Monosaccharide Analogs, Sialic Acid Metabolism and Apoptosis

描述(申请人提供)：用外源提供的单糖补充糖基化途径对从罕见的先天性疾病(CDGs)到癌症的各种人类疾病具有潜在的治疗价值。特别是，N-乙酰甘露糖胺(ManNAc)类似物可以调节细胞表面唾液酸的显示，是解决广泛癌症中发现的糖基化缺陷的有希望的候选者。糖类似物从目前的实验室工具到临床应用的转变最近得到了推动，因为我们实验室最近发现，糖类似物在用羟基保护基团修饰单糖以改善其药理性质时所观察到的毒性是由于细胞凋亡。这一观察结果，再加上作为癌症控制关键靶点的凋亡刺激物的出现，支持了加强研究唾液酸和细胞凋亡之间的联系的努力：具体目标1是表征ManNAc类似物在人类癌细胞凋亡中的作用，重点是N-酰基“R1”基团的作用。这一目标将通过以下方式实现：(A)测试类似物对O-GlcNAc蛋白修饰的影响；(B)逐步评估执行细胞凋亡的各种细胞途径对类似物的反应；(C)测试类似物对细胞用来构建一套参与凋亡的唾液糖结合物(包括Fas、TNFalphaR2、IGF和CD43)的生物合成机制的影响；以及(D)评估通过唾液酸生物合成途径的代谢流抑制是类似物诱导的细胞凋亡的原因还是下游反应。Aim#1中概述的一系列实验旨在全面评估ManNAc类似物对唾液酸代谢的影响，并将这些反应与细胞凋亡联系起来。这些化合物还通过O-羟基“R2”基团的修饰而引起细胞范围的反应。在本项目中，将在目标2中测试R2-基团的细胞响应，R2-基团在摄入后向细胞输送短链脂肪酸(SCFA)：具体目标#2是通过(A)分析细胞对模拟提供的短链脂肪酸(SCFA)的反应，(B)表征SCFA‘载体’分子的作用，(C)分析模拟处理细胞中神经酰胺和神经节苷脂的代谢，以及(D)测试R2-基团的代谢‘携带’。

DESCRIPTION (provided by applicant): The supplementation of glycosylation pathways with exogenously-supplied monosaccharide has potential therapeutic value for human diseases ranging from rare congenital disorders (CDGs) to cancer. In particular, N-acetylmannosamine (ManNAc) analogs that modulate cell surface sialic acid display are promising candidates for addressing glycosylation defects found in a wide range of cancers. The transition of sugar analogs from their current status as laboratory tools to clinical applications has recently been given a boost by our laboratory's recent finding that sugar analog-induced toxicity observed upon modification of monosaccharide with hydroxyl-protecting groups designed to improve their pharmacological properties is due to apoptosis. This observation, combined with the emergence of apoptosis stimulators as key targets for the control of cancer, support enhanced efforts to investigate the connections between sialic acid and apoptosis: Specific Aim #1 is to characterize the role of ManNAc analogs in apoptosis in human cancer cells, with a focus on the effects of the N-acyl "R1" group. This goal will be pursued by (A) testing the impact of the analogs on O-GlcNAc protein modification; (B) performing a step by step evaluation of the response of the various cellular pathways that execute apoptosis to analogs; (C) testing the effects of analogs on the 'biosynthetic machinery' used by cells to construct a set of the sialoglycoconjugates involved in apoptosis that include Fas, TNFalphaR2, IGF, and CD43; and (D) evaluating whether inhibition of metabolic flux through the sialic acid biosynthetic pathway is the cause, or a downstream response, to analog-induced apoptosis. The set of experiments outlined in Aim #1 are designed to comprehensively evaluate the effects of ManNAc analogs on sialic acid metabolism and link these responses to apoptosis. These compounds also elicit cell-wide responses by virtue of O-hydroxyl "R2" group modifications. In this project, cellular responses of R2-groups, which deliver short chain fatty acids (SCFAs) to a cell upon intake, will be tested in Aim #2: Specific Aim #2 is to characterize the role of R2-groups in apoptosis by (A) analyzing cellular response to analog-supplied SCFAs, (B) characterizing the effects of the SCFA 'carrier' molecule, (C) analyzing ceramide and ganglioside metabolism in analog-treated cells and (D) testing metabolic 'carry-through' of R2-groups.