经典的 TGF-β/Smad 信号通路参与了 EGFR 突变型非小细胞肺癌中 PD-L1 诱导的 EGFR-TKIs 原发性耐药。

The canonical TGF-β/Smad signalling pathway is involved in PD-L1-induced primary resistance to EGFR-TKIs in EGFR-mutant non-small-cell lung cancer.

机构信息

Department of Respiratory Medicine, the First Affiliated Hospital of Soochow University, Suzhou, 215006, People's Republic of China.

Suzhou Key Laboratory for Respiratory Diseases, Suzhou, 215006, China.

出版信息

Respir Res. 2019 Jul 22;20(1):164. doi: 10.1186/s12931-019-1137-4.

BACKGROUND

Approximately 30% of patients with epidermal growth factor receptor (EGFR)-activating mutations have no response to EGFR-tyrosine kinase inhibitors (TKIs) (primary resistance). However, little is known about the molecular mechanism involved in primary resistance to EGFR-TKIs in EGFR-mutant non-small cell lung cancer (NSCLC). Programmed death ligand-1 (PD-L1) plays important regulatory roles in intracellular functions and leads to acquired resistance to EGFR-TKIs in NSCLC. Here, we investigated the mechanistic role of PD-L1 in primary resistance to EGFR-TKIs in EGFR-mutant NSCLC cells.

METHODS

The expression levels of PD-L1 and the sensitivity to gefitinib in H1975, HCC827 and PC-9 cells were determined by quantitative real-time PCR analysis (qRT-PCR) and Cell Counting Kit-8 (CCK-8) assays, respectively. Molecular manipulations (silencing or overexpression) were performed to assess the effect of PD-L1 on sensitivity to gefitinib, and a mouse xenograft model was used for in vivo confirmation. Western blotting and qRT-PCR were used to analyse the expression of epithelial-mesenchymal transition (EMT) markers. The effect of PD-L1 on migratory and invasive abilities was evaluated using the Transwell assay and mice tail intravenous injection.

RESULTS

Higher expression of PD-L1 was related to less sensitivity to gefitinib in EGFR-mutant NSCLC cell lines. The overexpression or knockdown of PD-L1 presented diametrical sensitivity to gefitinib in vitro and in vivo. Furthermore, the overexpression of PD-L1 led to primary resistance to gefitinib through the induction of EMT, which was dependent on the upregulation of Smad3 phosphorylation. Moreover, in the mouse model, the knockdown of PD-L1 inhibited transforming growth factor (TGF)-β1-induced cell metastasis in vivo.

CONCLUSION

PD-L1 contributes to primary resistance to EGFR-TKI in EGFR-mutant NSCLC cells, which may be mediated through the induction of EMT via the activation of the TGF-β/Smad canonical signalling pathway.

背景

大约 30%的表皮生长因子受体（EGFR）激活突变患者对 EGFR 酪氨酸激酶抑制剂（TKI）无反应（原发性耐药）。然而，对于 EGFR 突变非小细胞肺癌（NSCLC）中 EGFR-TKI 原发性耐药的相关分子机制知之甚少。程序性死亡配体-1（PD-L1）在细胞内功能中发挥重要的调节作用，并导致 NSCLC 对 EGFR-TKI 的获得性耐药。在这里，我们研究了 PD-L1 在 EGFR 突变 NSCLC 细胞对 EGFR-TKI 原发性耐药中的机制作用。

方法

通过定量实时 PCR 分析（qRT-PCR）和细胞计数试剂盒-8（CCK-8）检测 H1975、HCC827 和 PC-9 细胞中 PD-L1 的表达水平和对吉非替尼的敏感性。进行分子操作（沉默或过表达）以评估 PD-L1 对吉非替尼敏感性的影响，并使用小鼠异种移植模型进行体内验证。Western blot 和 qRT-PCR 用于分析上皮-间充质转化（EMT）标志物的表达。使用 Transwell 测定法和小鼠尾静脉注射评估 PD-L1 对迁移和侵袭能力的影响。

结果

PD-L1 表达较高与 EGFR 突变 NSCLC 细胞系对吉非替尼的敏感性降低有关。PD-L1 的过表达或敲低在体外和体内均表现出对吉非替尼的截然相反的敏感性。此外，PD-L1 的过表达通过诱导 EMT 导致对吉非替尼的原发性耐药，这依赖于 Smad3 磷酸化的上调。此外，在小鼠模型中，PD-L1 的敲低抑制了转化生长因子（TGF）-β1 在体内诱导的细胞转移。