Combined PD-L1 and TIM3 blockade improves expansion of fit human CD8+ antigen-specific T cells for adoptive immunotherapy

Abstract

Antigen-specific Tcell expansion exvivo followed by adoptive transfer enables targeting of a multitude of microbial and cancer antigens. However, clinical-scale Tcell expansion from rare precursors requires repeated stimulation, which may lead toTcell dysfunction and limited therapeutic potential. We useda clinically compliant protocol to expand Epstein-Barr virus (EBV) and Wilms tumor 1 (WT1) antigen-specific CD8+ Tcells, and leveraged Tcell exhaustion-associated inhibitory receptor blockade to improve Tcell expansion. Several inhibitory receptors were expressed early by exvivo-expanded antigen-specific CD8+ Tcells, including PD-1 and TIM3, with co-expression matching evidence of Tcell dysfunction as the cultures progressed. Introduction of anti-PD-L1 and anti-TIM3 blockade in combination (but not individually) to the culture led to markedly improved antigen-specific Tcell expansion without inducing Tcell dysfunction. Single-cell RNA sequencing (RNA-seq) and Tcell receptor (TCR) repertoire profiling revealed that double blockade does not impart specific transcriptional programs in Tcells or alterations in TCR repertoires. However, combined blockade may affect gene expression in a minority of clonotypes in a donor-specific fashion. We conclude that antigen-specific CD8+ Tcell manufacturing can be improved by using TIM3 and PD-L1/PD-1 axis blockade in combination. This approach is readily applicable to several adoptive immunotherapy strategies.