Jones et al. investigated off-target binding of the 1G4 TCR in CD8+ T cells using an MHC-I ligandome screening approach. The native TCR exhibited less off-target binding than the high-affinity therapeutic version. TCR+CD4+ T cells reacted to a subpopulation of these hits, suggesting that some off-target hits were CD8+ T cell-independent. Blockade/KO of CD8α reduced the off-target hits to match the CD4+ T cell pool, which limited off-target killing. CD8α-/- cells maintained on-target cytotoxicity at high E:T ratios and controlled tumors better than CD4+ T cells, albeit slightly less effectively than CD8α+/+ cells. Analogous results were observed with the MART1-reactive DMF5 TCR.
Contributed by Morgan Janes
ABSTRACT: Engineered T cells have shown efficacy in cancer treatment. However, the promiscuity of TCR-engineered T cells may result in recognition of off-target epitopes, causing severe toxicities. A genetic screen of >3,000 proteomic epitopes in the MHCI ligandome uncovered off-target peptides for both, native and affinity-enhanced 1G4 TCR, which target cancer antigen NY-ESO-1/A02-expressing cells. We validated off-target peptides derived from the human proteome recognized by both TCRs, showing that the affinity-enhanced TCR has more off-targets. Multiple off-target epitopes were reactive only in CD8 T cells, not in CD4 T cells. We identified a previously undescribed class of CD8 receptor-dependent off-targets. CD8α-negative cells (CD8α-/-) 1G4 T cells had fewer off-target reactivities, enhancing on-target specificity in vitro and in vivo. We corroborated our findings with the DMF5 TCR, targeting MART1/A02. This research advances our understanding of the distinct roles that CD4 and CD8 proteins play in T cell antigen recognition, potentially leading to more specific, effective, and safer engineered T cell therapies.
Author Info: (1) Pharmacology Program, Weill Cornell Medicine, New York, NY 10021, USA; Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (2) Phys
Author Info: (1) Pharmacology Program, Weill Cornell Medicine, New York, NY 10021, USA; Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (2) Physiology Biophysics and Systems Biology Program, Weill Cornell Medicine, New York, NY 10021, USA; Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (3) Tri-Institutional MD-PhD Program, Sloan Kettering Institute, Weill Cornell Medicine and Rockefeller University, New York, NY 10065, USA; Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (4) Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (5) Pharmacology Program, Weill Cornell Medicine, New York, NY 10021, USA; Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Electronic address: scheinbd@mskcc.org.
