Evaluating the repertoire of immune checkpoint markers expressed on peripheral and ascites CD8⁺ T cells in ovarian cancer.

Abstract

5571 Background: Understanding the immune checkpoint marker repertoire can facilitate development of therapeutic strategies to improve efficacy of immune-based therapies. We used a novel high-dimensional flow cytometry panel to determine co-expression patterns of immune checkpoint markers and effector function of CD8+ T cells from peripheral blood and ascites of patients newly diagnosed with ovarian cancer. Methods: Peripheral blood and ascites samples were collected from patients with epithelial ovarian cancer (n=8). Cells isolated from peripheral blood and ascites were used for immune profiling by multiparameter flow cytometry of 5 inhibitory receptors (PD-1, LAG-3, TIM-3, TIGIT, and BTLA) on CD8+ T cells, along with 4 functional parameters (production of each of the following: TNF-α, IFN-γ, IL-2, and upregulation of CD107a). A complementary multiplex analysis on plasma and ascites fluid was performed to quantify 14 soluble checkpoint markers. Results: The concentrations of soluble PD-1, TIM-3, LAG-3, CTLA-4, BTLA, IDO, and CD137 were increased in ascites fluid compared to plasma from patients with ovarian cancer. Ascites CD8+ T cells co-express higher levels of inhibitory receptors than peripheral CD8+ T cells. In total, CD8+ T cells in ascites retained the ability to produce effector functions at levels similar to peripheral blood. However, IFN-γ production was retained in PD-1 only expressing CD8+ T cells and decreased in CD8+ T cells co-expressing multiple receptors. Conclusions: High-dimensional flow cytometry allowed for the phenotypic and functional characterization of CD8+ T cells from ovarian cancer patients. The profile of receptor co-expression was distinct in peripheral blood compared to ascites. Collectively, our study suggests that co-expression of factors beyond PD-1 influences CD8+ T cell activity. Thus blocking PD-1 and PD-L1 alone may not be sufficient for CD8+ T cells expressing multiple inhibitory receptors.

Mechanistic overview of immune checkpoints to support the rational design of their combinations in cancer immunotherapy

“…Costimulatory receptors on T cells further activate the T cells and amplify the immune response whereas checkpoint receptors inhibit the T cells and dampen the response. Tumor microenvironment (TME) is enriched with mediators of immunosuppression, which induce the expression of checkpoint receptors on the surface of T cells and promote a dysfunctional/exhausted phenotype in T cells, which is briefly described in the following section [59][60][61][62][63][64][65].…”

“…Progressive loss of T-cell effector functions, termed 'T-cell exhaustion' is commonly seen during chronic viral infections and in cancer. The significance of T-cell exhaustion in cancer has rapidly gained importance and several studies have identified the presence of 'exhausted' T cells in TME [59][60][61][62][63][64][65].…”