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B-cell maturation antigen (BCMA) is the lead antigen for chimeric antigen receptor (CAR) T-cell therapy in multiple myeloma (MM). A challenge is inter- and intra-patient heterogeneity in BCMA expression on MM cells and BCMA downmodulation under therapeutic pressure. Accordingly, there is a desire to augment and sustain BCMA expression on MM cells in patients that receive BCMA-CAR T-cell therapy. We used all-trans retinoic acid (ATRA) to augment BCMA expression on MM cells and to increase the efficacy of BCMA-CAR T cells in pre-clinical models. We show that ATRA treatment leads to an increase in BCMA transcripts by quantitative reverse transcription polymerase chain reaction and an increase in BCMA protein expression by flow cytometry in MM cell lines and primary MM cells. Analyses with super-resolution microscopy confirmed increased BCMA protein expression and revealed an even distribution of non-clustered BCMA molecules on the MM cell membrane after ATRA treatment. The enhanced BCMA expression on MM cells after ATRA treatment led to enhanced cytolysis, cytokine secretion and proliferation of BCMA-CAR T cells in vitro, and increased efficacy of BCMA-CAR T-cell therapy in a murine xenograft model of MM in vivo (NSG/MM.1S). Combination treatment of MM cells with ATRA and the γ- secretase inhibitor crenigacestat further enhanced BCMA expression and the efficacy of BCMA-CAR T-cell therapy in vitro and in vivo. Taken together, the data show that ATRA treatment leads to enhanced BCMA expression on MM cells and consecutively, enhanced reactivity of BCMA-CAR T cells. The data support the clinical evaluation of ATRA in combination with BCMA-CAR T-cell therapy and potentially, other BCMA-directed immunotherapies., EGG has been supported by Instituto de Salud Carlos III (PFIS - FI12/00189 and PI20/01792). LGRL has been a BITRECS fellow and has received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement No 754550 and from the “La Caixa” Foundation. SD has been supported by the IZKF Würzburg (Interdisziplinäres Zentrum für Klinische Forschung) and is a fellow of the Clinician Scientist Program of the Else-Kröner Forschungskolleg. The authors have been supported by the patient advocacy group 'Hilfe im Kampf gegen den Krebs e.V.', Würzburg, Germany and 'Forschung hilft' - Stiftung zur Förderung der Krebsforschung an der Universität Würzburg. Further, the authors have been supported by the European Union’s Horizon 2020 research and innovation program under grant agreements No 733297 (EURE-CART to MH, HE and SRP) and No 754658 (CARAMBA to MH, SD, SRP and HE), the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG, project number 324392634, TRR 221, subproject A03 to MH and HE), the Bavarian Center for Cancer Research (Bayerisches Zentrum für Krebsforschung, BZKF to MH, HE and SRP) and the Stifterverband für die Deutsche Wissenschaft e.V. (to KMK and MH). MS, MH and HE acknowledge funding by the German Ministry for Science and Education (BMBF, Bundesministerium für Bildung und Forschung, grant #13N15986). Further, the authors have been supported by the Innovative Medicines Initiative 2 Joint Undertaking under grant agreement No 116026 (T2EVOLVE). This Joint Undertaking receives support from the European Union's Horizon 2020 research and innovation program and EFPIA.

Supplementary Material 1

Interactions of malignant multiple myeloma (MM) plasma cells with the microenvironment control MM plasma-cell growth, survival, drug-resistance and dissemination. As microvascular density increases in the bone marrow in MM, we investigated whether bone marrow MM endothelial cells control disease progression via the junctional adhesion molecule-A (JAM-A). Membrane and cytoplasmic JAM-A levels were upregulated in MM endothelial cells in 111 patients with newly diagnosed MM and in 201 with relapsed/refractory MM compared to the levels in patients with monoclonal gammopathy of undetermined significance and healthy controls. Elevated membrane expression of JAM-A on MM endothelial cells predicted poor clinical outcome. Mechanistically, addition of recombinant JAM-A to MM endothelial cells increased angiogenesis, whereas inhibition of this adhesion molecule impaired angiogenesis and MM growth in two-dimensional and three-dimensional in vitro cell cultures and chorioallantoic membrane assays. To corroborate these findings, we treated MM-bearing mice with a JAM-A-blocking monoclonal antibody and demonstrated impaired MM progression, corresponding to decreased MM-related vascularity. These findings support the concept that JAM-A is an important mediator of MM progression through facilitating MM-associated angiogenesis. Elevated JAM-A expression on bone marrow endothelial cells is an independent prognostic factor for the survival of both patients with newly diagnosed MM and those with relapsed/refractory MM. Blocking JAM-A restricts angiogenesis in vitro, in utero and in vivo and represents a suitable druggable molecule to halt neo-angiogenesis and MM progression.