Abstract 515: Heterogeneity of PD-1hi CD8 T cells associates with response to PD-1 blockade in hepatocellular carcinoma
Authors: Magen Assaf , Hamon Pauline , Humblin Etienne , Schanoski Alessandra , Kim Joel , Berichel Jessica Le , Kenigsberg Ephraim , Schwartz Myron , Marron Thomas , Kamphorst Alice , Merad Miriam 
Published: July 2021

Abstract
Blockade of the PD-1 pathway is a therapeutic strategy to reinvigorate T cell responses against tumors, and when combined with other biologic therapies in the first line setting this achieves significant clinical response in about 25% of hepatocellular carcinoma (HCC) patients. We hypothesize that phenotypic diversity of tumor infiltrating T cells can explain, at least partially, the disparate clinical responses to immunotherapy. Here, we analyze the molecular diversity of T cells in tumor, adjacent tissue and tumor-draining lymph node (dLN) by single-cell RNA sequencing of tissue from 23 patients with early stage HCC treated by neoadjuvant PD-1 blockade (NCT03916627; final data to be submitted to AACR 2021 as late breaking abstract). We identify distinct subsets of PD-1hi CD8 T cells with varying degrees of exhaustion and effector gene programs. Compared to parallel analysis of untreated HCC tumors, we observed that PD-1 blockade resulted in expansion of PD-1hi CD8 T cells in the tumor, regardless of clinical response. PD-1hi CD8 T cell subsets were highly clonal and enriched in the tumor compared to adjacent tissue, suggesting specificity to tumor antigens. Remarkably, we find an association between a specific transcriptomic phenotype within PD-1hi CD8 T cells and response to PD-1 blockade. Using T cell receptor (TCR) sequencing to study the differentiation patterns between T cell states, we found that expanded clonotypes present among PD-1hi CD8 T cells were also found in CD8 effector cells; these data identify characteristic clonally related T cell populations that are enriched in clinical responders. Furthermore, we find the dLNs harbor clonotypes of PD-1hi CD8 T cells expanded in the tumor. These clonally expanded CD8 T cells in dLN display features of activation and exhaustion, suggesting a continuous role of dLN in anti-tumor responses. These results will be corroborated with 8 additional patient samples in which we will further analyze the molecular characteristics of tumor-specific CD8 T cells in the TME and dLN. Furthermore, our ongoing sequencing of pre- and post-treatment lesions will enable measurement of CD8 T cell clonal expansion and molecular programs during response or resistance to PD-1 blockade. These results should help unravel the contribution of dLN T cells to response to immunotherapy as well as identify T cell molecular programs at baseline that correlate with response.
Citation Format: Assaf Magen, Pauline Hamon, Etienne Humblin, Alessandra Schanoski, Joel Kim, Jessica Le Berichel, Ephraim Kenigsberg, Myron Schwartz, Thomas Marron, Alice Kamphorst, Miriam Merad. Heterogeneity of PD-1hi CD8 T cells associates with response to PD-1 blockade in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 515.