Abstract 1534: Targeting oxygen consumption with metformin and phenformin have differential effects on immune cells in the tumor microenvironment

Abstract Immune checkpoint blockade (ICB) has yielded durable clinical responses which led to its FDA approval as a first-line therapy for melanoma and other malignancies. However, only ∼20-40% of patients show benefit when used as monotherapies. This is mainly due to inherent or acquired resistance to these therapies within the tumor microenvironment (TME). There is increasing evidence in pre-clinical studies that tumor metabolism (e.g., glycolysis and oxygen consumption (OXPHOS)) presents a barrier to anti-tumor immunity. High levels of OXPHOS in tumor cells lead to tumor hypoxia. Hypoxia causes altered tumor physiology, increased immunosuppression, and resistance to several therapies such as ICB. We propose that blocking tumor oxygen consumption using drugs that target the mitochondrial complex I (metformin and phenformin) will enhance the efficacy of immunotherapies such as ICB in preclinical models. In this study, we examined how low oxygen levels (hypoxia) influence T cell effector function such as cytotoxic activity and cytokine production in vitro. We co-cultured mouse B16 melanoma cells in the presence of tumor specific T cells and incubated at 37ºC in either normoxia (∼21% O2) or hypoxia (3% O2) and assessed killing and intracellular cytokine production 24 - 48 hours later. We show that the ability of tumor antigen-specific T cells to kill tumor cells or make effector cytokine such as IFNγ and TNFα is significantly reduced in hypoxia settings. We also examined the effects of metformin and phenformin on T cell priming and activation. When T cells were activated in the presence of each drug in vitro, there was enhanced expression of T cell activation markers (CD25, Granzyme B) as well as an increase in their differentiation into central memory T cells. We treated mice bearing melanoma and breast cancers with metformin and phenformin and tested whether these drugs could delay tumor growth and improve overall survival in vivo as a monotherapy or in combination with anti-PD1. While both drugs can significantly delay tumor growth as monotherapies, only metformin showed improved anti-tumor efficacy when combined with anti-PD-1 in vivo. We performed flow cytometry on mice bearing melanoma and breast cancers treated with metformin and phenformin in vivo. We found that these drugs have differential effects on the immune system in the tumors and periphery in the two cancer models. These studies show that while both drugs target tumor oxygen consumption (via inhibition of complex I), and can delay tumor growth in vivo, they have differential effects on the TME and will need further considerations when combining with ICBs and other immunotherapies. In addition, these studies will advance our understanding of how tumor metabolism impacts anti-tumor immune responses and could lead to better design of combination therapies for cancer patients treated with diabetes management drugs. Citation Format: Sadna Budhu, Anais Assouvie, Juan Zurita, Mamadou A. Bah, Svena Verma, Mehdi Benzaoui, Inna Serganova, Jason Koutcher, Vladimir Ponomarev, Jedd D. Wolchok, Taha Merghoub. Targeting oxygen consumption with metformin and phenformin have differential effects on immune cells in the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1534.