Inhibition of Autophagy Enhances Anticancer Effects of Atorvastatin in Digestive Malignancies

Department of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan.
Cancer Research (Impact Factor: 9.33). 09/2010; 70(19):7699-709. DOI: 10.1158/0008-5472.CAN-10-1626
Source: PubMed


Preclinical and clinical studies have shown that statins, the 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors with cholesterol-lowering properties, exhibited anticancer effects. However, the underlying mechanisms remain ill defined. In this study, we showed that atorvastatin could inhibit the growth of hepatocellular carcinoma (HCC) and colorectal carcinoma (CRC) cells via induction of apoptosis. Atorvastatin also induced autophagy that is a physiologic process involved in the turnover of intracellular organelles. Atorvastatin-induced autophagy was found to be inhibited by AMP-activated protein kinase (AMPK) small interfering RNA. Examination of HCC patients showed the positive correlation between AMPK activity and autophagic marker (beclin-1). Atorvastatin-induced AMPK activation could induce p21 expression, which was also positively correlated with beclin-1 expression in CRC patients. AMPK/p21 signaling caused endoplasmic reticulum (ER) stress response leading to the induction of autophagy. Inhibition of autophagy by an autophagic inhibitor bafilomycin A1 or genetic knockout of autophagy-related gene 5 enhanced atorvastatin-induced cytotoxicity and apoptosis. In summary, activation of AMPK by atorvastatin enhances p21 expression and ER stress response, leading to autophagy, which promotes survival of cancer cells. Combinations of atorvastatin with bafilomycin A1 provide a novel and promising strategy to improve the treatment of digestive malignancies.

