Acyl-CoA synthetase ACSVL3 in Malignant Glioma: Metabolism and Oncogenic Cellular

描述（由申请人提供）：恶性胶质瘤占脑肿瘤的很大比例。由于这些肿瘤通常难以治疗，因此需要新的和新颖的治疗方法。ACSVL 3是一种脂肪酸（FA）代谢酶，其水平在人类恶性胶质瘤中高度上调，并在培养的胶质瘤细胞中由致癌受体酪氨酸激酶（RTK）信号转导诱导。ACSVL 3是26种酰基辅酶A合成酶（ACS）之一，其“激活”FA以参与生物合成、降解和调节下游代谢途径。快速肿瘤生长需要高速率的膜脂质合成;这些脂质在致癌细胞质信号传导中也具有关键功能。使用完善的人类胶质瘤临床前模型，我们发现使用RNA干扰的ACSVL 3敲低（KD）降低了人类胶质瘤细胞的体外恶性表型。我们建立了体内胶质瘤异种移植物中ACSVL 3表达与肿瘤发生之间的相关性。我们进一步通过磷脂酰肌醇-3激酶（PI 3 K）/Akt和磷脂酶c-3（PLC-3）/二酰基甘油（DAG）途径建立了ACSVL 3表达与致癌第二信使信号传导之间的关系。我们假设ACSVL 3产生特定的FA-CoA产物，通过（i）产生特定的结构脂质（ii）改变直接参与细胞信号传导的脂质，或（iii）改变参与与特定致癌信号蛋白的膜相互作用的脂质来影响肿瘤发生。基于我们的初步发现，我们还假设靶向ACSVL 3将在治疗恶性胶质瘤中具有治疗价值，并提出以下具体目的：（1）鉴定ACSVL 3缺失对人神经胶质瘤细胞系的体外表型的影响，（2）鉴定ACSVL 3缺失对恶性神经胶质瘤细胞中脂质代谢的影响，（3）阐明ACSVL 3 KD如何改变恶性胶质瘤细胞中的信号转导;（4）确定ACSVL 3缺失如何抑制胶质瘤致瘤性。在目的1中，我们将研究对照和KD人脑胶质瘤细胞中的细胞增殖、凋亡和自噬，以确定ACSVL 3在肿瘤发生中的特异性。这些研究也将扩展到其他胶质瘤模型，可能证明在随后的目标，如内源性表达肿瘤抑制基因PTEN的细胞系有用。ACSVL 3在脂质代谢中的功能对于正常细胞或癌细胞都是未知的。目标2提出了填补这一知识空白的研究，也将确定与恶性表型相关的脂质途径。在目的3中，详细分析了PI 3 K/Akt和PLC-？/将在对照和ACSVL 3 KD神经胶质瘤细胞中进行DAG信号传导途径。在目的4中，将在皮下和颅内异种移植物中探索ACSVL 3 KD对体内肿瘤发生、脂质代谢改变和RTK信号传导的影响。这些研究的结果将建立胶质瘤中ACSVL 3上调的机制基础，查明该酶在致癌RTK信号传导和脂质代谢中的作用，并验证靶向该蛋白在恶性胶质瘤中的治疗潜力。 公共卫生相关性：恶性神经胶质瘤在儿童和成人人群中占脑肿瘤的显著百分比，并且需要新的和新颖的治疗方法。在本申请中，我们将研究脂肪酸代谢的酶，我们已经确定为独特的人类恶性胶质瘤上调。这些研究的结果将提供一个关键的评估机制，这种酶有助于恶性表型，并在神经胶质瘤中靶向这种酶的潜在治疗效益。

DESCRIPTION (provided by applicant): Malignant glioma accounts for a significant percentage of brain tumors. As these tumors are typically refractory to treatment, there is a need for new and novel therapeutic approaches. Levels of ACSVL3, an enzyme of fatty acid (FA) metabolism, were found to be highly upregulated in human malignant gliomas and were induced by oncogenic receptor tyrosine kinase (RTK) signaling in cultured glioma cells. ACSVL3 is one of 26 acyl-CoA synthetases (ACS) that "activate" FAs for their participation in biosynthetic, degradative, and regulatory downstream metabolic pathways. Rapid tumor growth requires high rates of membrane lipid synthesis; these lipids also have key functions in oncogenic cytoplasmic signaling. Using a well-established preclinical model of human glioma, we found that ACSVL3 knockdown (KD) using RNA interference decreased the in vitro malignant phenotype of human glioma cells. We established a correlation between ACSVL3 expression and tumorigenesis in glioma xenografts in vivo. We further established a relationship between ACSVL3 expression and oncogenic second messenger signaling via the phosphatidyl inositol-3 kinase (PI3K)/Akt and phospholipase c-3 (PLC-3)/diacylglycerol (DAG) pathways. We hypothesize that ACSVL3 generates specific FA-CoA products that influence oncogenesis by (i) generating specific structural lipids (ii) altering lipids involved directly in cell signaling, or (iii) altering lipids involved in membrane interactions with specific oncogenic signaling proteins. Based on our preliminary findings, we also hypothesize that targeting ACSVL3 will be of therapeutic value in treating malignant glioma, and propose the following Specific Aims: (1) To identify the consequences of ACSVL3 depletion on the in vitro phenotype of human glioma cell lines, (2) To identify the consequences of ACSVL3 depletion on lipid metabolism in malignant glioma cells, (3) To elucidate how ACSVL3 KD alters signal transduction in malignant glioma cells, and (4) To determine how ACSVL3 depletion inhibits glioma tumorigenicity. In Aim 1 we will investigate cell proliferation, apoptosis, and autophagy in control and KD human glioma cells to determine the specificity for ACSVL3 in oncogenesis. These studies will also be extended to other glioma models that may prove useful in subsequent Aims, such as cell lines that endogenously express the tumor suppressor PTEN. The function of ACSVL3 in lipid metabolism is not known for either normal or cancer cells. Aim 2 proposes studies to fill this gap in knowledge that will also identify lipid pathways that correlate with the malignant phenotype. In Aim 3, detailed analyses of the PI3K/Akt and PLC-?/ DAG signaling pathways in control and ACSVL3 KD glioma cells will be carried out. In Aim 4, effects of ACSVL3 KD on in vivo tumorigenesis, alterations in lipid metabolism, and RTK signaling will be explored, both in subcutaneous and in intracranial xenografts. The results of these studies will establish the mechanistic basis for ACSVL3 upregulation in glioma, pinpoint the role of this enzyme in oncogenic RTK signaling and lipid metabolism, and validate the therapeutic potential of targeting this protein in malignant glioma. PUBLIC HEALTH RELEVANCE: Malignant glioma accounts for a significant percentage of brain tumors in both the pediatric and adult populations, and there is a need for new and novel therapeutic approaches. In this application we will investigate an enzyme of fatty acid metabolism that we have identified as uniquely upregulated in human malignant glioma. Results of these studies will provide a critical assessment of the mechanism by which this enzyme contributes to the malignant phenotype, and the potential therapeutic benefit of targeting this enzyme in glioma.