Purpose

Metastasis of lung adenocarcinoma (LADC) is a crucial factor determining patient survival. Repurposing of the antipsychotic agent penfluridol has been found to be effective in the inhibition of growth of various cancers. As yet, however, the anti-metastatic effect of penfluridol on LADC has rarely been investigated. Herein, we addressed the therapeutic potential of penfluridol on the invasion/metastasis of LADC cells harboring different epidermal growth factor receptor (EGFR) mutation statuses.

Methods

MTS viability, transwell migration and invasion, and tumor endothelium adhesion assays were employed to determine cytotoxic and anti-metastatic effects of penfluridol on LADC cells. Protease array, Western blot, immunohistochemistry (IHC), immunofluorescence (IF) staining, and expression knockdown by shRNA or exogenous overexpression by DNA plasmid transfection were performed to explore the underlying mechanisms, both in vitro and in vivo.

Results

We found that nontoxic concentrations of penfluridol reduced the migration, invasion and adhesion of LADC cells. Protease array screening identified matrix metalloproteinase-12 (MMP-12) as a potential target of penfluridol to modulate the motility and adhesion of LADC cells. In addition, we found that MMP-12 exhibited the most significantly adverse prognostic effect in LADC among 39 cancer types. Mechanistic investigations revealed that penfluridol inhibited the urokinase plasminogen activator (uPA)/uPA receptor/transforming growth factor-β/Akt axis to downregulate MMP-12 expression and, subsequently, reverse MMP-12-induced epithelial–mesenchymal transition (EMT). Subsequent analysis of clinical LADC samples revealed a positive correlation between MMP12 and mesenchymal-related gene expression levels. A lower survival rate was found in LADC patients with a SNAl1^high/MMP12^high profile compared to those with a SNAl1^low/MMP12^low profile.

Conclusions

Our results indicate that MMP-12 may serve as a useful biomarker for predicting LADC progression and as a promising penfluridol target for treating metastatic LADC.

中文翻译：

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通过重新利用五氟利多阻断 MMP-12 调节的上皮间质转化通过 uPA/uPAR/TGF-β/Akt 通路抑制肺腺癌转移

目的

肺腺癌（LADC）的转移是决定患者生存的关键因素。已发现抗精神病药五氟利多的再利用可有效抑制各种癌症的生长。然而，迄今为止，很少研究五氟利多对 LADC 的抗转移作用。在此，我们探讨了五氟利多对具有不同表皮生长因子受体 (EGFR) 突变状态的 LADC 细胞侵袭/转移的治疗潜力。

方法

MTS 活力、transwell 迁移和侵袭以及肿瘤内皮粘附测定用于确定五氟利多对 LADC 细胞的细胞毒性和抗转移作用。进行了蛋白酶阵列、蛋白质印迹、免疫组织化学 (IHC)、免疫荧光 (IF) 染色和 shRNA 表达敲低或 DNA 质粒转染的外源过表达，以探索体外和体内的潜在机制。

结果

我们发现五氟利多的无毒浓度降低了 LADC 细胞的迁移、侵袭和粘附。蛋白酶阵列筛选将基质金属蛋白酶 12 (MMP-12) 确定为五氟利多的潜在靶标，以调节 LADC 细胞的运动性和粘附性。此外，我们发现 MMP-12 在 LADC 的 39 种癌症类型中表现出最显着的不良预后作用。机制研究表明，五氟利多通过抑制尿激酶纤溶酶原激活物 (uPA)/uPA 受体/转化生长因子-β/Akt 轴来下调 MMP-12 表达，从而逆转 MMP-12 诱导的上皮-间质转化 (EMT)。随后对临床 LADC 样本的分析显示MMP12 与和间充质相关的基因表达水平。与SNAl1^低/ MMP12^低的患者相比，SNAl1^高/ MMP12^高的 LADC患者的存活率较低。

结论

我们的研究结果表明，MMP-12 可作为预测 LADC 进展的有用生物标志物和作为治疗转移性 LADC 的有希望的五氟利多靶标。