Financial support statement: This study was supported by the Deutsche Krebshilfe (111147).Data availability: GEO accession numbers for miRNA expression array data (Exiqon) and gene expression profiling data (HZI) are NCBI GEO: GSE152920 and GSE152950. Authors contributions: A.B conceived the idea, designed the study, and provided the conceptual framework for the study. R-M.K, QH, and A.B performed all experiments and analyzed the data. L.vD and J.D helped with the cloning and the in vitro experiments. H-C.T helped with the xenograft experiments. Q.Y helped with animal experiments. O.O helped with the metabolic assays and luciferase reporter assays for the revised manuscript. AG provided us with the TRE-Myc, TRE-RAS, and LAP-tTA transgenic mouse models, which we bred and crossed to obtain the double transgenic LT2/MYC and LT2/RAS mice. A.B wrote the manuscript with the help of M.O, AD.S. and A.G. Y.X and R.Q helped with collection and analyses of human HCC tumor and matched non-tumor biopsies (China). A.B, M.P.M, M.O, and AD.S, provided conceptual evaluation of the project.
In contrast to normal tissue, cancer cells display profound alterations in protein synthesis and degradation. Therefore, proteins that regulate endoplasmic reticulum (ER) homeostasis are being increasingly recognized as potential therapeutic targets. The ubiquitin-proteasome system and autophagy are crucially important for proteostasis in cells. However, interactions between autophagy, the proteasome, and ER stress pathways in cancer remain largely undefined. This study demonstrated that withaferin-A (WA), the biologically active withanolide extracted from Withania somnifera, significantly increased autophagosomes, but blocked the degradation of autophagic cargo by inhibiting SNARE-mediated fusion of autophagosomes and lysosomes in human pancreatic cancer (PC) cells. WA specifically induced proteasome inhibition and promoted the accumulation of ubiquitinated proteins, which resulted in ER stress-mediated apoptosis. Meanwhile, the impaired autophagy at early stage induced by WA was likely activated in response to ER stress. Importantly, combining WA with a series of ER stress aggravators enhanced apoptosis synergistically. WA was well tolerated in mice, and displayed synergism with ER stress aggravators to inhibit tumor growth in PC xenografts. Taken together, these findings indicate that simultaneous suppression of 2 key intracellular protein degradation systems rendered PC cells vulnerable to ER stress, which may represent an avenue for new therapeutic combinations for this disease.
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