Structures of Two Melanoma-Associated Antigens Suggest Allosteric Regulation of Effector Binding

The MAGE (melanoma associated antigen) protein family are tumour-associated proteins normally present only in reproductive tissues such as germ cells of the testis. The human genome encodes over 60 MAGE genes of which one class (containing MAGE-A3 and MAGE-A4) are exclusively expressed in tumours, making them an attractive target for the development of targeted and immunotherapeutic cancer treatments. Some MAGE proteins are thought to play an active role in driving cancer, modulating the activity of E3 ubiquitin ligases on targets related to apoptosis. Here we determined the crystal structures of MAGE- A3 and MAGE-A4. Both proteins crystallized with a terminal peptide bound in a deep cleft between two tandem-arranged winged helix domains. MAGE-A3 (but not MAGE-A4), is pre- dominantly dimeric in solution. Comparison of MAGE-A3 and MAGE-A3 with a structure of an effector-bound MAGE-G1 suggests that a major conformational rearrangement is required for binding, and that this conformational plasticity may be targeted by allosteric binders

The effect of novel therapies in high-molecular-risk multiple myeloma.

Multiple myeloma is a heterogeneous disease with a prognosis that varies with patient factors, disease burden, tumor biology, and treatments. Certain molecular abnormalities confer a worse prognosis and thus are considered high-risk. These include t(4;14), del(17p), t(14;16), t(14;20), hypodiploidy, and gain(1q)/del(1p). In our previous review in 2013, we discussed the effect of available therapies on prognosis in these high-risk patients. Since then, seven phase 3 clinical trials in relapsed myeloma with 1 to 3 lines of therapy have been conducted, resulting in the approval of panobinostat, ixazomib, daratumumab, and elotuzumab, as well as additional data on carfilzomib. In our current review of these studies, all the novel therapies resulted in an improvement in progression-free survival for high-risk patients, but none of the trials provided clear statistical evidence that they overcame high-risk status. Moreover, there are several limitations in the currently available data. For example, the patients Revised International Staging System score is generally not reported, and even when it is reported, it is usually at the time of initial diagnosis rather than at the time of study entry. Furthermore, the methodology used to determine risk suffers from technologic issues. Finally, the clonal and allele burden and concurrent molecular abnormalities can affect risk status and prognosis. To determine the optimal therapy for high-risk patients, future clinical trials should provide standardized risk assessments for all patients in addition to hazard ratios for Kaplan-Meier survival curves of high-risk patients vs those of standard-risk patients to determine if high-risk status has truly been overcome by a novel agent

Bispecific Antibodies in Multiple Myeloma: Present and Future.

UNLABELLED: Despite many recent advances in therapy, there is still no plateau in overall survival curves in multiple myeloma. Bispecific antibodies are a novel immunotherapeutic approach designed to bind antigens on malignant plasma cells and cytotoxic immune effector cells. Early-phase clinical trials targeting B-cell maturation antigen (BCMA), GPRC5D, and FcRH5 have demonstrated a favorable safety profile, with mainly low-grade cytokine release syndrome, cytopenias, and infections. Although dose escalation is ongoing in several studies, early efficacy data show response rates in the most active dose cohorts between 61% and 83% with many deep responses; however, durability remains to be established. Further clinical trial data are eagerly anticipated. SIGNIFICANCE: Overall survival of triple-class refractory multiple myeloma remains poor. Bispecific antibodies are a novel immunotherapeutic modality with a favorable safety profile and impressive preliminary efficacy in heavily treated patients. Although more data are needed, bispecifics will likely become an integral part of the multiple myeloma treatment paradigm in the near future. Studies in earlier lines of therapy and in combination with other active anti-multiple myeloma agents will help further define the role of bispecifics in multiple myeloma

Subcutaneous daratumumab and hyaluronidase-fihj in newly diagnosed or relapsed/refractory multiple myeloma.

Daratumumab, a human immunoglobulin G1 kappa monoclonal antibody that targets CD38, is currently approved as monotherapy and in varying combinations with approved anti-myeloma regimens in both newly diagnosed multiple myeloma and relapsed refractory multiple myeloma. Originally developed for intravenous administration, the subcutaneous formulation of daratumumab (daratumumab and hyaluronidase-fihj) was recently approved by the US Federal Drug Administration and European Commission in 2020. In clinical trials, compared with the intravenous formulation, subcutaneous daratumumab (Dara-SC) has significantly shorter administration time (median first dose 7 h versus 3-5 min, respectively), lower rates of infusion-related reactions (median first dose 50% versus less than 10%, respectively), and lower volume of infusion (median 500-1000 ml versus 15 ml, respectively). Otherwise, the pharmacokinetics, safety profile, and efficacy are comparable. This review summarizes the pivotal trials that led to the approval of Dara-SC, highlights important clinical considerations for the use of Dara-SC, and provides practical guidelines for the administration of Dara-SC in the clinic

Risk stratification of smoldering multiple myeloma: predictive value of free light chains and group-based trajectory modeling.

