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Intratumoral Hypoxia Reduces IFN-γ-Mediated Immunity and MHC Class I Induction in a Preclinical Tumor Model.

ABSTRACT: Tumor hypoxia occurs because of an increased demand for oxygen by the rapidly growing tumor cells, together with reduction in the oxygen supply due to malformed and nonfunctional tumor vasculature. The effects of tumor hypoxia on radiotherapy (RT) are well known; however, recent findings suggest it may also suppress immunotherapy, although the mechanisms governing this observation remain undetermined. Our laboratory and others have shown that IFN-γ conditions the tumor milieu and is important for the efficacy of RT. Thus, we hypothesized that hypoxia could inhibit IFN-γ-mediated antitumor responses, resulting in decreased RT efficacy. This inhibition could involve the production and/or the cellular response to IFN-γ. To test this, we used murine tumor cell lines B16F0 and Colon38. We observed that hypoxia inhibited upregulation of IFN-γ-dependent MHC class I expression by tumor cells along with the gene expression of IFN-γ-dependent chemokines CXCL9 and CXCL10, essential for immune cell infiltration. Furthermore, CD8⁺ T cells, an important source of IFN-γ, which mediate effector antitumor responses, had reduced ability to proliferate and generate IFN-γ under hypoxic conditions in vitro. Interestingly, reoxygenation restored the cytokine-producing capability of these cells. Studies performed in vivo using a mouse tumor model and the hypoxia marker EF5 demonstrated that RT could reverse the hypoxia within treated tumors. This study has identified a unique mechanism of hypoxia-induced immune suppression involving the downregulation of IFN-γ production and cellular responsiveness to this essential cytokine. These results suggest that therapies that target and reduce tumor hypoxia can potentially boost antitumor immune responses.

SUBMITTER: Murthy A

PROVIDER: S-EPMC8195309 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Interferon-induced transmembrane protein (IFITM1) plays a dual role in restriction of RNA viruses and in metastatic cancer cell growth. The signal transduction events that are orchestrated by IFITM1 are not well defined. We set out to identify IFITM1 interacting proteins to begin to define its mechanism of action. Affinity purification of SBP-tagged IFITM1, coupled to SWATH-mass spectrometry, identified significantly higher confidence interacting proteins from IFN- treated cells, relative to non-treated cells. This promoted an examination of the protein synthetic machinery in order to determine whether IFITM1 can impact on ribosomal proteome from IFN- treated cells. Isogenic ifitm1 null and ifitm1-ifitm3 null-SiHa cell panels were generated using CRISPR gRNAs to define signaling events that are linked to IFN--dependent protein synthesis. Ultracentrifugation sedimentation of cell lysates from interferon gamma treated wt and ifitm1-ifitm3 null-SiHa cells was carried out to isolate ribosomal constituents. Although SWATH-mass spectrometry of the 40S, 60S, and 80S fractions demonstrated changes in selected protein content, a significant reduction in A254 (RNA) in the 80S ribosomal fractions suggested a relatively select defect in 80S ribosomal biogenesis in ifitm1-ifitm3 null cells. The localization of IFITM1 to ribosomal protein components prompted an analysis of IFN--dependent protein synthesis using pulse SILAC. STAT1 and B2M were two dominant proteins whose synthesis increased equivalently in wt or ifitm1-ifitm3 null cells. However, MHC class I molecules and ISG15 were the most highly suppressed IFN--responsive proteins in the ifitm1-ifitm3 double null cells and this was confirmed using ifitm1 single null cells. Transient depletion of IFITM1 using targeted siRNA also depleted MHC class I molecules as well as IFITM3, STAT1, B2M, and ISG15. These data have implications for the function of IFITM1 in mediating IFN--stimulated protein synthesis and associated antigen presentation during either oncogenic or anti-viral signalling.

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Intratumoral Hypoxia Reduces IFN-γ-Mediated Immunity and MHC Class I Induction in a Preclinical Tumor Model.

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