来自各种类型肿瘤的异质细胞外囊泡（EV）被认为可以诱导引流淋巴结（LN）中转移前“壁龛”的形成，从而促进淋巴转移。为了确定所涉及的 EV 的特定亚群，我们对膀胱癌 (BCa) 组织来源的 EV 进行了高分辨率蛋白质组学分析并结合纳流式细胞术，以确定包含整合素 α6 的肿瘤来源的 EV 的新子集 (ITGA6 EV)并在820例BCa患者的多中心临床分析中揭示了ITGA6 EV与引流淋巴结转移前生态位的形成和淋巴转移呈正相关。 BCa 衍生的 ITGA6 EV 在体内和体外诱导 E-选择素 (SELE) 标记的转移前淋巴管重塑，并通过将负载 circRNA-LIPAR 递送至淋巴内皮细胞，促进引流淋巴结中的转移。从机制上讲，LIPAR将ITGA6连接到RAB5A的开关II结构域并维持RAB5A GTP结合激活状态，从而维持通过内体运输装载LIPAR的ITGA6 EV的产生。 ITGA6 EVs 通过 ITGA6-CD151 相互作用靶向淋巴管，并释放 LIPAR 诱导 SELE 过度表达标记的转移前淋巴管重塑。重要的是，我们构建了工程化的ITGA6 EV来抑制淋巴转移前生态位，从而抑制了临床前模型中的淋巴转移并延长了生存期。总的来说，我们的研究揭示了 BCa 衍生的 ITGA6 EV 介导转移前生态位的机制，并提供了一种基于工程化 EV 的对抗 BCa 淋巴转移的策略。© 2024 作者。 《Journal of Extracellular Vesicles》由 Wiley periodicals LLC 代表国际细胞外囊泡学会出版。

Heterogeneous extracellular vesicles (EVs) from various types of tumours are acknowledged for inducing the formation of pre-metastatic "niches" in draining lymph nodes (LNs) to promote lymphatic metastasis. In order to identify the specific subpopulations of EVs involved, we performed high-resolution proteomic analysis combined with nanoflow cytometry of bladder cancer (BCa) tissue-derived EVs to identify a novel subset of tumour-derived EVs that contain integrin α6 (ITGA6+EVs) and revealed the positive correlation of ITGA6+EVs with the formation of pre-metastatic niche in draining LNs and lymphatic metastasis in multicentre clinical analysis of 820-case BCa patients. BCa-derived ITGA6+EVs induced E-selectin (SELE)-marked lymphatic remodelling pre-metastatic niche and promoted metastasis in draining LNs through delivering cargo circRNA-LIPAR to lymphatic endothelial cells in vivo and in vitro. Mechanistically, LIPAR linked ITGA6 to the switch II domain of RAB5A and sustained RAB5A GTP-bound activated state, thus maintaining the production of ITGA6+EVs loaded with LIPAR through endosomal trafficking. ITGA6+EVs targeted lymphatic vessels through ITGA6-CD151 interplay and released LIPAR to induce SELE overexpression-marked lymphatic remodelling pre-metastatic niche. Importantly, we constructed engineered-ITGA6 EVs to inhibit lymphatic pre-metastatic niche, which suppressed lymphatic metastasis and prolonged survival in preclinical models. Collectively, our study uncovers the mechanism of BCa-derived ITGA6+EVs mediating pre-metastatic niche and provides an engineered-EV-based strategy against BCa lymphatic metastasis.© 2024 The Author(s). Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.