Vander Mause, E. R. | bioRxiv Science

bioRxiv2021

Low-affinity CAR T cells exhibit reduced trogocytosis, preventing fratricide and antigen-negative tumor escape while preserving anti-tumor activity

Chimeric antigen receptor (CAR) T cells using the high-affinity CD19 binding domain FMC63 are an effective treatment for patients with relapsed and aggressive B cell lymphoma. However, antigen loss and poor CAR T cell persistence remain common causes for relapse in these patients. Using primary patient samples, we now show that FMC63-based CAR T cells confer rapid antigen loss in all major tumor types currently approved for treatment with CD19 CAR T cells via trogocytosis, the stripping of antigen from tumor cells by CAR T cells. We show that CAR T cell-mediated trogocytosis can be dramatically reduced across a wide range of B cell malignancies by replacing FMC63 with a low affinity CD19 antibody. This reduction in trogocytosis does not alter the direct anti-tumor activity of CD19 CAR T cells but prevents the emergence of antigen-negative tumor cells and significantly increases CAR T cell viability by reducing fratricide of CD19 CAR T cells following trogocytosis. TEASERA reduction in CAR affinity does not affect tumor killing but prolongs T cell persistence and prevents antigen-negative tumor escape.

bioengineering↗

bioRxiv2021

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 low affinity variants of the fully human CD229-specific antibody 2D3. Parental 2D3 originally 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, engineered to also express CJUN to increase CAR T cell expansion, maintain the parental antibodys anti-tumor activity but lack its targeting of healthy T cells in vitro and in vivo. In addition, we found that low affinity 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 and other lymphoid malignancies. One sentence summaryRational T cell engineering yields low affinity CD229 CAR T cells overexpressing CJUN, which maintain the parental cells anti-tumor activity but eliminate killing of healthy T cells, increasing CAR T cell expansion, and decreasing trogocytosis.

bioengineering↗