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

恶性胶质瘤在脑肿瘤中占很大比例。因为这些肿瘤通常是难治性的 在治疗方面，需要新的和新的治疗方法。脂肪酶ACSVL3的水平 酸(FA)代谢，被发现在人类恶性胶质瘤中高度上调，并由 培养的胶质瘤细胞中致癌受体酪氨酸激酶(RTK)的信号转导。ACSVL3是26个酰基辅酶A之一 “激活”脂肪酸参与生物合成、降解和调节的合成酶(AC) 下游代谢途径。肿瘤的快速生长需要高速率的膜脂合成；这些 脂类在致癌细胞质信号转导中也起着关键作用。使用一个完善的临床前模型 在人脑胶质瘤中，我们发现使用RNA干扰的ACSVL3基因敲除(KD)在体外减少了 人脑胶质瘤细胞的恶性表型。我们建立了ACSVL3表达与 脑胶质瘤异种移植瘤体内致瘤性研究。进一步建立了ACSVL3表达与ACSVL3之间的关系 和致癌的第二信使信号通过磷脂酰肌醇-3激酶(PI3K)/Akt和 磷脂酶C-β(PLC-β)/二酰甘油(DAG)途径。我们假设ACSVL3产生特定的 通过(I)产生特定的结构脂(Ii)改变脂类来影响肿瘤发生的FA-CoA产物 直接参与细胞信号传递，或(Iii)改变与特定细胞膜相互作用的脂类 致癌信号蛋白。根据我们的初步发现，我们还假设以ACSVL3为目标 将在治疗恶性胶质瘤方面具有治疗价值，并提出以下具体目标：(1) 确定ACSVL3缺失对人脑胶质瘤细胞系体外表型的影响，(2) 确定ACSVL3缺失对恶性胶质瘤细胞脂代谢的影响，(3)阐明 ACSVL3KD如何改变恶性胶质瘤细胞的信号转导，以及(4)确定ACSVL3如何 耗竭抑制胶质瘤的致瘤性。在目标1中，我们将研究细胞增殖、凋亡和自噬。 在对照和KD人脑胶质瘤细胞中确定ACSVL3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，这些模型可能被证明在后续的AIMS中有用，例如 内源性表达肿瘤抑制基因PTEN。ACSVL3在脂类代谢中的作用尚不清楚 无论是正常细胞还是癌细胞。Aim 2建议进行研究，以填补这一知识空白，也将识别脂质 与恶性表型相关的通路。在目标3中，详细分析了PI3K/Akt和PLC-？ 将在对照组和ACSVL3KD胶质瘤细胞中进行DAG信号通路的研究。在目标4中， ACSVL3KD在体内肿瘤发生、脂代谢改变和RTK信号转导方面的作用将被探索，两者都 在皮下和颅内异种移植中。这些研究的结果将奠定机理基础 对于胶质瘤中ACSVL3的上调，明确该酶在致癌RTK信号和脂质中的作用 代谢，并验证靶向该蛋白在恶性胶质瘤中的治疗潜力。

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-¿ (PLC-¿)/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.