Abstract
BackgroundChemo-resistance is one of the major challenges in the therapy of small cell lung cancer (SCLC). Multiple mechanisms are thought to be involved in chemo-resistance during SCLC treatment, but unfortunately, these mechanisms have not been well elucidated. Herein, we investigated the role of miRNA in the resistance of SCLC cells to doxorubicin (Dox).MethodsMiRNA microarray analysis revealed that several miRNAs, including miR-7-5p, were specifically decreased in Dox-resistant SCLC cells (H69AR) compared to parental cells (H69). The expression level of miR-7-5p was confirmed by qRT-PCR in Dox-resistant cells (H69AR and H446AR cells) and their parental cells. Bioinformatic analysis indicated that poly ADP-ribose polymerase 1 (PARP1) is a direct target of miR-7-5p. The binding sites of miR-7-5p in the PARP1 3′ UTR were verified by luciferase reporter and Western blot assays. To investigate the role of miR-7-5p in the chemo-resistance of SCLC cells to doxorubicin, mimic or inhibitor of miR-7-5p was transfected into SCLC cells, and the effect of miR-7-5p on homologous recombination (HR) repair was analyzed by HR reporter assays. Furthermore, the expression of HR repair factors (Rad51 and BRCA1) induced by doxorubicin was detected by Western blot and immunofluorescent staining in H446AR cells transfected with miR-7-5p mimic.ResultsThe expression level of miR-7-5p was remarkably reduced (4-fold) in Dox-resistant SCLC cells (H69AR and H446AR cells) compared with that in parental cells (H69 and H446 cells). Poly ADP-ribose polymerase 1 (PARP1) is a direct target of miR-7-5p, and PARP1 expression was downregulated by miR-7-5p. MiR-7-5p impeded Dox-induced HR repair by inhibiting the expression of HR repair factors (Rad51 and BRCA1) that resulted in resensitizing SCLC cells to doxorubicin.ConclusionsOur findings provide evidence that miR-7-5p targets PARP1 to exert its suppressive effects on HR repair, indicating that the alteration of the expression of miR-7-5p may be a promising strategy for overcoming chemo-resistance in SCLC therapy.
Highlights
Chemo-resistance is one of the major challenges in the therapy of small cell lung cancer (SCLC)
MiR-7-5p is negatively correlated with dox resistance in SCLC cells and enhances doxorubicin cytotoxicity Two Dox-resistant SCLC cell lines (H69AR and H446AR) were analyzed by a Cell counting kit-8 (CCK-8) assay
MiR-7-5p reduces BRCA1 and Rad51 expression and disrupts homologous recombination (HR) repair induced by doxorubicin in doxresistant SCLC cells by targeting poly ADP-ribose polymerase 1 (PARP1) Since PARP1 is a target of miR-7-5p and plays a crucial role in the HR repair pathway, we investigated the role of the PARP1 inhibitor ABT-888 (Veliparib) in HR repair in H446 and H446AR cells
Summary
Chemo-resistance is one of the major challenges in the therapy of small cell lung cancer (SCLC). Acquired chemo-resistance is considered the major drawback of current chemotherapeutic regimens, but the molecular details have not been completely elucidated. DNA-damaging agents are the most widely used chemotherapeutic drugs [3]. DNA-damaging agents, such as doxorubicin (Adriamycin, Dox), prevent cell division and lead to cell death by inhibiting the religation of DNA strands in double-strand breaks (DSBs) [4]. Cancer cells may acquire chemo-resistance by altering the cell survival signaling pathway and repairing the DNA damage [5]. The DNA damage response (DDR) is a molecular mechanism that cancer cells have exploited to activate DNA repair pathways and prevent DNA damage-induced cell death [6]
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