Abstract Poly ADP-ribose polymerase (PARP) is an NAD-dependent DNA repair enzyme. Inhibition of PARP in BRCA-mutant cancers has been a clinically approved therapy for several years. While the PARP inhibitor, olaparib, has been approved for the treatment of BRCA-mutant breast and ovarian cancers, it has not been used in other cancers because its mechanism of action relies on synthetic lethality. We have found that olaparib may also have use in other cancers due to its ability to modulate the immune system directly. Specifically, we have found that olaparib is able to increase the proliferation, activation, and cytotoxicity of T cells in vitro and reduce tumor growth in vivo at low doses. Recent papers have proposed that olaparib is able to enhance T cell function through the stimulator of interferon genes (STING) pathway by enhancing antigen presenting cell activity and resulting in more granzyme B expression and enhanced tumor killing. Using purified T cells from both wild type and STING knockout mice, we have shown that olaparib can also cause an increase in granzyme B and perforin expression, an increase in T cell proliferation, and enhanced tumor cell killing in a STING-independent manner. Because PARP is an NAD-consuming enzyme, we examined the role of NAD+ in the activation T cells and the production of granzyme B and perforin. By using an inhibitor of NAD+ recycling, FK866, we show that T cells have reduced NAD+ levels, reduced granzyme B and perforin expression, and even fail to activate in the presence of CD3 and CD28 when NAD+ is limited; however, adding olaparib to these cells rescues the effects of FK866 and allows proliferation and granzyme B production. In vivo we can see that even at olaparib doses that are significantly below those which induce synthetic lethality of tumor cells, olaparib is able to inhibit tumor growth in models of both colorectal and breast cancer. This suggests that olaparib can modulate the activity of T cells directly and has implications for enhancing the anti-tumor activity of these cells both alone and in combination with other forms of immunotherapy. Citation Format: Matthew J. Dean, Liqin Zheng, Phaethon Philbrook, Augusto C. Ochoa, A. Hamid Boulares. Poly ADP-ribose polymerase inhibition enhances T cell cytotoxicity and anti-tumor function by altering NAD+ levels [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 2081.