Abstract LB217: Ascorbic acid rescues antitumor immune response by reversing hypoxia-induced CD8+T cell dysfunction
Authors: Jafarzadeh Nazli , Gaur Pankaj , Sarhan Dareen , Shobaki Nour , Traboulsi Wael , Ramlaoui Zainab , Verma Vivek , Gupta Seema , Khleif Samir N. 
Published: June 2022

Abstract
Growing evidence suggests that hypoxia is one of the most important phenomena of solid tumors that supports tumor progression as well as therapeutic resistance. When T cells infiltrate the tumor microenvironment (TME), they encounter hypoxia and, in this condition, cells upregulate an evolutionary conserved Hypoxia-inducible factor 1 (HIF-1) transcription factor to adapt to the hypoxic stress. Hypoxia has been reported to alter specific aspects of CD8+ T cell biology through various mechanisms but in large part by the HIF1-α, leading to immune suppression. Therefore, identification and development of drugs that target hypoxia is critical to augment immune cells' function and enhance the anti-tumor response of immunotherapy. Over the past decades, several cellular and pre-clinical studies have shown that ascorbic acid (AA) possesses anti-tumor properties and acts as an adjuvant to various cancer therapies. AA, as a proven redox potential modulator, has proved to also be effective in targeting oxygen sensing regulator (reducing Hif1-α levels) and epigenetic reprogramming (increasing ten-eleven translocation (TET) methylcytosine dioxygenase enzymatic activity) in cancer cells. Most recently, immunomodulating potential of AA has been recognized as it enhanced the efficacy of checkpoint blockade by increasing immune cell infiltration within the TME. However, the exact mechanisms by which AA exerts immunotherapeutic effects in cancer have not been fully understood. Here, we hypothesized that AA exerts its immunotherapeutic effects by targeting tumor-induced hypoxic stress in CD8 T cells. Indeed, we found that AA induces a robust anti-tumor response by decreasing tumor volume and increasing mice survival. Immune profiling in the TME revealed that AA increases the infiltration of CD8+ T cells with enhanced cytotoxic cytokine production and proliferation. To further evaluate the underlying mechanism, we challenged CD8+ T cells with a cognate antigen ex vivo under the hypoxic/normoxic conditions. We found that hypoxia negatively affected the effector functions and proliferation of CD8 T cells. Interestingly, AA could reverse these effects, bringing the function and proliferation of cells back to the normoxic level by increasing the expression of Granzyme B, Interferon gamma and Ki-67. This reversal of hypoxia-induced adverse effects on CD8 T cells was achieved by AA-mediated decrease in the Hif1-α levels (a hypoxia mediator) and increase in TET-2 protein expression and function (characterized by increased 5-hydroxymethylcytosine (5-hmc) levels). Moreover, AA treatment could significantly enhance the metabolic fitness of CD8+ T cells (characterized by improved glycolytic ability and mitochondrial function) that were exposed to severe hypoxia. In conclusion, our data indicate that AA treatment shows enhanced anti-tumor effects via restoring cytotoxic T cell functions under hypoxic TME. These results also provide new insights into the immunomodulatory effects of AA via targeting hypoxia in CD8 T cells. Given that hypoxia in the TME is a barrier for generation of effective immune response, our results suggest that AA treatment is an effective strategy against cancer and may further enhance the anti-tumor effects of immunotherapeutic agents.
Citation Format: Nazli Jafarzadeh, Pankaj Gaur, Dareen Sarhan, Nour Shobaki, Wael Traboulsi, Zainab Ramlaoui, Vivek Verma, Seema Gupta, Samir N. Khleif. Ascorbic acid rescues antitumor immune response by reversing hypoxia-induced CD8+T cell dysfunction [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB217.