We investigated the predictive role for serum free light chain ratio (FLCr) ≥100, bone marrow plasma cell (BMPC) ≥60%, and evolving biomarkers through group-based trajectory modeling (GBTM) as high-risk defining events in 273 smoldering multiple myeloma (SMM) patients with a median follow-up of 74 months. FLCr ≥100 was confirmed as a marker for high-risk progression with a median time to progression (TTP) of 40 months with a 44% risk of progression of disease (PD) at 2 years; however, 44% of FLCr ≥100 also did not progress during follow-up. For patients with BMPC ≥60% by core biopsy, the median TTP was 31 months with a 2-year PD of 41%. GBTM established high-risk trajectories for evolving hemoglobin (eHb; characterized as a 1.57 g/dL decrease in hemoglobin), evolving m-protein (eMP; 64% increase in m-protein), and evolving differences in FLC (edFLC; 169% increase in dFLC) within 1 year of diagnosis associated with a decreased median TTP and an increased 2 year rate of PD. Of all the variables examined, we identify a model where immunoparesis, eHb, eMP, and edFLC were significant predictors for ultra-high-risk progression with a median TTP of only 13 months with 3 or more variables present. Our results not only confirm a more modest 2 year PD associated with FLCr ≥100 and BMPC ≥60 but also suggest that eHb, eMP, and edFLC may help identify an ultra-high-risk SMM group

The Safety and Efficacy of Radiation Therapy with Concurrent Dexamethasone, Cyclophosphamide, Etoposide, and Cisplatin-Based Systemic Therapy for Multiple Myeloma.

INTRODUCTION: The concurrent delivery of radiation therapy (RT) with salvage chemotherapies in the management of relapsed and refractory multiple myeloma (MM) is an area of ongoing investigation. This study examined the safety and efficacy of palliative RT given in the setting of concurrent dexamethasone, cyclophosphamide, etoposide, and cisplatin (DCEP). PATIENTS AND METHODS: Fifty-five patients with MM received RT to 64 different sites within three weeks of receiving DCEP from 2010 to 2020. A median dose of 20 Gray (range 8-32.5 Gy) was delivered in a median of 5 fractions (range 1-15). Patients received a median of 1 cycle (range 1-5) of DCEP. Rates of hematologic and RT toxicity were recorded along with pain, radiographic, and laboratory responses to treatment. RESULTS: RT was completed in 98% of patients. 21% of patients experienced RTOG grade 3+ hematologic toxicity before RT, which increased to 35% one-month post-RT (P = .13) before decreasing to 12% at 3 to 6 months (P = .02). The most common toxicity experienced was thrombocytopenia. Grade 1 to 2 non-hematologic RT-related toxicity was reported in 15% of patients while on treatment and fell to 6% one-month after completing RT. Pain resolved in 94% of patients with symptomatic lesions at baseline. Stable disease or better was observed in 34/39 (87%) of the targeted lesions on surveillance imaging. CONCLUSION: RT administered concurrently with DCEP was well-tolerated by most of the patients in this series, with low rates of hematologic and RT-related toxicity. RT was also very effective, with the vast majority of patients demonstrating resolution of their pain and a significant response on follow-up imaging

MAGE-A inhibit apoptosis and promote proliferation in multiple myeloma through regulation of BIM and p21Cip1.

The type I Melanoma Antigen Gene (MAGE) A3 is a functional target associated with survival and proliferation in multiple myeloma (MM). To investigate the mechanisms of these oncogenic functions, we performed gene expression profiling (GEP) of p53 wild-type human myeloma cell lines (HMCL) after MAGE-A knockdown, which identified a set of 201 differentially expressed genes (DEG) associated with apoptosis, DNA repair, and cell cycle regulation. MAGE knockdown increased protein levels of pro-apoptotic BIM and of the endogenous cyclin-dependent kinase (CDK) inhibitor p21Cip1. Depletion of MAGE-A in HMCL increased sensitivity to the alkylating agent melphalan but not to proteasome inhibition. High MAGEA3 was associated with the MYC and Cell Cycling clusters defined by a network model of GEP data from the CoMMpass database of newly diagnosed, untreated MM patients. Comparative analysis of CoMMpass subjects based on high or low MAGEA3 expression revealed a set of 6748 DEG that also had significant functional associations with cell cycle and DNA replication pathways, similar to that observed in HMCL. High MAGEA3 expression correlated with shorter overall survival after melphalan chemotherapy and autologous stem cell transplantation (ASCT). These results demonstrate that MAGE-A3 regulates Bim and p21Cip1 transcription and protein expression, inhibits apoptosis, and promotes proliferation

Multi-omic analysis of the tumor microenvironment shows clinical correlations in Ph1 study of atezolizumab +/- SoC in MM

