Triple MAPK inhibition salvaged a relapsed post-BCMA CAR-T cell therapy multiple myeloma patient with a BRAF V600E subclonal mutation

Abstract

Background: Multiple Myeloma (MM) is a progressive plasma cell neoplasm characterized by heterogeneous clonal expansion. Despite promising response rates achieved with anti-BCMA CAR-T cell therapy, patients may still relapse and there are currently no clear therapeutic options in post-CAR-T settings. In this report, we present a case of a post-BCMA CAR-T relapsed/refractory (RR) MM patient with skin extramedullary disease (EMD) in which a novel MAPK inhibition combinatorial strategy was implemented based on next-generation sequencing and in vitro experiments.

Case presentation: A 61-year-old male with penta-refractory MM penta- (IgA lambda), ISS stage 3 with hyperdiploidy, gain of 1q21 and del13 was treated with anti-BCMA CAR-T cell therapy, achieving a best response of VGPR. He progressed after 6 months and was salvaged for a short period with autologous stem cell transplantation. Eventually, he progressed with extramedullary disease manifested as subcutaneous nodules. Based on whole-exome sequencing, we identified a BRAF (V600E) dominant subclone in both bone marrow and cutaneous plasmacytoma. Following in vitro experiments, and according to our previous studies, we implemented a triple MAPK inhibition strategy under which the patient achieved a very good partial response for 110 days, which allowed to bridge him to subsequent clinical trials and eventually achieve a stringent complete response (sCR).

Conclusion: Here, we show the applicability, effectiveness, and tolerability the triple MAPK inhibition strategy in the context of post-BCMA CAR-T failure in specific subset of patients. The triple therapy could bridge our hospice bound RRMM patient with BRAF (V600E) to further therapeutic options where sCR was achieved. We will further evaluate triple MAPK inhibition in patients with BRAF V600E in a precision medicine clinical trial launching soon.

Keywords: BCMA CAR-T relapse; BRAF (V600E); Clonality; MAPK inhibition; Multiple myeloma; RNA-seq; Whole-exome sequencing.

Fig. 1

Timeline depicting the patient’s clonal evolution, treatment regimens, and responses since diagnosis. Timeline is represented in days since the establishment of MM diagnosis. Treatment regimens are indicated with black arrows descending toward the upper side of the timeline. Diagnostic whole-exome sequencing (WES), RNA-sequencing (RNA-seq) or targeted panel are indicated as blue bars descending from the timeline. Red arrows indicate period of disease progression (PD), and different shades of green arrows indicate very good partial response (VGPR), complete response (CR), and stringent complete response (sCR). Lower part of the graph depicts the patient’s lambda LC levels (mg/L) since the beginning of the triple therapy regimen (regorafenib, dabrafenib and encorafenib) until sCR achieved after antibody trial 2. ASCT: Autologous stem cell transplantation; VCd: Velcade (Bortezomib), cyclophosphamide and dexamethasone. VRd: Velcade (Bortezomib), lenalidomide, and dexamethasone; Vd: Velcade (Bortezomib) and dexamethasone; anti-BCMA CAR-T: anti-B cell maturation antigen chimeric antigen receptor T cells; VDCEP: Velcade (Bortezomib), dexamethasone, cyclophosphamide, etoposide, and cisplatin; BCNU: Carmustine

Fig. 2

Triple MAPK inhibition effectively reduces phosphorylated ERK in BRAF (V600E) CD138⁺ plasma cells A WB of magnetic bead selected CD138⁺ plasma cells from RRMM patient’s BMA after 48 h in vitro treatment with encorafenib (ENC; 50 nM) and binimetinib (BIN; 250 nM), regorafenib alone (REG; 1μΜ), or combination of the three drugs. B Travera analysis on RRMM patient CD138 + cells showing sensitivity to trametinib (TRAM) in combination with dabrafenib (DAB) and regorafinib (REG) at varying concentrations. C Relative pERK protein expression after quantification and normalization to actin

Fig. 3

Temporal evolution and trajectory of MAPK alterations. A Clusters of subclonal mutations sampled over time with the mutational VAF represented as a percentage on the y axis show the trajectory of the subclone harboring the BRAF V600E mutation over time in response to treatment and the later emergence of a subclonal KRAS Q61R mutation after the end of triple MAPK inhibition. B A reconstructed phylogenetic tree of subclones across all time points shows that the BRAF V600E is subclonal and ancestral to the clone that gives rise to a KRAS Q61R mutation. C RNA expression shows an increase in MAPK pathway activation, measured as a combined z-score, over time in response to treatment with triple MAPK inhibition therapy

Fig. 4

CD138⁺ BMA plasma cells shift their dependency to PI3K/AKT pathway with increased sensitivity to copanlisib A WB of magnetic bead selected CD138 + plasma cells from RRMM patient’s BMA after 48 h in vitro treatment with with encorafenib (ENC; 50 nM) and binimetinib (BIN; 250 nM), regorafenib alone (REG; 1μΜ), combination of the three drugs or copanlisib alone (25 nM). B Rlative protein expression of pERK, pS6, pAKT and BRAF (V600E) after quantification and normalization to actin C CD138 + viability measurement after 48 h in vitro treatment with with encorafenib (ENC; 50 nM) and binimetinib (BIN; 250 nM), regorafenib alone (REG; 1 μΜ), combination of the three drugs or copanlisib alone (25 nM). D RNA expression shows elevated PI3K/Akt pathway activation, measured as a combined z-score, throughout the course of treatment with triple MAPK inhibition therapy