Metformin improves cancer immunotherapy by directly rescuing tumor-infiltrating CD8 T lymphocytes from hypoxia-induced immunosuppression

Abstract

BackgroundDespite their revolutionary success in cancer treatment over the last decades, immunotherapies encounter limitations in certain tumor types and patients. The efficacy of immunotherapies depends on tumor antigen-specific CD8 T-cell viability and functionality within the immunosuppressive tumor microenvironment, where oxygen levels are often low. Hypoxia can reduce CD8 T-cell fitness in several ways and CD8 T cells are mostly excluded from hypoxic tumor regions. Given the challenges to achieve durable reduction of hypoxia in the clinic, ameliorating CD8 T-cell survival and effector function in hypoxic condition could improve tumor response to immunotherapies.MethodsActivated CD8 T cells were exposed to hypoxia and metformin and analyzed by fluorescence-activated cell sorting for cell proliferation, apoptosis and phenotype. In vivo, metformin was administered to mice bearing hypoxic tumors and receiving either adoptive cell therapy with tumor-specific CD8 T cells, or immune checkpoint inhibitors; tumor growth was followed over time and CD8 T-cell infiltration, survival and localization in normoxic or hypoxic tumor regions were assessed by flow cytometry and immunofluorescence. Tumor oxygenation and hypoxia were measured by electron paramagnetic resonance and pimonidazole staining, respectively.ResultsWe found that the antidiabetic drug metformin directly improved CD8 T-cell fitness in hypoxia, both in vitro and in vivo. Metformin rescued murine and human CD8 T cells from hypoxia-induced apoptosis and increased their proliferation and cytokine production, while blunting the upregulation of programmed cell death protein 1 and lymphocyte-activation gene 3. This appeared to result from a reduced production of reactive oxygen species, due to the inhibition of mitochondrial complex I. Differently from what others reported, metformin did not reduce tumor hypoxia, but rather increased CD8 T-cell infiltration and survival in hypoxic tumor areas, and synergized with cyclophosphamide to enhance tumor response to adoptive cell therapy or immune checkpoint blockade in different tumor models.ConclusionsThis study describes a novel mechanism of action of metformin and presents a promising strategy to achieve immune rejection in hypoxic and immunosuppressive tumors, which would otherwise be resistant to immunotherapy.

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“…The drug is a type 2 diabetes treatment that has been found to diminish intratumoral accumulation of myeloid-derived suppressor cells and downregulate PD-L1 expression in tumor cells. 130 When mice with hypoxic tumors were treated with metformin and tumor-specific CD8+ T cells, Metformin was found to rescue CD8+ T cells from hypoxia-induced apoptosis and improve infiltration into hypoxic areas of the tumor without actually reducing tumor hypoxia. 130 …”

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“…Therefore, data on transcriptional and epigenetic levels are needed to determine the mechanisms underlying the regulatory effects of metformin on the AMPK/mTORC1 axis and corresponding metabolic reprogramming. Undeniably, metformin, which has been evaluated in preclinical studies, may be a metabolically activating small‐molecule drug that increases tumour immunity 172,173 . In contrast, TCR‐induced zinc finger protein 91 (ZEP91) translocation promotes assembly of the serine/threonine protein phosphatase 2A complex, which regulates mTOR signalling.…”