MM-378: Inhibitory and Stimulatory Interplay Between Myeloma Tumor and T Cells

Context: T cells engineered to express chimeric antigen receptors (CAR-Ts) offer hope for improved outcomes in multiple myeloma. Patients relapse despite 80% or better response rates from CAR-Ts in clinical trials. Improvements in CAR design may come from a better understanding of the interplay between T cells and the tumor microenvironment. Objective: To describe this interplay, we deeply characterized the surfaceome of the ANBL-6 cell line shown by us to best represent the transcriptome of patients newly diagnosed with multiple myeloma. Design: ANBL-6 cells were cultured in RPMI Complete with 2 ng/mL IL-6 and were stained with antibodies directed toward 28 unique cell-surface antigens known to mark myeloma cells, as well as those known to have effects on T cells that are stimulatory, inhibitory, or both. Flow cytometry was employed to assess expression of cell-surface antigens on ANBL-6 cells incubated for 24 hours in and out of the presence of IFNγ, which is expressed by activated T cells, at 100 U/mL. Results: Canonical myeloma markers CD38 and CD138 were expressed. BCMA, BAFF, and the receptor to BAFF (BAFF-R) were not. CD19, CD44, CD56, CD72, CD307, CS1/SLAMF7, GPRC5D, and TACI were expressed. CD19, CD72, CD56, and CS1/SLAMF7 increased with IFNγ. TACI decreased. Stimulatory 4-1BB-L, CD48, CD86, ICOS-L, and OX40-L were present. 4-1BB-L increased with IFNγ. The inhibitory cell-surface antigens CD112, CEACAM, FasL, PD-L1, and PD-L2 were present. CEACAM, FasL, and PD-L1 all increased with IFNγ. Stimulatory and inhibitory CD155 and CD270 were both present and did not change with IFNγ. Inhibitory CD200 and stimulatory CD70 and CD80 were not expressed. Conclusions: ANBL-6 expressed five known T-cell stimulatory antigens and five known inhibitory antigens. One of five stimulatory antigens increased with IFNγ. Three of five inhibitory proteins increased with IFNγ. Thus, stimulated CAR-Ts may cause their own inhibition. These in vitro data in a single cell line offer insight into myeloma tumor dynamics but may not fully reflect in vivo tumor behavior. Characterization of multiple cell lines and primary tumor samples in vitro and in vivo may improve CAR design ranging from having better suited costimulatory domains to switching receptors that transform inhibitory ligands into costimulatory signals.