CAR-modified marrow infiltrating lymphocytes efficiently target malignant plasma cells with very low antigen density.

B cell maturation antigen (BCMA) is the primary target for chimeric antigen receptor (CAR)-T cell therapies and other immunotherapies in multiple myeloma (MM). Frequent relapses and the need for sequential treatments underscore the importance of alternative targets. We previously developed a dual-antigen-specific CAR recognizing both BCMA and transmembrane activator and CAML interactor (TACI). Here, we efficiently expressed this CAR in patient-derived marrow-infiltrating lymphocytes (MILs) to investigate MM cell killing via both endogenous T cell receptor (TCR) specificities and the CAR. While the patients' MIL TCRs did not recognize or kill autologous malignant plasma cells (PCs), the CAR consistently mediated efficient PC killing even at low or undetectable antigen levels by flow cytometry. Quantification of BCMA and TACI molecules on target cells by direct stochastic optical reconstruction microscopy (dSTORM) revealed that both BCMA and TACI were significantly underestimated by flow cytometry. Analysis by dSTORM allowed us to delineate the dual-antigen sensitivity of the CAR, indicating that CAR-MILs were polyfunctional and killed targets at very low antigen densities. Our findings support the further exploration of TACI as target in MM and underline the value of super-resolution microscopy in assessing antigen density thresholds for CAR-T cells.