Metastatic Units 介导卵巢癌腹腔转移的分子机制及靶向阻遏

Peritoneal dissemination and massive ascites are the hallmarks of high-grade serous ovarian cancer (HGSOC) that distinguish it from its low-grade counterpart (LGSOC), however, the mechanisms underlying this discrepant biological behavior remain unclear. To complete peritoneal metastases, ascitic tumor cells (ATCs) must escape from the primary tumor site as either single cells or spheroids, adhere to the mesothelial layer covering the abdominal cavity, and subsequently invade into the favored extracellular matrix (ECM)-rich compartment. Unfortunately, little attention has been devoted to analyzing phenotypic alterations in ATCs throughout HGSOC progression, much less for investigating the metastatic cascade of LGSOC patients. Considering the crucial role of ATCs during ovarian cancer peritoneal metastasis, we sought to performed a comparative analysis of tumor epithelial cells isolated from matched primary tumors, ascites, and metastases of HGSOC and LGSOC patients. We uncovered that ATCs with ITGA5 high phenotype are a distinguishing characteristic of HGSOC and was propagated by an intimate interaction with cancer-associated fibroblasts（CAFs）in compact heterotypic spheroids referred to as metastatic units (MUs). More specially, our results demonstrate that the discriminative effects of ascites stromal CAFs on tumor cells and the resultant MU architecture, as well as disparate ITGA5 expression in ATCs, might explain the distinct differences of peritoneal metastasis in HGSOC and LGSOC. Therefore, in the preliminary unpublished data, we hypothesize that ITGA5 high phenotype was maintained by their interactions with CAFs in metastatic units (MUs) that form within the malignant ascites. Further studies need to figure out how this MUs developed and how to affect the peritoneal metastasis of ovarian cancer. Furthermore, we should clarify how this specialized MUs microenvironment amplifies tumor-stroma signaling to promote ovarian cancer cell survival and metastasis. Also, targeting CAFs to disrupt MUs and hamper ovarian cancer peritoneal propagation strategy need to be considered. Actually, we previously constructed the novel oncolytic adenovirus which combined the characteristics of oncolysis with targeted gene amplication. This tumor-selective replication of the E1A mutant adenovirus would activate the native adenovirus E3 promoters to express the shFAP preferentially in CAFs and knockdown the FAP protein, resulting in depletion of MUs and ultimate ovarian cancer peritoneal metastasis. If this project is completed on schedule, we believe it will shed the new light on the research of ovarian cancer peritoneal metastasis, span the key technique barrier in adenovirus gene therapy and ultimately provide a new strategy to prevent the metastasis of ovarian cancer.

腹腔广泛转移是卵巢高级别浆液癌有别于低级别浆液癌的标志，但其内在机制尚不明了。前期研究中，课题组从寻找临床实际存在的卵巢高、低级别浆液癌腹腔转移差异的分子机制着手，利用反应临床真实状况的配对样本的高通量测序分析，结合腹水微环境的流式细胞分选和PDTX模型体内验证结果，大胆提出执行卵巢高级别浆液癌腹腔播散的功能是腹水中CAFs与ATCs结合形成的异质性球体MUs这一创新概念，揭秘卵巢癌腹水大的微环境中CAFs与ATCs间的“cross-talk”形成MUs小的微环境，来维持ATCs的ITGA5 high 表型导致卵巢癌腹腔转移的复杂机制，进而以活化CAFs的关键分子FAP为切入点，以困扰临床治疗实际的卵巢癌腹腔转移为突破口，有的放矢地构建以FAP为靶点，靶向CAFs的新型选择性复制溶瘤病毒，开展针对卵巢癌腹腔转移基因治疗的新探索，从而为解决卵巢癌治疗棘手的转移问题提供理论和实践先导。

高级别浆液卵巢癌死亡率高一个主要原因是早期发生腹腔转移播散，相比而言，低级别卵巢癌播散转移更慢，预后更好。尽管二者之间已有明确的突变差异，但这两者不同播散转移能力进一步的分子和生物学基础并不清楚。以往研究表明，腹水中单个肿瘤细胞或多细胞球体是卵巢癌播散转移的重要环节，但其参与这种高转移潜能的机制其实并不清楚。我们的研究首次发现高级别浆液性卵巢癌腹水中肿瘤细胞与CAFs形成一个特殊的高转移潜能的结构和功能单位，并命名为转移单元。.我们通过分离原代高级别浆液性卵巢癌的配对原位、转移和腹水细胞并发现腹水肿瘤细胞转移能力最强。转录组测序发现高级别卵巢癌腹水肿瘤细胞ITGA5高表达与其高转移能力有关。通过比较高低级别卵巢癌腹水细胞结构和成分，我们首次发现高级别浆液性卵巢癌腹水中肿瘤细胞与CAFs形成高转移潜能的异质性球体，并命名为转移单元。进一步细胞因子芯片分析发现转移单元中肿瘤细胞ITGA5高表达是通过CAFs依赖的EGF分泌实现的。CAFs与肿瘤细胞的相互作用使肿瘤细胞ITGA5维持高表达并进一步使转移单元具有更强的转移潜能并参与卵巢癌的早期腹膜转移。相反，低级别浆液性卵巢癌缺乏CAFs及其构成的转移单元，一定程度上解释了两者临床进展和预后的不同。这种生物结构的发现对阐明高级别卵巢癌转移机制和针对该结构设计新型临床诊疗策略具有重要意义。.在此基础上，我们进一步构建了构建新型溶瘤腺病毒Adv5/dE1A-Bim/tBid拟从治疗角度清除卵巢癌腹腔肿瘤病灶，探索一种靶向肿瘤转移治疗的新途径，提升腺病毒基因治疗的临床应用价值。.