Determining mechanisms of resistance to αPD-1/PD-L1 immune-checkpoint immunotherapy is key to developing new treatment strategies. Cancer-associated fibroblasts (CAF) have many tumor-promoting functions and promote immune evasion through multiple mechanisms, but as yet, no CAF-specific inhibitors are clinically available. Here we generated CAF-rich murine tumor models (TC1, MC38, and 4T1) to investigate how CAFs influence the immune microenvironment and affect response to different immunotherapy modalities [anticancer vaccination, TC1 (HPV E7 DNA vaccine), αPD-1, and MC38] and found that CAFs broadly suppressed response by specifically excluding CD8+ T cells from tumors (not CD4+ T cells or macrophages); CD8+ T-cell exclusion was similarly present in CAF-rich human tumors. RNA sequencing of CD8+ T cells from CAF-rich murine tumors and immunochemistry analysis of human tumors identified significant upregulation of CTLA-4 in the absence of other exhaustion markers; inhibiting CTLA-4 with a nondepleting antibody overcame the CD8+ T-cell exclusion effect without affecting Tregs. We then examined the potential for CAF targeting, focusing on the ROS-producing enzyme NOX4, which is upregulated by CAF in many human cancers, and compared this with TGFβ1 inhibition, a key regulator of the CAF phenotype. siRNA knockdown or pharmacologic inhibition [GKT137831 (Setanaxib)] of NOX4 "normalized" CAF to a quiescent phenotype and promoted intratumoral CD8+ T-cell infiltration, overcoming the exclusion effect; TGFβ1 inhibition could prevent, but not reverse, CAF differentiation. Finally, NOX4 inhibition restored immunotherapy response in CAF-rich tumors. These findings demonstrate that CAF-mediated immunotherapy resistance can be effectively overcome through NOX4 inhibition and could improve outcome in a broad range of cancers. SIGNIFICANCE: NOX4 is critical for maintaining the immune-suppressive CAF phenotype in tumors. Pharmacologic inhibition of NOX4 potentiates immunotherapy by overcoming CAF-mediated CD8+ T-cell exclusion. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/9/1846/F1.large.jpg.See related commentary by Hayward, p. 1799.

中文翻译：

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通过克服肿瘤相关的成纤维细胞介导的CD8 T细胞排斥，NOX4抑制增强了免疫治疗。

确定对αPD-1/ PD-L1免疫检查点免疫治疗的耐药性机制是开发新治疗策略的关键。癌症相关的成纤维细胞（CAF）具有许多促进肿瘤的功能，并通过多种机制促进免疫逃逸，但目前尚无临床上可利用的CAF特异性抑制剂。在这里，我们生成了富含CAF的鼠肿瘤模型（TC1，MC38和4T1），以研究CAF如何影响免疫微环境并影响对不同免疫疗法的反应[抗癌疫苗接种，TC1（HPV E7 DNA疫苗），αPD-1和MC38 ]，发现CAF通过特异性地从肿瘤中排除CD8 + T细胞（不是CD4 + T细胞或巨噬细胞）来广泛抑制反应。富含CAF的人类肿瘤中也相似地存在CD8 + T细胞排斥。来自富含CAF的鼠类肿瘤的CD8 + T细胞的RNA测序和人类肿瘤的免疫化学分析表明，在没有其他衰竭标记的情况下，CTLA-4明显上调；用非消耗性抗体抑制CTLA-4的作用克服了CD8 + T细胞的排斥作用，而不影响Treg。然后，我们研究了CAF靶向的潜力，重点是在许多人类癌症中被CAF上调的产生ROS的酶NOX4，并将其与CAF表型的关键调控因子TGFβ1抑制进行了比较。siRNA抑制或NOX4“标准化” CAF的药理抑制[GKT137831（Setanaxib）]到静止的表型，并促进肿瘤内CD8 + T细胞浸润，克服了排斥作用；抑制TGFβ1可以预防CAF分化，但不能逆转。最后，NOX4抑制恢复富含CAF的肿瘤的免疫治疗反应。这些发现表明，通过NOX4抑制可以有效克服CAF介导的免疫治疗耐药性，并可以改善多种癌症的预后。意义：NOX4对于维持肿瘤中免疫抑制CAF表型至关重要。通过克服CAF介导的CD8 + T细胞排斥，NOX4的药理抑制作用可增强免疫疗法。图形摘要：http://cancerres.aacrjournals.org/content/canres/80/9/1846/F1.large.jpg。请参阅Hayward的相关评论，第1页。1799年。NOX4对于维持肿瘤中免疫抑制CAF表型至关重要。通过克服CAF介导的CD8 + T细胞排斥，NOX4的药理抑制作用可增强免疫疗法。图形摘要：http://cancerres.aacrjournals.org/content/canres/80/9/1846/F1.large.jpg。请参阅Hayward的相关评论，第1页。1799年。NOX4对于维持肿瘤中免疫抑制CAF表型至关重要。通过克服CAF介导的CD8 + T细胞排斥，NOX4的药理抑制作用可增强免疫疗法。图形摘要：http://cancerres.aacrjournals.org/content/canres/80/9/1846/F1.large.jpg。请参阅Hayward的相关评论，第1页。1799年。