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    • "We have previously shown that statin-induced autophagy is cytoprotective in glioma cells, and that inhibition of autophagy in statin-treated glioma cells enhances their apoptotic death (Misirkic et al., 2012). These results were consistent with similar observations in digestive malignancies, ovarian and bladder cancer, and human airway mesenchymal cells (Ghavami et al., 2011, 2014, 2012; Kang et al., 2014; Robinson et al., 2013; Yang et al., 2010). In concord with these data, our present study shows that inhibition of autophagy reveals a dramatic apoptosis-inducing effects of statins and Ro 48-8071 in leukemic cells, an effect not seen with protein prenylation inhibitors, or in normal human leukocytes. "
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    ABSTRACT: Statins exhibit anti-leukemic properties due to suppression of the mevalonate pathway by the inhibition of 3-hydroxy-3-methylglutaryl coenzyme A reductase, and subsequent depletion of cholesterol, farnesylpyrophosphate, and geranylgeranylpyrophosphate. We investigated the role of autophagy, a controlled intracellular self-digestion, in the anti-leukemic action of statins. Treatment with low concentrations (≤6µM) of statins, cholesterol depletion, and specific inhibition of cholesterol synthesis and protein farnesylation or geranylgeranylation, all inhibited proliferation of leukemic cell lines and primary leukemic cells without inducing overt cell death. Statins and agents that selectively reduce intracellular cholesterol levels, but not the inhibition of protein farnesylation or geranylgeranylation, induced autophagy in leukemic cells. The observed autophagic response was associated with the reduction of phosphorylated Akt levels in the lipid rafts, accompanied by a decrease in the activation of the main autophagy suppressor mammalian target of rapamycin (mTOR) and its substrate ribosomal p70S6 kinase (p70S6K). No significant autophagy induction and downregulation of mTOR/p70S6K activation were observed in normal leukocytes. Autophagy suppression by bafilomycin A1 or RNA interference-mediated knockdown of beclin-1 and microtubule-associated protein 1 light chain 3B induced apoptotic death in statin-treated leukemic cells, an effect attenuated by the addition of mevalonate or squalene, but not farnesylpyrophosphate or geranylgeranylpyrophosphate. Therefore, while the inhibition of cholesterol synthesis, protein farnesylation, and geranylgeranylation all contributed to anti-leukemic effects of statins, the inhibition of cholesterol synthesis was solely responsible for the induction of cytoprotective autophagy. These data indicate that combined treatment with statins and autophagy inhibitors might be potentially useful in anti-leukemic therapy.
    Full-text · Article · Sep 2015 · European journal of pharmacology
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    • "Baf, 3-MA or CHX not only suppressed Rott induced phosphorylation of AMPK but also attenuated the expression of LC3 and Atg12. All these observations are in agreement with several studies showing that activation of AMPK leads to autophagy [33,45,61]. "
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    ABSTRACT: Autophagy is an indispensable lysosomal self-digestion process involved in the degradation of aggregated proteins and damaged organelles. Autophagy is associated with the several pathological processes, including cancer. Cancer stem cells (CSCs) play significant roles in cancer initiation, progression and drug resistance. Recent studies have demonstrated the antitumor activities of plant-derived chemopreventive agent rottlerin (Rott). However, the molecular mechanism by which Rott induces autophagy in breast CSCs has not been investigated. The objectives of this study were to examine the molecular mechanism by which Rott induces autophagy which leads to apoptosis in breast CSCs. Treatment of breast CSCs with Rott for 24 h resulted in a concentration dependent induction of autophagy, followed by apoptosis as measured by flow cytometry. Electron microscopy confirmed the presence of autophagosomes in Rott treated breast CSCs. Western blot analysis showed that Rott treatment increased the expression of LC3, Beclin-1 and Atg12 that are accumulated during autophagy. Prolonged exposure of breast CSCs to Rott caused apoptosis which was associated with the suppression of phosphorylated Akt and mTOR, upregulation of phosphorylated AMPK, and downregulation of anti-apoptosis Bcl-2, Bcl-XL, XIAP and cIAP-1. Knock-down of Atg7 or Beclin-1 by shRNA inhibited Rott-induced autophagy at 24 h. Our study also demonstrates that pre-treatment of breast CSCs with autophagosome inhibitors 3-methyladenine and Bafilomycin, as well as protein synthesis inhibitor cycloheximide inhibited Rott-induced autophagy and apoptosis. Rott induces autophagy via extensive cytoplasmic vacuolization in breast CSCs. Molecular docking results between C2-domain of protein kinase C-delta and Rott indicated that both hydrogen bonding and hydrophobic interactions contributed significantly for ligand binding with minimum binding affinity of [almost equal to] 7.5 Kcal/mol. Although, autophagy inhibitors suppress the formation of cytoplasmic vacuolization and autophagy in breast CSCs, the potency of Rott to induce autophagy and apoptosis might be based on its capability to activate several pathways such as AMPK and proteasome inhibition. A better understanding of the relationship between autophagy and apoptosis would eventually allow us to discover novel drugs for the treatment of breast cancer by eliminating CSCs.
    Full-text · Article · Dec 2013 · Molecular Cancer
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    • "How p53 triggers ER stress is still unclear. Our recent study demonstrates that cytoplasmic p21 induces ER stress38. However, Zeb-induced p21 only partially contributed to ER stress (Supplementary Figure 7), indicating that other p53 target genes might be involved. "
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    ABSTRACT: Aberrant DNA hypermethylation is frequently found in tumor cells and inhibition of DNA methylation is an effective anticancer strategy. In this study, the therapeutic effect of DNA methyltransferase (DNMT) inhibitor zebularine (Zeb) on colorectal cancer (CRC) was investigated. Zeb exhibited anticancer activity in cell cultures, tumor xenografts and mouse colitis-associated CRC model. It stabilizes p53 through ribosomal protein S7 (RPS7)/MDM2 pathways and DNA damage. Zeb-induced cell death was dependent on p53. Microarray analysis revealed that genes related to endoplasmic reticulum (ER) stress and unfolded protein response (UPR) were affected by Zeb. Zeb induced p53-dependent ER stress and autophagy. Pro-survival markers of ER stress/UPR (GRP78) and autophagy (p62) were increased in tumor tissues of CRC patients, AOM/DSS-induced CRC mice and HCT116-derived colonospheres. Zeb downregulates GRP78 and p62, and upregulates a pro-apoptotic CHOP. Our results reveal a novel mechanism for the anticancer activity of Zeb.
    Full-text · Article · Nov 2013 · Scientific Reports
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