Immunotherapy is transforming treatment of multiple myeloma patients in all stages of their disease. This review will discuss recent developments in immunotherapy in multiple myeloma with a focus on antibodies, antibody-drug conjugates, and T-cell-redirection strategies. RECENT FINDINGS: CD38-targeting antibodies have single agent activity in multiple myeloma, and especially when combined with other drugs, are improving the clinical outcome of patients with newly diagnosed or relapsed/refractory multiple myeloma. Also the SLAMF7-targeting antibody, elotuzumab, improves the survival of relapsed/refractory multiple myeloma patients, when it is combined with either lenalidomide or pomalidomide. Several novel immunotherapies, such as chimeric antigen receptor T cells, antibody-drug conjugates, and bispecific antibodies, are active in patients who developed resistance to all currently available antimultiple myeloma drugs, including immunomodulatory drugs, proteasome inhibitors, and CD38 antibodies. These new immunotherapeutic agents frequently target B-cell maturation antigen, which is highly and uniformly expressed on multiple myeloma cells. However, other targets, such as GPRC5D, are also being investigated. SUMMARY: Immunotherapy is incorporated into first-line and relapse regimens, and is improving the survival of both newly diagnosed and relapsed/refractory multiple myeloma patients.

Preclinical Activity of JNJ-7957, a Novel BCMA×CD3 Bispecific Antibody for the Treatment of Multiple Myeloma, Is Potentiated by Daratumumab.

PURPOSE: Multiple myeloma (MM) patients with disease refractory to all available drugs have a poor outcome, indicating the need for new agents with novel mechanisms of action. EXPERIMENTAL DESIGN: We evaluated the anti-MM activity of the fully human BCMA×CD3 bispecific antibody JNJ-7957 in cell lines and bone marrow (BM) samples. The impact of several tumor- and host-related factors on sensitivity to JNJ-7957 therapy was also evaluated. RESULTS: We show that JNJ-7957 has potent activity against 4 MM cell lines, against tumor cells in 48 of 49 BM samples obtained from MM patients, and in 5 of 6 BM samples obtained from primary plasma cell leukemia patients. JNJ-7957 activity was significantly enhanced in patients with prior daratumumab treatment, which was partially due to enhanced killing capacity of daratumumab-exposed effector cells. BCMA expression did not affect activity of JNJ-7957. High T-cell frequencies and high effector:target ratios were associated with improved JNJ-7957-mediated lysis of MM cells. The PD-1/PD-L1 axis had a modest negative impact on JNJ-7957 activity against tumor cells from daratumumab-naïve MM patients. Soluble BCMA impaired the ability of JNJ-7957 to kill MM cells, although higher concentrations were able to overcome this negative effect. CONCLUSIONS: JNJ-7957 effectively kills MM cells ex vivo, including those from heavily pretreated MM patients, whereby several components of the immunosuppressive BM microenvironment had only modest effects on its killing capacity. Our findings support the ongoing trial with JNJ-7957 as single agent and provide the preclinical rationale for evaluating JNJ-7957 in combination with daratumumab in MM.

CD38-targeted therapy with daratumumab reduces autoantibody levels in multiple myeloma patients.

Autoantibody-producing plasma cells are frequently resistant to conventional immunosuppressive treatments and B-cell depletion therapy. As a result of this resistance, autoreactive plasma cells survive conventional therapy, resulting in persistent autoantibody production and inflammation. CD38 is highly and uniformly expressed on normal and malignant plasma cells. Daratumumab is the first in class CD38-targeting monoclonal antibody approved for the treatment of multiple myeloma (MM). To evaluate the potential activity of daratumumab in antibody-mediated autoimmune disorders by targeting autoantibody-producing plasma cells, we evaluated serum levels of autoantibodies in MM patients during daratumumab treatment. We found that 6 out of 41 (15%) had detectable autoantibodies before initiation of daratumumab therapy, and that these autoantibodies rapidly disappeared in 5 out of 6 patients during daratumumab treatment. Our data provide support for the evaluation of daratumumab in patients with autoantibody-dependent autoimmune disorders.