Sorafenib Resistance Contributed by IL7 and MAL2 in Hepatocellular Carcinoma Can Be Overcome by Autophagy-Inducing Stapled Peptides

SIMPLE SUMMARY: Drug resistance remains a major challenge in the treatment of hepatocellular carcinoma (HCC). Our objective was to investigate the genetic mechanisms involved in the development of Sorafenib-associated drug resistance in HCC cells. To achieve this, transposon insertional mutagenesis was utilized as a forward genetic tool to generate Sorafenib-resistant HCC cell lines and identify potential drug resistant genes. Our studies identified two genes, interleukin 7 (IL7) and mal, T cell differentiation protein 2 (MAL2), which conferred resistance to Sorafenib by activating the pro-survival JAK/STAT and PI3K/AKT signaling pathways. Additionally, autophagy inducing stapled peptides exhibited an anti-proliferative effect on both wild-type and drug-resistant HCC cells. When combined with Sorafenib, these peptides demonstrated efficacy against drug-resistant cells. Therefore, targeting autophagy could present a novel approach for overcoming Sorafenib-associated drug resistance in HCC induced by IL7- and MAL2-overexpression. ABSTRACT: Drug resistance poses a great challenge in systemic therapy for hepatocellular carcinoma (HCC). However, the underlying molecular mechanisms associated with resistance to anti-cancer drugs, such as Sorafenib, remain unclear. In this study, we use transposon insertional mutagenesis to generate Sorafenib-resistant HCC cell lines in order to identify potential drug resistant causative genes. Interleukin 7 (IL7) and mal, T cell differentiation protein 2 (MAL2) were identified as candidate genes that promote survival by activating JAK/STAT and PI3K/AKT signaling pathways. Sorafenib-resistant cells exhibited higher clonogenic survival and lower drug sensitivity due to IL7 and MAL2 upregulation. Higher anti-apoptotic effect, clonogenic survival and increased PI3K/AKT/STAT3 activities were observed in IL7 and MAL2 co-overexpressing cells compared with controls or cells overexpressing IL7 or MAL2 individually. Given the critical role of MAL2 in endocytosis, we propose that MAL2 might facilitate the endocytic trafficking of IL7 and its cognate receptors to the plasma membrane, which leads to upregulated JAK/STAT and PI3K/AKT signaling pathways and Sorafenib resistance. Additionally, our previous studies showed that an autophagy-inducing stapled peptide promoted the endolysosomal degradation of c-MET oncogene and overcame adaptive Sorafenib resistance in c-MET(+) HCC cells. In this study, we demonstrate that these stapled peptides readily induced autophagy and inhibited the proliferation of both wild-type and Sorafenib-resistant HCC cells co-overexpressing both IL7 and MAL2. Furthermore, these peptides showed synergistic cytotoxicity with Sorafenib in drug-resistant HCC cells co-overexpressing both IL7 and MAL2. Our studies suggest that targeting autophagy may be a novel strategy to overcome IL7/MAL2-mediated Sorafenib resistance in HCC.