Chemoprevention via modulation of autophagy by sphingolipids

描述(由申请人提供)： 自噬是癌症化学预防和化疗的一个很有前途的新靶点，因为它有可能预防、逆转或延缓癌前病变的进展，并通过“II型”或“自噬”细胞死亡来杀死肿瘤细胞。与癌症预防和治疗相关的自噬诱导剂包括白藜芦醇等天然产品和抗癌药物芬维甲胺(4-羟基维甲酰胺)等合成化合物；然而，自噬与癌症之间的联系是复杂的，因为它也为癌细胞在营养限制和代谢压力时期提供了一种生存手段。因此，最佳药剂或药剂组合更倾向于“致命性”而非“保护性”自噬。这项资助申请涉及最近发现的一种诱导自噬的机制，该机制似乎控制了这些结果之间的切换。其机制涉及神经酰胺(Cer)、二氢神经酰胺(DHCer)和1-磷酸鞘氨醇(S1P)诱导的自噬，而S1P是决定自噬是致死性还是保护性的关键因素。这项研究的基本假设是，在诱导自噬的过程中，(DH)Cer被结合到自噬小体中，随后(在自噬溶酶体中)被水解为鞘氨醇和鞘氨醇，这两种物质是细胞毒性的，除非被鞘氨醇激酶磷酸化。因此，通过自噬细胞死亡进行化学预防和化疗的最有效方案将是有利于产生(DH)Cer和狮身人面像碱基，而不是狮身人面像碱基1-磷酸。这种干预策略将使用表达GFP标记的LC3的MCF7细胞来促进自噬的监测，对这些脂介质中变化的神经鞘脂体进行分析的方法，以及在细胞被两种典型的饮食和化疗药物(分别为白藜芦醇和4HPR)处理时可以操纵鞘脂代谢的工具，以及为提供机械信息而选择的其他化合物。研究结果可能定义癌症化学预防和化疗的新概念，并在实验室和临床随访研究中确定用于评估这些和其他药物的有用工具和生物标记物。

DESCRIPTION (provided by applicant): Autophagy is a promising new target for cancer chemoprevention and chemotherapy because it has the potential to prevent, reverse, or retard progression of precancerous lesions as well as to kill tumor cells by "type II" or "autophagic" cell death. Inducers of autophagy that have been associated with cancer prevention and treatment include natural products such as resveratrol and synthetic compounds such as the anti- cancer drug fenretinide (4-hydroxy-phenylretinamide); nonetheless, the link between autophagy and cancer is complex because it also affords a means of survival for cancer cells during times of nutrient limitation and metabolic stress. Hence, the optimal agent, or combination of agents, would favor "lethal" versus "protective" autophagy. This grant application concerns a recently discovered mechanism for induction of autophagy that appears to control the switch between these outcomes. The mechanism involves the induction of autophagy by ceramides (Cer), dihydroceramides (DHCer), and sphingosine 1-phosphate (S1P), with the latter being a critical determinant of whether autophagy is lethal or protective. The underlying hypothesis of this research is that in the induction of autophagy, (DH)Cer are incorporated into autophagosomes and subsequently hydrolyzed (in autophagolysosomes) to sphinganine and sphingosine, which are cytotoxic unless phosphorylated by sphingosine kinase. Thus, the most effective scenario for chemoprevention and chemotherapy via autophagic cell death would be to favor the production of (DH)Cer and sphingoid bases over sphingoid base 1-phosphates. This intervention strategy will be explored using MCF7 cells that express GFP-tagged LC3 to facilitate the monitoring of autophagy, methods for "sphingolipidomic" analysis of changes in these lipid mediators, and tools that can manipulate sphingolipid metabolism as the cells are treated with two prototypic dietary and chemotherapeutic agents (resveratrol and 4HPR, respectively) as well as other compounds selected for mechanistic information they will provide. The findings of the studies could define a new concept in cancer chemoprevention and chemotherapy, and identify useful tools and biomarkers for evaluation of these and additional agents in laboratory and clinical follow- up studies.