CDH11+/ITGA5+外泌体介导RUNX2高表达乳腺癌促进骨转移前微环境形成的作用和机制

Although most breast cancer metastases in bone cause osteolytic lesions, both of the reports and our previous study indicate that breast cancer induces pre-metastatic osteogenic niche. The previous study provided evidences that RUNX2 confers cancer cells (seeds) bone chemoattractant and colonization advantages by transcriptional regulation of ITGA5, and experimental clues that RUNX2 high-expressed breast cancer cells induce the formation of pre-metastatic osteogenic niche (soil). CDH11 and ITGA5 were present abundantly in exosomes secreted by RUNX2 high-expressed cancer cells. Further studies revealed that CDH11+ exosomes were uptaken by a set of osteoblast precursors, and ITGA5+ exosomes induced osteoblast differentiation. Thus, we propose a hypothesis: as soon as CDH11+/ITGA5+ exosomes, secreted by RUNX2 high-expressed cancer cells, are uptaken by CDH11 high-expressed osteoblasts, exosome ITGA5 initiates pre-metastatic niche formation by educating pro-osteoblasts toward matured “spy” osteoblasts which can further induce pre-metastatic osteogenic niche formation by instigating the surrounding osteoblasts differentiation. In this study, we attempt to confirm the mediate role of CDH11 in mutual recognition of exosomes and osteoblasts; to investigate the function of ITGA5 in intiating pre-metastatic “spy” osteoblasts; to explore the mechanism of “spy” osteoblasts inducing the pre-metastatic osteogenic niche by instigating the surrounding osteoblasts differentiation; to make clear the facilitating role of pre-metastatic osteogenic niche in osteolytic bone metastasis; and further to elucidate the role and mechanism of breast cancer CDH11+/ITGA5+ exosomes initiates the RUNX2-driven pre-metastatic niche formation in the bone. The results can enrich the “seed and soil” hypothesis, and provide therapeutic targets for anti-bone metastases.

虽然乳腺癌形成的骨转移灶大都为溶骨性病灶，但文献和前期研究表明乳腺癌诱导成骨性转移前微环境形成。前期研究发现RUNX2通过转录调控ITGA5增强“种子”癌细胞在成骨性微环境中的定植能力；且RUNX2高表达癌细胞可以诱导成骨性转移前微环境“土壤”形成。进而基于外泌体的前期研究推测RUNX2高表达癌细胞的CDH11+/ITGA5+外泌体通过血液到达骨组织后被CDH11高表达成骨细胞摄入；ITGA5诱导成骨细胞分化成为“间谍”细胞；“间谍”细胞再驯化周围的成骨细胞形成成骨性转移前微环境。本项目拟证实CDH11介导外泌体与成骨细胞相互识别；探讨RUNX2高表达癌细胞的ITGA5+外泌体诱导“间谍”成骨细胞形成；探索“间谍”细胞促进转移前微环境形成的机制；明确成骨性转移前微环境促进溶骨性骨转移的作用；阐明CDH11+/ITGA5+外泌体介导RUNX2高表达乳腺癌促进骨转移前微环境形成的作用和机制。

乳腺癌具有嗜骨性转移的特点。虽然乳腺癌形成的骨转移灶大都为溶骨性病灶，但文献和前期研究表明乳腺癌诱导成骨性转移前微环境形成。乳腺癌发生器官特异性转移的分子机制目前仍基于“种子—土壤”学说：原发癌向不同远处器官的定向转移能力由肿瘤细胞（种子）的器官特异性转移潜能和远处器官的微环境（土壤）共同决定。前期研究发现RUNX2高表达乳腺癌细胞高表达ITGA5，从而增强“种子”癌细胞在成骨性微环境中的定植能力。本研究采用体外细胞学实验和体内动物学实验发现RUNX2高表达乳腺癌细胞外泌体可以诱导成骨细胞分化形成适宜RUNX2高表达癌细胞定植的成骨性转移前微环境“土壤”。进一步采用蛋白质质谱筛选并用分子生物学方法和细胞生物学方法证实外泌体携带的CDH11和ITGA5是诱导成骨性转移前微环境形成的关键分子：外泌体通过自身携带的CDH11特异性识别CDH11高表达成骨细胞，并被成骨细胞摄入；外泌体ITGA5则诱导摄入外泌体的成骨细胞分化形成“间谍”细胞；“间谍”细胞再驯化周围的CDH11低表达成骨细胞形成成骨性转移前微环境。成骨性转移前微环境中富含BSP、OPN等骨基质蛋白，作为ITGA5的配体募集RUNX2高表达癌细胞至骨组织，进一步协助癌细胞定植并激活破骨细胞形成溶骨性转移病灶。综上，本项目发现了乳腺癌细胞RUNX2通过外泌体诱导转移前微环境“土壤”形成的新功能；创新性提出“间谍”细胞概念和外泌体高效利用模式：癌细胞外泌体先诱导部分成骨细胞形成“间谍”细胞，“间谍”细胞进而驯化周围未摄入外泌体的成骨细胞协同形成转移性微环境；阐明了CDH11 high/ITGA5 high外泌体介导RUNX2高表达乳腺癌诱导成骨性转移前微环境形成的作用和机制。