Bispecific targeting of CD20 and CD19 increases polyfunctionality of chimeric antigen receptor T-cell products in B-cell malignancies.

Abstract

Selective immune pressure contributes to relapse due to target antigen downregulation in patients treated with anti-CD19 chimeric antigen receptor (CAR) T cells. Bispecific lentiviral anti-CD20/anti-CD19 (LV20.19) CAR T cells may prevent progression/relapse due to antigen escape. Highly polyfunctional T cells within a CAR T-cell product have been associated with response in single-antigen-targeted anti-CD19 CAR T cells. The authors performed a single-cell proteomic analysis to assess polyfunctional cells in our LV20.19 CAR T-cell product. Analysis was limited to those treated at a fixed dose of 2.5 × 10⁶ cells/kg (n = 16). Unused pre-infusion CAR T cells were thawed, sorted into CD4/CD8 subsets and stimulated with K562 cells transduced to express CD19 or CD20. Single-cell production of 32 individual analytes was measured and polyfunctionality and polyfunctional strength index (PSI) were calculated. Fifteen patients had adequate leftover cells for analysis upon stimulation with CD19, and nine patients had adequate leftover cells for analysis upon stimulation with CD20. For LV20.19 CAR T cells, PSI was 866-1109 and polyfunctionality was 40-45%, which were higher than previously reported values for other CAR T-cell products. Stimulation with either CD19 or CD20 antigens resulted in similar levels of analyte activation, suggesting that this product may have efficacy in CD19- patient populations.