Abstract 4993: Label-free metabolic imaging of T cell response to the tumor microenvironment

Abstract

Abstract Numerous factors including low pH, low glucose, and high lactic acid levels in tumors create an immunosuppressive tumor microenvironment. This immunosuppression is partially due to the metabolic requirements of T cell activation, which are difficult to meet in the tumor microenvironment. The goal of this study is to use label-free optical metabolic imaging (OMI) of the metabolic co-factors NAD(P)H and FAD in CD3-conjugated (activated) and control T cells to monitor metabolic changes of T cells within tumor micro environmental conditions. Single-cell measurements include the optical redox ratio (fluorescence intensity of NAD(P)H divided by FAD), which measures the oxidation-reduction state of the cell, and the mean fluorescence lifetimes (τm) of NAD(P)H and FAD, which measure enzyme-binding activity in the cell. Jurkat T cells were conjugated with CD3 antibody over a two-day incubation period to achieve activation. Control and CD3-conjugated cells were exposed to environments with increasing concentrations of lactic acid (0-20 mM), hydrochloric acid (HCl; pH 6.4- 7.4), and glucose deprivation to mimic the tumor microenvironment. OMI monitored metabolic changes on a single-cell level without confounding factors associated with cell surface marker labeling. Treatment with lactic acid resulted in a significant dose-dependent decrease in the FAD τm and optical redox ratio (p<0.005). Along with changes in metabolism, OMI also resolved impaired mobility of the T cells with increasing lactic acid concentrations. Lactic acid affects both pH and metabolism, so cells were separately exposed to different concentrations of HCl (pH 6.4 - pH 7.4) to monitor pH effects alone. In control cells, the optical redox ratio decreased with decreasing pH (p<0.005). Conversely, in the presence of CD3, the optical redox ratio increased with decreasing pH (p<0.005). CD3 conjugation in glucose deprived environments also resulted in increases in both optical redox ratio and NAD(P)H τm. Cellular viability assays indicate apoptosis only in the lowest pH environment (pH 6.4, p<0.005). These studies indicate that OMI is a promising label-free approach to investigate the effects of the tumor microenvironment on T cell metabolism, activation, and drug response. Citation Format: Ava VanDommelen, Amani Gillette, Manish S. Patankar, Melissa Skala. Label-free metabolic imaging of T cell response to the tumor microenvironment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4993.