Heilos Daniela, Röhrl Clemens, Pirker Christine, Englinger Bernhard, Baier Dina, Mohr Thomas, Schwaiger Michaela, Iqbal Shahid Muhammad, van Schoonhoven Sushilla, Klavins Kristaps, Eberhart Tanja, Windberger Ursula, Taibon Judith, Sturm Sonja, Stuppner Hermann, Koellensperger Gunda, Dornetshuber-Fleiss Rita, Jäger Walter, Lemmens-Gruber Rosa, Berger Walter
Institute of Cancer Research, Department of Internal Medicine I, Medical University of Vienna, Comprehensive Cancer Center of the Medical University of Vienna, Vienna, Austria.
Department of Pharmacology and Toxicology, University of Vienna, Vienna, Austria.
Oncotarget. 2018 May 22;9(39):25661-25680. doi: 10.18632/oncotarget.25432.
Destruxins, secondary metabolites of entomopathogenic fungi, exert a wide variety of interesting characteristics ranging from antiviral to anticancer effects. Although their mode of action was evaluated previously, the molecular mechanisms of resistance development are unknown. Hence, we have established destruxin-resistant sublines of HCT116 colon carcinoma cells by selection with the most prevalent derivatives, destruxin (dtx)A, dtxB and dtxE. Various cell biological and molecular techniques were applied to elucidate the regulatory mechanisms underlying these acquired and highly stable destruxin resistance phenotypes. Interestingly, well-known chemoresistance-mediating ABC efflux transporters were not the major players. Instead, in dtxA- and dtxB-resistant cells a hyper-activated mevalonate pathway was uncovered resulting in increased cholesterol synthesis rates and elevated levels of lanosterol, cholesterol as well as several oxysterol metabolites. Accordingly, inhibition of the mevalonate pathway at two different steps, using either statins or zoledronic acid, significantly reduced acquired but also intrinsic destruxin resistance. Vice versa, cholesterol supplementation protected destruxin-sensitive cells against their cytotoxic activity. Additionally, an increased cell membrane adhesiveness of dtxA-resistant as compared to parental cells was detected by atomic force microscopy. This was paralleled by a dramatically reduced ionophoric capacity of dtxA in resistant cells when cultured in absence but not in presence of statins. Summarizing, our results suggest a reduced ionophoric activity of destruxins due to cholesterol-mediated plasma membrane re-organization as molecular mechanism underlying acquired destruxin resistance in human colon cancer cells. Whether this mechanism might be valid also in other cell types and organisms exposed to destruxins e.g. as bio-insecticides needs to be evaluated.
destruxins是昆虫病原真菌的次生代谢产物，具有从抗病毒到抗癌等多种有趣的特性。尽管其作用模式先前已被评估，但耐药性产生的分子机制尚不清楚。因此，我们通过用最常见的衍生物destruxin（dtx）A、dtxB和dtxE进行筛选，建立了HCT116结肠癌细胞的抗destruxin亚系。应用了各种细胞生物学和分子技术来阐明这些获得性且高度稳定的抗destruxin表型背后的调控机制。有趣的是，众所周知的介导化疗耐药的ABC外排转运蛋白并不是主要因素。相反，在对dtxA和dtxB耐药的细胞中，发现甲羟戊酸途径过度激活，导致胆固醇合成速率增加以及羊毛甾醇、胆固醇和几种氧甾醇代谢物水平升高。相应地，使用他汀类药物或唑来膦酸在两个不同步骤抑制甲羟戊酸途径，显著降低了获得性以及固有性抗destruxin能力。反之，补充胆固醇可保护对destruxin敏感的细胞免受其细胞毒性作用。此外，通过原子力显微镜检测发现，与亲代细胞相比，dtxA耐药细胞的细胞膜黏附性增加。当在无他汀类药物但不在有他汀类药物的情况下培养时，dtxA在耐药细胞中的离子载体能力显著降低。总之，我们的结果表明，由于胆固醇介导的质膜重组，destruxins的离子载体活性降低，这是人类结肠癌细胞获得性抗destruxin耐药性的分子机制。这种机制在暴露于destruxins的其他细胞类型和生物体（例如作为生物杀虫剂）中是否也有效，还有待评估。
