Platinum-based doublets with a third-generation agent are the recommended option for many patients with non-small cell lung cancer (NSCLC) with no contraindications to platinum compounds. Unfortunately, the clinical effectiveness of such chemotherapy is limited by intrinsic or acquired resistance. Circulating exosomal miRNAs were isolated and used to perform HiSeq deep-sequencing analyses on serum pool samples from platinum-resistant or platinum-sensitive patients, and six exosomal miRNAs were further validated for their predictive utility by qRT-PCR in 170 serum samples of patients with advanced NSCLC. Gain- and loss-of-function experiments clarified the responsiveness regulating role of the clinically relevant miRNA. IHC analyses were performed to evaluate the association between basal autophagy in lung cancer tissues and responsiveness in 203 patients with NSCLC receiving platinum-based chemotherapy. Six circulating exosomal miRNAs (miR-425-3p, miR-1273h, miR-4755-5p, miR-9-5p, miR-146a-5p, and miR-215-5p) were found to be differentially expressed with the largest fold change in platinum-resistant patients compared with platinum-sensitive patients. High miR-425-3p proved to be a potent predictive biomarker for low responsiveness and poor progression-free survival (PFS). Mechanistically, miR-425-3p upregulated the autophagic levels via targeting AKT1, leading to the decrease in therapeutic response. Concordantly, high levels of basal autophagy in lung cancer tissues correlate with low responsiveness in patients with NSCLC within the early and advanced disease stages. Our study highlights circulating exosomal miR-425-3p as a potential biomarker for improved predictions of the clinical response to platinum-based chemotherapy in patients with NSCLC. This study provides the first evidence that miR-425-3p in NSCLC patient-derived exosomes can be a marker for predicating the clinical response to platinum-based chemotherapy.

IntroductionExosomes are important mediators of intercellular communication. Previously, we characterized circulating exosomal miR-425-3p as a non-invasive prognostic marker for predicting clinical response to platinum-based chemotherapy in patients with non-small cell lung cancer (NSCLC).MethodsCirculating exosomal miR-425-3p was validated by qRT-PCR in paired serum samples from NSCLC patients during the course of platinum-based chemotherapy. Cell coculture was performed to examine the effects of exosomal miR-425-3p on the sensitivity of recipient A549 cells to cisplatin. Using bioinformatics, ChIP and luciferase reporter assays, the transcription factor essential for miR-425-3p expression was identified. Autophagic activity in the recipient cells was determined by Western blot and fluorescence microscopy.ResultsHigher levels of exosomal miR-425-3p were found in serum samples from the patients in tolerance versus those at baseline. An upward trend in the expression of circulating exosomal miR-425-3p was revealed during chemotherapy. Furthermore, the expression of exosomal miR-425-3p could be induced by cisplatin in NSCLC cells. Exosomes isolated from either cisplatin-treated or cisplatin-resistant NSCLC cells conferred chemoresistance to sensitive A549 cells in a miR-425-3p-dependent manner. Cisplatin-induced c-Myc was found to directly bind the miR-425-3p promoter and transactivated its expression. Exosomal miR-425-3p facilitated autophagic activation in the recipient cells by targeting AKT1, eventually leading to chemoresistance.DiscussionOur results suggest that apart from a prognostic marker of treatment response, exosomal miR-425-3p might be a potential dynamic biomarker to tailor cisplatin resistance in NSCLC patients during the treatment and represent a promising therapeutic target for therapy-resistant NSCLC.

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Lung cancer is the most common cause of cancer-related death worldwide and the platinum-based drugs such as cisplatin have been used as the first line of the treatment. However, the clinical effectiveness of such chemotherapy is limited by intrinsic or acquired resistance. In this study, we found that cisplatin induced autophagy that attenuated the sensitivity of both A549 and Lewis lung cancer (LLC) cells to cisplatin. In contrast, the clinical drug andrographolide (Andro) suppressed autophagy and enhanced cisplatin-mediated apoptosis in these cells. Using two murine lung cancer models, including a subcutaneously inoculated LLC model and an orthotopic LLC implantation model, we investigated the therapeutic efficacy of the combined treatment of cisplatin and Andro. Compared with the sole cisplatin treatment, combining cisplatin with Andro potentially inhibited tumor growth, reduced the incidence of lung metastases, and relieved renal tubular damage. Moreover, the combined treatment prolonged the life span of tumor-bearing mice. TUNEL and immunohistochemistry assays showed the increase in apoptotic cells and the decrease in both conversion of LC3B-I to LC3B-II and Atg5 protein expression in the tumor tissues from mice with the combined treatment. These results suggest that Andro offers an ideal candidate of autophagy inhibitors in clinical application, and combination of cisplatin with Andro could be a promising strategy for the treatment of lung cancer.