SCC-31

The mammalian target of rapamycin (mTOR) is a key regulatory molecular target to treat cancer, and MTI-31 is a potent mTOR inhibitory agent for the therapeutically target of the renal cell carcinoma (RCC). However, the therapeutic efficacy of MTI-31 is limited by multiple factors, including autophagy. MTI-31 can activate cells to generate autophagy, which may in turn indirectly affect cell proliferation and apoptosis. We aimed to observe changes in cell protective autophagy via the ERK pathway and explore the potential mechanism underlying drug resistance of RCC cells to MTI-31. Different concentrations of 786-O and RCC4 cells were co-cultured with MTI-31 for distinct durations. The result of autophagy marker detection by Western blot showed that MTI-31 could induce RCC cells to produce autophagy in a dose and time-dependent manner. After treating the RCC cells with the autophagy inhibitor chloroquine (CQ), CCK8 and Western blot assays demonstrated that CQ could effectively enhance cell apoptosis induced by MTI-31 and that the autophagy induced by MTI-31 was cytoprotective. In addition, CCK8 and Western blot demonstrated that MTI-31 exerted its effect by activating the ERK pathway rather than the JNK or p38 pathway. The use of the ERK inhibitor AZD6244 to block the ERK pathway could effectively promote cell apoptosis induced by MTI-31. AZD6244 attenuated the autophagy induced by MTI-31 and increased the cytotoxicity of MTI-31. Western blot also demonstrated that MTI-31-induced autophagy was mediated by the downstream regulators of ERK pathways, including Beclin-1 and Bcl-2. It demonstrated that the MTI-31 mediated activation ERK pathway is associated with the induction of autophagy, and autophagy can attenuate the cytotoxicity of MTI-31 on RCC cells. In summary, inhibition of ERK pathway-mediated autophagy can rectify drug resistance to MTI-31 effectively.

At most of the times, patients who are diagnosed with kidney cancer should be provided with systemic treatment as drug resistance is a challenging issue in the treatment of this disease. The progression of the cancer can be inhibited with the help of mTOR inhibitors namely RAD001 (everolimus) and MTI-31. In literature, it has been revealed that these mTOR inhibitors have the potential to stimulate autophagy. This degradation pathway boosts the survival rate of the cancerous cells that are subjected to anti-cancer therapy. In this study, CCK8, colony formation assays, and ethynyl deoxyuridine (EdU) analysis were conducted to detect cell proliferation. Furthermore, Transwell assays were also conducted for cell migration analysis. In addition to these, the researchers also performed the flow cytometry process to identify the cells that are undergoing apoptosis. In vivo, experiments were conducted to measure the growth of tumors and metastasis. In this study, the treatment provided through a combination of MTI-31 and RAD001 significantly inhibited the kidney cancer cells' proliferation and tumor growth. Furthermore, there was a notable reduction in the migration and invasion of kidney cancer cells upon the neighboring cells. The outcomes from the mechanistic studies infer that the combination of MTI-31 and RAD001 increases the LC3 levels, which in turn translates into the activation of autophagy. To conclude, the combination of MTI-31 and RAD001 improves the anti-cancerous impact produced by RAD001 in vivo through the promotion of autophagy.