Abstract 518: Hypoxia and high interstitial pressure present within the tumor microenvironment can promote T cell suppression and exhaustion

Abstract

Abstract Introduction: The tumor microenvironment is thought to be hostile to the optimal functionality of immune cells. In addition to hypoxia, tumors have a higher interstitial pressure than normal tissue. We examined the impact of hypoxia and increased pressure on human T cell function in vitro. We hypothesize that hypoxia and high pressure induced a more profound and durable exhaustion of activated T cells. Methods: We cultivated human healthy PBMCs stimulated with phytohemagglutinin (PHA) or CD3/CD28 beads under tumor hypoxic conditions (1%O2, 2PSI) and compared it to normoxic in vitro culture conditions (20% O2, 0 PSI). We utilized the AVATAR system (Xcellbio), a novel bioreactor with highly customable cultivation conditions that allows for the modulation of oxygen and pressure. Functionality and phenotypic changes induced by different cultivation conditions were assessed by flow cytometry and cytokine release assays. Results: Upon stimulation of PBMCs for 3 days, we observed that hypoxia markedly reduced IFNγ and TNFα cytokine release. Additionally, hypoxia induced an overall decrease in surface expression of the activation and metabolic markers CD71 (transferrin) and CD98 (neutral amino acids transporter), while promoting an increased expression of the inhibitory receptor PD1. This was true for both CD4+ and CD8+ T cells. Following this initial 3-day “priming” stimulation, cells spent 2-days in a “resting period” and underwent a second stimulation under a different condition (cells under normoxia were cultured under hypoxia and vice versa). We found that cells initially primed under hypoxic conditions maintained higher levels of PD1 expression after the second stimulation, regardless of whether the latter condition was normoxic or hypoxic. Conclusions: Taken together these observations suggest that T cell priming in hypoxic and high pressure conditions present in tumors induce an irreversible state of T cell exhaustion and point out a dominant effect of hypoxia over normoxia. Our data suggest that environmental culture conditions greatly affect T cell responses and should be taken into account when studying T cell functionality, especially in the investigation of novel therapeutic agents targeting the immune system. Citation Format: Chiara Rancan, Rachel Marusinec, James Lim, Lawrence Fong. Hypoxia and high interstitial pressure present within the tumor microenvironment can promote T cell suppression and exhaustion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 518.