The tumor-promoting fibrotic stroma rich in tumor-associated fibroblasts (TAF) is drawing increased therapeutic attention. Intriguingly, a trial with the antifibrotic drug nintedanib in non–small cell lung cancer reported clinical benefits in adenocarcinoma (ADC) but not squamous cell carcinoma (SCC), even though the stroma is fibrotic in both histotypes. Likewise, we reported that nintedanib inhibited the tumor-promoting fibrotic phenotype of TAFs selectively in ADC. Here we show that tumor fibrosis is actually higher in ADC-TAFs than SCC-TAFs in vitro and patient samples. Mechanistically, the reduced fibrosis and nintedanib response of SCC-TAFs was associated with increased promoter methylation of the profibrotic TGFβ transcription factor SMAD3 compared with ADC-TAFs, which elicited a compensatory increase in TGFβ1/SMAD2 activation. Consistently, forcing global DNA demethylation of SCC-TAFs with 5-AZA rescued TGFβ1/SMAD3 activation, whereas genetic downregulation of SMAD3 in ADC-TAFs and control fibroblasts increased TGFβ1/SMAD2 activation, and reduced their fibrotic phenotype and antitumor responses to nintedanib in vitro and in vivo . Our results also support that smoking and/or the anatomic location of SCC in the proximal airways, which are more exposed to cigarette smoke particles, may prime SCC-TAFs to stronger SMAD3 epigenetic repression, because cigarette smoke condensate selectively increased SMAD3 promoter methylation. Our results unveil that the histotype-specific regulation of tumor fibrosis in lung cancer is mediated through differential SMAD3 promoter methylation in TAFs and provide new mechanistic insights on the selective poor response of SCC-TAFs to nintedanib. Moreover, our findings support that patients with ADC may be more responsive to antifibrotic drugs targeting their stromal TGFβ1/SMAD3 activation. Significance: This study implicates the selective epigenetic repression of SMAD3 in SCC-TAFs in the clinical failure of nintedanib in SCC and supports that patients with ADC may benefit from antifibrotic drugs targeting stromal TGFβ1/SMAD3.

中文翻译：

---

肿瘤相关的成纤维细胞的表观遗传SMAD3抑制损害肺鳞状细胞癌的纤维化和对抗纤维化药物Nintedanib的反应

富含肿瘤相关成纤维细胞（TAF）的促肿瘤纤维化基质正在引起越来越多的治疗关注。有趣的是，一项针对非小细胞肺癌的抗纤维化药物nintedanib的试验报告了腺癌（ADC）而非鳞状细胞癌（SCC）的临床获益，即使两种组织间质均呈纤维化。同样，我们报道了nintedanib在ADC中选择性抑制TAFs的促肿瘤纤维化表型。在这里，我们显示，在体外和患者样品中，ADC-TAF中的肿瘤纤维化实际上高于SCC-TAF。从机制上讲，与ADC-TAFs相比，SCC-TAFs的纤维化减少和nintedanib反应与促纤维化TGFβ转录因子SMAD3的启动子甲基化增加有关，这引起TGFβ1/ SMAD2激活的代偿性增加。一致地，用5-AZA强迫SCC-TAF的整体DNA去甲基化可以挽救TGFβ1/ SMAD3的活化，而ADC-TAF和对照成纤维细胞中SMAD3的基因下调增加了TGFβ1/ SMAD2的活化，并降低了其对nintedanib的纤维化表型和抗肿瘤反应。体内。我们的研究结果还支持吸烟和/或更接近香烟烟雾颗粒的近端气道中SCC的解剖位置，可能使SCC-TAF增强SMAD3表观遗传抑制力，因为香烟烟雾冷凝物选择性地增加了SMAD3启动子甲基化。我们的结果表明，肺癌中肿瘤纤维化的组织型特异性调节是通过TAF中SMAD3启动子甲基化的差异介导的，并为SCC-TAF对Nintedanib的选择性不良反应提供了新的机制。此外，我们的发现支持ADC患者可能对靶向其基质TGFβ1/ SMAD3活化的抗纤维化药物反应更强。启示：这项研究暗示SCC-TAFs中SMAD3的选择性表观遗传抑制与Sintnanib在SCC中的临床失败有关，并支持ADC患者可受益于靶向基质TGFβ1/ SMAD3的抗纤维化药物。