Multiple myeloma (MM) remains incurable, and treatment of relapsed/refractory (R/R) disease is challenging. There is an unmet need for more targeted therapies in this setting; deep cellular and molecular phenotyping of the tumor and microenvironment in MM could help guide such therapies. This phase 1b study (NCT02431208) evaluated the safety and efficacy of the anti-programmed death-ligand 1 monoclonal antibody atezolizumab (Atezo) alone or in combination with the standard of care (SoC) treatments lenalidomide (Len) or pomalidomide (Pom) and/or daratumumab (Dara) in patients with R/R MM. Study endpoints included incidence of adverse events (AEs) and overall response rate (ORR). A novel unsupervised integrative multi-omic analysis was performed using RNA sequencing, mass cytometry immunophenotyping, and proteomic profiling of baseline and on-treatment bone marrow samples from patients receiving Atezo monotherapy or Atezo+Dara. A similarity network fusion (SNF) algorithm was applied to preprocessed data. Eighty-five patients were enrolled. Treatment-emergent deaths occurred in 2 patients; both deaths were considered unrelated to study treatment. ORRs ranged from 11.1% (Atezo+Len cohorts, n=18) to 83.3% (Atezo+Dara+Pom cohort, n=6). High-dimensional multi-omic profiling of the tumor microenvironment and integrative SNF analysis revealed novel correlations between cellular and molecular features of the tumor and immune microenvironment, patient selection criteria, and clinical outcome. Atezo monotherapy and SoC combinations were safe in this patient population and demonstrated some evidence of clinical efficacy. Integrative analysis of high dimensional genomics and immune data identified novel clinical correlations that may inform patient selection criteria and outcome assessment in future immunotherapy studies for myeloma

Phase 1 study combining elotuzumab with autologous stem cell transplant and lenalidomide for multiple myeloma.

BACKGROUND: Autologous stem cell transplantation (ASCT) after induction therapy improves disease-free survival for patients with multiple myeloma (MM). While the goal of ASCT is to render a minimal disease state, it is also associated with eradication of immunosuppressive cells, and we hypothesize that early introduction of immunotherapy post-ASCT may provide a window of opportunity to boost treatment efficacy. METHODS: We conducted a phase 1 clinical trial to investigate the application of autologous lymphocyte infusion and anti-SLAMF7 monoclonal antibody, elotuzumab, after ASCT in patients with newly diagnosed MM previously treated with induction therapy. In addition to CD34+ stem cells, peripheral blood mononuclear cells were harvested prior to transplant and infused on day 3 after stem cell infusion to accelerate immune reconstitution and provide autologous natural killer (NK) cells that are essential to the mechanism of elotuzumab. Elotuzumab was administered starting on day 4 and then every 28 days after until 1 year post-ASCT. Cycles 4-12 were administered with standard-of-care lenalidomide maintenance. RESULTS: All subjects were evaluated for safety, and 13 of 15 subjects completed the treatment protocol. At 1 year post-ASCT, the disease status of enrolled subjects was as follows: five stringent complete responses, one complete response, six very good partial responses, one partial response, and two progressive diseases. The treatment plan was well tolerated, with most grade 3 and 4 AEs being expected hematologic toxicities associated with ASCT. Correlative analysis of the immune microenvironment demonstrated a trend toward reduced regulatory T cells during the first 3 months post-transplant followed by an increase in NK cells and monocytes in patients achieving a complete remission. CONCLUSIONS: This phase 1 clinical trial demonstrates that early introduction of immunotherapy after ASCT is well tolerated and shows promising disease control in patients with MM, accompanied by favorable changes in the immune microenvironment. TRIAL REGISTRATION NUMBER: NCT02655458

Dual Targeting of CDK4 and ARK5 Using a Novel Kinase Inhibitor ON123300 Exerts Potent Anticancer Activity against Multiple Myeloma.

Multiple myeloma is a fatal plasma cell neoplasm accounting for over 10,000 deaths in the United States each year. Despite new therapies, multiple myeloma remains incurable, and patients ultimately develop drug resistance and succumb to the disease. The response to selective CDK4/6 inhibitors has been modest in multiple myeloma, potentially because of incomplete targeting of other critical myeloma oncogenic kinases. As a substantial number of multiple myeloma cell lines and primary samples were found to express AMPK-related protein kinase 5(ARK5), a member of the AMPK family associated with tumor growth and invasion, we examined whether dual inhibition of CDK4 and ARK5 kinases using ON123300 results in a better therapeutic outcome. Treatment of multiple myeloma cell lines and primary samples with ON123300 in vitro resulted in rapid induction of cell-cycle arrest followed by apoptosis. ON123300-mediated ARK5 inhibition or ARK5-specific siRNAs resulted in the inhibition of the mTOR/S6K pathway and upregulation of the AMPK kinase cascade. AMPK upregulation resulted in increased SIRT1 levels and destabilization of steady-state MYC protein. Furthermore, ON123300 was very effective in inhibiting tumor growth in mouse xenograft assays. In addition, multiple myeloma cells sensitive to ON123300 were found to have a unique genomic signature that can guide the clinical development of ON123300. Our study provides preclinical evidence that ON123300 is unique in simultaneously inhibiting key oncogenic pathways in multiple myeloma and supports further development of ARK5 inhibition as a therapeutic approach in multiple myeloma