Ceramide in angiogenic integrin signaling

描述（由申请人提供）：血管生成，新毛细血管的发芽，是癌症生长所必需的。在小鼠模型中抑制血管生成关键细胞表面受体整合素av-beta3/av-beta5，诱导内皮细胞凋亡，破坏血管生成并抑制脑肿瘤生长整合素av- β 3/av- β 5抑制在癌症治疗中的潜在效用正变得越来越明显，RGDfV是一种整合素功能阻断肽，完成了成人复发性胶质瘤的i期临床试验。尽管av-beta3/av-beta5整合素抑制在癌症治疗，特别是脑肿瘤治疗中的应用越来越多，但这种抑制的分子机制尚不清楚，这妨碍了未来试验中合理设计联合治疗。我们已经表明，整合素抑制增加细胞内神经酰胺，促凋亡脂质第二信使。目前尚不清楚这种神经酰胺的增加是否需要整合素av- β 3/av- β 5抑制的抗血管生成作用。我们最近的数据表明，酸性鞘磷脂酶在这一机制中起关键作用。我们推测，由酸性鞘磷脂酶产生并由整合素- av-beta3/av-beta5抑制诱导的神经酰胺增加是内皮细胞凋亡和RGDfV在体内抗脑肿瘤作用所必需的。我们的具体目的是：1)确定神经酰胺、整合素av-beta3/av-beta5与细胞凋亡的分子相互作用；2)通过颅内小鼠脑肿瘤模型比较RGDfV对WT和ASMase缺陷小鼠的抑制作用。在体外实验中，我们将使用asmase缺陷或正常的内皮细胞涂覆玻璃体连接蛋白，比较它们对RGDfV的神经酰胺和凋亡反应。在体内，我们将在ASMase基因敲除小鼠和野生型幼鼠中植入颅内脑肿瘤，并测定RGDfV或对照处理小鼠在存活、肿瘤生长和血管生成方面的差异。证明在av- β 3/av- β 5整合素介导的内皮细胞凋亡中需要酸性鞘磷脂酶，将是神经酰胺在整合素信号传导中因果作用的第一个证据。整合素信号传导对神经酰胺代谢的要求具有潜在的深远意义，为整合素抑制药物与优化神经酰胺反应的药物联合使用提供了分子基础，从而提高了抗癌效果。

DESCRIPTION (provided by applicant): Angiogenesis, the sprouting of new capillaries, is essential for cancer growth. Inhibition of integrins av-beta3/av-beta5, cell surface receptors critical in angiogenesis, induces endothelial apoptosis, disrupts angiogenesis and inhibits brain tumor growth in a mouse model. The potential utility of integrin av-beta3/av-beta5 inhibition in cancer therapy is becoming apparent with the encouraging responses to RGDfV, an integrin-function-blocking peptide that completed a phase-I clinical trial in adults with recurrent gliomas. Despite the emerging use of av-beta3/av-beta5 integrin inhibition in cancer therapy, especially against brain tumors, the molecular mechanism of this inhibition is poorly understood, precluding rational design of combination therapies for future trials. We have shown that integrin inhibition increases intracellular ceramide, a pro-apoptotic lipid second messenger. It is not known whether this ceramide increase is required for the anti-angiogenic action of integrin av-beta3/av-beta5 inhibition. Our recent data suggest that acid sphingomyelinase is critical in this mechanism. We hypothesize that the ceramide increase, generated by acid sphingomyelinase and induced by integrin- av-beta3/av-beta5 inhibition is required for endothelial apoptosis and for the effect of RGDfV against brain tumors in vivo. Our Specific Aims are: 1) To determine molecular interactions between ceramide, integrins av-beta3/av-beta5 and apoptosis, and 2) To compare the inhibitory effect of RGDfV in WT and ASMase deficient mice using an intracranial mouse brain tumor model. In vitro we will use ASMase-deficient or -competent endothelial cells plated on vitronectin and compare their ceramide and apoptotic responses to RGDfV. In-vivo we will implant intracranial brain tumors in ASMase knockout mice and wild type littermates and determine differences in survival, tumor growth and angiogenesis in RGDfV- or control-treated mice between the groups. Demonstration of a requirement for acid sphingomyelinase in av-beta3/av-beta5 integrin-mediated endothelial apoptosis will be the first evidence for a causal role of ceramide in integrin signaling. Requirement for ceramide metabolism in integrin signaling has potentially far-reaching implications by providing a molecular rationale for combining integrin-inhibiting drugs with agents geared to optimize the ceramide response, resulting in improved anti-cancer efficacy.