lung tumor exosome-mediated inflammation promotes tumor progression and growth

本研究的总体目标是阐明肺肿瘤外泌体促进炎症反应，即巨噬细胞的激活，促进肺肿瘤进展和生长的机制。我们的初步数据表明，用TC-1肺肿瘤细胞产生的外泌体或用致癌物氨基甲酸乙酯预处理的NJ小鼠肺组织分离的外泌体预处理A/J小鼠，肿瘤生长更快，肺癌进展更早。聚氨酯处理导致活化形式的TRAF2募集到外泌体，具有活化TRAF2的外泌体被巨噬细胞的骨髓来源的前体所吸收，导致其成熟并随后迁移到肺部。激活的TRAF2募集到外泌体需要降解肿瘤细胞中的炎症抑制蛋白CYLD；此外，TRAF2可以介导CYLD泛素化，从而促进其降解。然而，siRNA TRAF2敲除的结果表明，TRAF2是CYLD泛素化所必需的，但不是充分的。我们现在已经确定了两种与TRAF2相互作用的外泌体蛋白，Itch和Jabi，它们有可能增强肺肿瘤细胞中CYLD的泛素化。我们建议：(1)通过确定敲除TC-1肺癌细胞中的TRAF2是否足以抑制巨噬细胞分化和（ii）预防TC-1细胞外泌体介导的聚氨酯诱导肺癌的增强，来证实肺癌外泌体TRAF2在促进聚氨酯诱导肺癌中的作用。(2)确定TC-1外泌体中的Jabi和itch是否能够促进CD1 1bGr1细胞向活化的巨噬细胞分化。(3)外泌体在体内靶向细胞的鉴定。所产生的数据应该允许开发一个高度创新的外泌体介导的慢性炎症和促进肺肿瘤生长的细胞和分子基础模型，并提出新的预防和治疗策略。

The overall goal of this proposal is to elucidate the mechanisms by which lung tumor exosomes promote an inflammatory response, namely the activation of macrophages, and promote the progression and growth of lung tumors. Our preliminary data indicate that pretreatment of A/J mice with exosomes produced by TC-1 lung tumor cells or exosomes isolated from the lung tissue of NJ mice that have been pretreated with the carcinogen urethane results in more rapid tumor growth and earlier progression of lung carcinomas. Urethane treatment results in the recruitment of the activated form of TRAF2 to the exosomes, and the exosomes with activated TRAF2 are taken up by the bone marrow-derived precursors of macrophages, leading to their maturation and subsequent migration to the lung. The recruitment of the activated TRAF2 to the exosomes requires degradation of the inflammation suppressor protein, CYLD, in the tumor cells; moreover, TRAF2 can mediate ubiquitination of CYLD thereby promoting its degradation. The results of siRNA TRAF2 knockout indicated, however, that TRAF2 is required but is not sufficient for ubiquitination of CYLD. We have now identified two exosomal proteins that interact with TRAF2, Itch and Jabi, which have the potential to enhance the ubiquitination of CYLD in lung tumor cells. We propose to: (1) Confirm the role of lung tumor exosomal TRAF2 in the promotion of urethane-induced lung carcinomas by determining whether knockout of TRAF2 in TC-1 lung tumor cells is sufficient for (0 inhibition of macrophage differentiation and (ii) prevention of TC-1 cell exosome-mediated enhancement of urethane induced lung cancer. (2) Determine if Jabi and itch in the TC-1 exosomes are capable of promoting the differentiation of CD1 1bGr1 cells into activated macrophages. (3) Identification of cells targeted in vivo by exosomes. The data generated should permit the development a highly innovative model of the cellular and molecular basis for exosome-mediated chronic inflammation and promotion of lung tumor growth and suggest novel preventive and therapeutic strategies.