Fast off-rate CD229 chimeric antigen receptor T cells efficiently target multiple myeloma, spare T cells, and exhibit reduced trogocytosis

T cells expressing chimeric antigen receptors have shown remarkable therapeutic activity against different types of cancer. However, their wider use has been hampered by the potential for life-threatening toxicities due to the unintended targeting of healthy cells expressing low levels of the targeted antigen. We have now developed an affinity-tuning approach for the generation of minimally modified, low-affinity antibody variants derived from existing high-affinity antibodies. Using this approach, we engineered a low affinity variant of the fully human CD229-specific antibody 2D3. Parental 2D3 efficiently targeted multiple myeloma cells but also healthy T cells expressing low levels of CD229. We demonstrate that CAR T cells based on a low affinity variant of 2D3 maintain the parental antibody's anti-tumor activity, but lack its targeting of healthy T cells. In addition, variant CD229 CAR T cells show reduced trogocytosis potentially augmenting CAR T cell persistence. The fast off-rate CAR produced using our affinity tuning approach eliminates a key liability of CD229 CAR T cells and paves the way for the effective and safe treatment of patients with multiple myeloma. ### Competing Interest Statement ERV, TL, and DA are inventors on provisional patent application 63/285843 describing low-affinity CD229 antibodies and CAR T cells. SVR, DA, and TL are inventors on PCT application US2017/42840 'Antibodies and CAR T Cells for the Treatment of Multiple Myeloma' describing the therapeutic use of CD229 CAR T cells for the treatment of multiple myeloma.