PHOSPHATIDYLCHOLINE BIOSYNTHESIS AND ANTICANCER AGENT MILTEFOSINE

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 米替福新是一种具有抗癌和抗菌活性的磷酸胆碱类似物。尽管其目前的医学应用，其作用模式仍然知之甚少。我们将使用人类寄生虫利什曼原虫作为模型系统，以加深我们对米替福新作用模式的理解。磷脂酰胆碱是哺乳动物细胞和利什曼原虫中最丰富的细胞脂质，由细胞外胆碱摄取的从头途径和一种或几种磷脂酰乙醇胺甲基转移酶对磷脂酰乙醇胺的三倍甲基化产生。与癌细胞相似，利什曼原虫对米替福新高度敏感，米替福新可减少磷脂酰乙醇胺合成磷脂酰胆碱 在癌细胞和利什曼原虫中的作用本申请的目标是鉴定参与从磷脂酰乙醇胺合成磷脂酰胆碱的利什曼酶，并评估它们是否是米替福新的靶点。虽然胆碱对利什曼原虫的生存能力是不利的，但我们的初步数据表明，利什曼原虫的两个磷脂酰乙醇胺甲基转移酶直系同源物LdPEM 1和LdPEM 2。Donovani，是必不可少的。我们的假设是LdPEM 1和LdPEM 2的甲基化途径代表了磷脂酰胆碱生物合成的主要途径，并且其中至少一种被米替福新抑制。我们的假设将通过执行以下特定目的来检验：1，确定LdPEM 1和LdPEM 2酶活性和米替福新的抑制谱; 2，确定LdPEM 1的过表达是否与LdPEM 2的表达相关。 和/或LdPEM 2调节利什曼原虫对米替福新的敏感性;和3，证实LdPEM 1和LdPEM 2在利什曼原虫生存力中的重要性。这些研究预计将显着提高我们的知识磷脂酰胆碱在利什曼原虫的生物合成和米替福新的行动模式，并提供更有效的抗癌药物的发展的分子基础。该项目的目标是更好地了解抗癌药物米替福新的作用机制。拟议的研究可能对癌症的治疗具有重要意义，因此， 直接关系到公共卫生。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Miltefosine is a phosphocholine analog with anticancer and antimicrobial activities. Despite its current medical application, its mode of action is still poorly understood. We will use the human parasite Leishmania as a model system to deepen our understanding of miltefosine's mode of action. Phosphatidyicholine, the most abundant cellular lipid in mammalian cells and in Leishmania, is produced from the de novo pathway from the uptake of extracellular choline, and from the three-fold methylation of phosphatidylethanolamine by one or several phosphatidylethanolamine methyltransferase(s). Similar to cancer cells, Leishmania is highly sensitive to miltefosine, which decreases phosphatidyicholine biosynthesis from phosphatidylethanolamine in both cancer cells and Leishmania. The goal of this application is to identify the leishmanial enzymes involved in phosphatidylcholine biosynthesis from phosphatidylethanolamine and assess whether they are the targets of miltefosine. While choline is dispensable for Leishmania viability, our preliminary data suggest that two phosphatidylethanolamine methyltransferase orthologs, LdPEM1 and LdPEM2 of L. donovani, are essential. Our hypothesis is that the methylation pathway made by LdPEM1 and LdPEM2 represents the primary route for phosphatidyicholine biosynthesis, and that at least one of them is inhibited by miltefosine. Our hypothesis will be tested by performing the following Specific Aims: 1, determine LdPEM1 and LdPEM2 enzymatic activities and inhibition profiles by miltefosine; 2, determine whether overexpression of LdPEM1 and/or LdPEM2 modulate(s) Leishmania sensitivity to miltefosine; and 3, confirm the importance of LdPEM1 and LdPEM2 in Leishmania viability. These studies are expected to significantly enhance our knowledge on phosphatidyicholine biosynthesis in Leishmania and on miltefosine's mode of action, and to provide the molecular basis for the development of more potent anticancer drugs. The goal of this project is to gain a better understanding of the mechanism of action of the anticancer drug miltefosine. The proposed studies may have critical implications for the treatment of cancer, and thus, relates directly to public health.