RESUMEN
OBJECTIVE: Apurinic endonuclease 1 (APE1) has been suggested as an oncogene of lung tumours and our bioinformatics analysis identified the association between Erlotinib resistance and interleukin-6 (IL-6). Thus, we performed this work to delineate the mechanistic actions of APE1/IL-6 signalling in Erlotinib resistance of non-small cell lung cancer (NSCLC). METHODS: We selected human NSCLC cell lines HCC827 and PC9 to establish Erlotinib-resistant HCC827R and PC9R cells. Cancer stem cells (CSCs) were isolated from Erlotinib-sensitive HCC827P and PC9P cells (PCSCs) and from HCC827R and PC9R cells (RCSCs). Further, extracellular vesicles (EVs) were separated from PCSCs (PCSC-EVs) and RCSCs (RCSC-EVs) and co-cultured with RCSCs with or without short hairpin RNA (shRNA)-targeting APE1 (APE1 shRNA) transduction. In addition, functional assays were conducted to determine the effect of APE1 shRNA on malignant phenotypes of cancer cells in vitro and in vivo and the activation of IL-6/STAT3 signalling. RESULTS: It was found that NSCLC cells could internalize both RCSC-EVs and PCSC-EVs. RCSC-EVs augmented the resistance of NSCLC cells to Erlotinib. The overexpression of APE1 occurred in NSCLC tissues, and IL-6 was enriched in serum samples of patients with NSCLC. APE1 shRNA was demonstrated to restrict the Erlotinib resistance of NSCLC cells by inactivating the IL-6/STAT3 signalling. Additionally, shAPE1-loaded RCSC-EVs suppressed the Erlotinib resistance of NSCLC via the IL-6/STAT3 axis both in vitro and in vivo, as reflected by impeded malignant phenotypes and xenograft tumour formation. CONCLUSIONS: Collectively, these data indicate that APE1 confers Erlotinib resistance by activating the IL-6/STAT3 signalling, suggesting targeting APE1 as a possible therapeutic target in Erlotinib-resistant NSCLC.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Vesículas Extracelulares , Neoplasias Pulmonares , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , ADN-(Sitio Apurínico o Apirimidínico) Liasa/genética , ADN-(Sitio Apurínico o Apirimidínico) Liasa/metabolismo , ADN-(Sitio Apurínico o Apirimidínico) Liasa/uso terapéutico , Clorhidrato de Erlotinib/metabolismo , Clorhidrato de Erlotinib/farmacología , Clorhidrato de Erlotinib/uso terapéutico , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/patología , Humanos , Interleucina-6/metabolismo , Interleucina-6/uso terapéutico , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , ARN Interferente Pequeño/metabolismo , ARN Interferente Pequeño/uso terapéutico , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/metabolismo , Factor de Transcripción STAT3/uso terapéuticoRESUMEN
SOX2 is related to drug resistance in many types of cancer, including lung cancer. Herein, we investigated the role of SOX2 and its regulatory signaling in cisplatin-treated non-small-cell lung cancer (NSCLC). The effects of SOX2 on cell viability, proliferation, and apoptosis were evaluated in vitro. Western blotting, real-time quantitative PCR, immunohistochemistry, and luciferase reporter assays were used to investigate the underlying mechanism. Kaplan-Meier survival analysis and the log-rank test were used to assess the relationship between SOX2 expression and patient survival. A549/CDDP cells had marked resistance to cisplatin and stronger colony formation ability than A549 cells. The expression of SOX2 protein or mRNA in A549/CDDP was higher than that in A549. Knockdown of SOX2 in A549/CDDP-induced apoptosis by inhibiting colony formation and decreasing viability, but overexpression of SOX2 reversed these effects. Interestingly, Genomatix software predicted that the APE1 promoter has some SOX2 binding sites, while the SOX2 promoter has no APE1 binding sites. Furthermore, luciferase reporter assays proved that SOX2 could bind the promoter of APE1 in 293T cells. We further verified that SOX2 expression was not affected by shAPE1 in A549/CDDP. As expected, colony formation was obviously inhibited and apoptosis was strongly enhanced in A549/CDDP treated with SOX2 siSOX2 alone or combined with CDDP compared with control cells. Meaningfully, patients with low expression of SOX2, and even including its regulating APE1, survived longer than those with high expression of SOX2, and APE1. siSOX2 overcomes cisplatin resistance by regulating APE1 signaling, providing a new target for overcoming cisplatin resistance in NSCLC.