抑制肿瘤转移的新靶点: iPLA2在整合素和基质金属蛋白酶再循环中的新颖作用

With increasing incidence and mortality, cancer is the leading cause of death in China and is a major public health problem. The migration of cancer cells away from the primary tumor into surrounding tissue is an important initial step in tumor progression. To date, the major challenge to the inhibition of migration, invasion and metastasis is far from being solved. Enhanced levels of expression of certain integrins and Matrix Metalloproteinases (MMPs), and a consequent increase in specific integrin signals, have been linked to cancer cell metastasis. Although there have been attempts to target integrins and MMPs directly using peptide or nonpeptide antagonist, they were not successful due to cytotoxicity resulted from essential nature of integrins and MMPs for normal cell function. As more evidences on the significance of integrin and MMP recycling in cancer metastasis were presented recently, there is very limited understanding toward ways of controlling integrin trafficking. Our recent studies revealed a novel function of Ca2+-independent phospholipase A2 (iPLA2) that is specifically required for the recycling of c-Src and integrin. This is consistent with recent observations that increased activity and expression of several PLA2 isoforms in various human cancers, suggesting their involvement in development and progression of tumor. A primary goal of this proposal is to understand mechanistic details of how iPLA2 controls the endocytic recycling of integrin αvβ3 and MT1-MMP, which would allow us to find a way to control their trafficking. Our approach to block integrin or MMP recycling by iPLA2 inhibition would give us an economically attractive and less cytotoxic therapeutic option for preventing tumor metastasis.

癌症已经成为在中国死亡的主要原因之一，也成为了最重要的社会医疗问题。 然而 如何抑制肿瘤迁移、侵润和转移这一主要问题还远未解决。研究表明整合素和基质金属蛋白酶（MMPs）的过量表达与恶性肿瘤细胞的发展存在着密切的联系。尽管已有前人进行了针对整合素和MMPs的抑制剂设计，但却因为其具有细胞毒性而失败 。越来越多的研究证明了整合素和MMP再循环对于癌症转移的意义，但是如何对整合素再循环进行控制却鲜为人知。我们最近的研究揭示了钙非依赖性磷脂酶A2（iPLA2）的一项全新功能: iPLA2是c-Src和整合素再循环过程的特异性必要因素. 因而我们的首要目标是探究iPLA2如何控制αvβ3型整合素和 MT1-MMP再循环的细节机理，进而给予我们一条控制后两者运输的全新思路。我们将着眼于利用抑制iPLA2来阻断整合素或MMP的再循环，从而为阻断癌症转移的经济适用且低细胞毒性新途径的建立打下良好的基础。

钙非依赖性磷脂酶A2（第六组磷脂酶A2；iPLA2）催化甘油磷脂在sn-2酯键处水解，形成非酯化脂肪酸和2-溶血磷脂。iPLA2含有三部分特殊结构，在N端包含7-8个锚蛋白重复序列，中部有一个链接区域，而C端则是其催化结构域。研究表明上调iPLA2会导致肿瘤发生，并且大多数的人源黑色素瘤细胞系在mRNA和蛋白水平上都会高表达iPLA2。一些同类的研究也表明MT1-MMP(又称MMP14)是肿瘤细胞进行组织侵袭过程的一个重要组成部分。基于这些研究，我们提出假设：iPLA2活性可能是调节MT1-MMP循环所必需的，这将直接影响肿瘤的侵袭能力。荧光免疫检验和MT1-MMP内吞实验显示，当iPLA2活性被药理抑制剂抑制时，内吞后的MT1-MMP仍保留在核周区域，这一现象说明MT1-MMP的内吞作用并不需要iPLA2活性，但是从内吞循环小室向细胞表面循环却需要其活性。shRNA对iPLA2基因敲除后，iPLA2活性的降低显著抑制了侵袭性伪足的形成并在荧光标记的胶原蛋白胶或基质胶浸润实验中显著抑制了黑色素瘤细胞的浸润能力。同时我们建立了C57/BL6小鼠自发高转移性B16-F10黑素瘤的同系模型，通过在小鼠耳朵背面的皮肤和软骨之间注射癌细胞以研究iPLA2抑制剂对肿瘤浸润的影响。结果显示在没有iPLA2药理抑制剂时，大多数小鼠的锁骨上淋巴结上观察到了转移的黑素瘤。然而，iPLA2抑制剂给药显著阻断了肿瘤转移，这一结论源于对对照组和FK-iPLA2抑制剂给药组的fisher检验。这些结果表明药理抑制剂对iPLA2活性的抑制可以阻断MT1-MMP的循环和侵袭性伪足的形成，以显著降低黑色素瘤的侵袭和转移能力。

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