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. 2022 Jan 28;10(1):e003826. doi: 10.1136/jitc-2021-003826

Figure 6.

Figure 6

MSLN Tmod system can be extended to autologous T cells. (A) MSLN SS1 CAR Tmod construct kills MSLN(+) A*02(−) tumor HeLa target cells but no longer blocks in the presence of autologous A*02 as a result of cis-binding. B2M knockout (KO) by CRISPR restores blocker availability as demonstrated by binding to A*02 tetramer. (B) B2M KO by CRISPR in A*02(+) T cell donors restores blocking on MSLN(+)A*02(+) “normal” HeLa target cells similar to A*02(−) T cell donors. E:T=1.2:1 (C) Representative co-culture images at 48 hours for figure 6B. (D) Similar to SS1 Tmod, CAR3 Tmod has reduced binding to A*02 tetramer in A*02(+) T cells. B2M knockdown (KD) with shRNA restores blocker availability. (E) B2M shRNA also restores blocking of cytotoxicity on MSLN(+)A*02(+) “normal” HeLa cells. (F) Interestingly, CAR3 paired with a humanized A*02 blocker retains the ability to block killing of “normal” cells in A*02(+) donor T cells, even in the absence of B2M KO or KD.