淋巴细胞特异性转录因子Aiolos在肺癌转移中的功能和调控研究

Parenchymal cells have strong anchorage requirements which prevent tissue disorganization and restrict long-range movement of differentiated cells. Pathologic loss of such requirements is required for metastatic behavior in transformed cancer cells. Our preliminary data showed that adaptor protein, p66Shc, localizes to focal adhesions, reports cell detachment and induces anoikis. p66Shc is frequently repressed in metastatic human lung cancer cells and when re-expressed, it completely blocks metastasis of Lewis Lung Carcinoma cells （which do not express p66Shc） in mice, indicating its ability to act as a powerful metastasis suppressor in vivo. Interestingly, the lymphocyte specific zinc finger transcription factor Aiolos, which is coded by IKZF3, is ectopically expressed in solid tumors and plays a critical role in suppressing p66Shc expression. In addition, the expression of Aiolos in human lung cancer tissues predicts survival disadvantage. These preliminary data provide a potential link between hematopoietic specific regulatory network and anchorage independent behavior in metastatic cancer. Here, the objective of this proposal is to systematically focus on the function and regulation of Aiolos in human solid tumors derived from lung using multiple techniques including RNA interference, immunohistochemistry, 3C, matrigel 3D culture, single cell RT-PCR and CHIP. Specifically, we will dissect the role of ectopic expression of Aiolos in cell adhesion related behavior and its mechanism, the connection between Aiolos expression level with distal metastasis of lung cancer, and the environmental factors that contribute to the ectopic expression of Aiolos. Hopefully we may gain significant insight into the change of regulatory network associated with tumorigenesis and metastasis in solid tumors. Also, a better understanding of the mechanisms underlying anoikis resistance could help to prevent solid tumor progression and metastasis.

逃逸失巢凋亡是实体肿瘤细胞转移过程中的重要分子生物学事件之一，我们前期工作表明，正常表皮细胞中定位于粘附斑的衔接蛋白p66Shc能够在细胞脱离基质时感受张力变化启动失巢凋亡，该蛋白在肺癌细胞中的表达缺失是导致肺癌细胞发生转移的主要因素，而生理状态下仅表达于淋巴细胞的转录因子Aiolos在肺癌细胞中的异位表达是抑制p66Shc转录的原因，这一发现暗示着实体肿瘤细胞很可能通过获得造血细胞特有蛋白而改变自身的特性包括对失巢凋亡的敏感性，进而影响肿瘤的发生发展和侵袭转移。本项目将利用细胞三维培养、单细胞RT-PCR、3C等多种技术研究Aiolos在肺癌发生发展中的功能和调控，包括Aiolos异位表达对细胞粘附相关特性的影响及机理以及与临床肺癌远端转移的相关性、并探讨导致Aiolos在肺癌细胞中异位表达的表观遗传机制。研究结果将有助于理解实体肿瘤转移的分子机制,为实体肿瘤的靶向诊疗提供实验基础。

失巢凋亡是指依赖胞外基质贴附生长的细胞在脱离基质后引发的细胞凋亡，对维持正常机体组织稳态和防止异位细胞生长起重要作用。失巢凋亡抵抗是具有转移潜能的实体肿瘤细胞必须具备的重要生物学特性之一。然而，关于可转移的实体肿瘤细胞通过何种机制逃逸失巢凋亡并不清楚。.我们比较了不同转移能力的肺癌细胞系的转录谱，发现转移能力强的小细胞肺癌细胞中表达正常情况下仅表达于淋巴细胞的转录因子Aiolos，Aiolos的生理功能是促进B淋巴细胞离开中枢免疫器官（骨髓），分散至外周血。我们利用免疫组化方法研究了Aiolos在临床肺癌组织样本中的表达情况，发现Aiolos在多种实体肿瘤细胞中表达，Aiolos的高表达预示着肿瘤病人较短的存活期。随后我们在不表达Aiolos的肿瘤细胞中过表达Aiolos，通过鼠尾静脉导入小鼠体内，发现Aiolos的表达能够促进肿瘤细胞的转移。进一步研究发现Aiolos能够下调一系列细胞粘附相关蛋白的表达，破坏肿瘤细胞-细胞间连接、肿瘤细胞-胞外基质间粘附，促进肿瘤细胞从表皮细胞片层结构中游离，并通过破坏失巢凋亡关键蛋白p66Shc基因染色质高级结构来抑制p66Shc基因转录，使得游离的肿瘤细胞产生失巢凋亡抵抗，最终促进肿瘤细胞发生远端转移。这些研究结果将调控实体肿瘤转移的转录调控机制与淋巴细胞发育的转录调控机制联系起来，并提出实体肿瘤细胞通过“co-opt”淋巴细胞转录调控途径，获得淋巴细胞的某些特性（如失巢凋亡抵抗等），实现远端扩散的新观点。.这一结果发表在Cancer Cell，IF=23.523，该工作被选为当期的featured article。Cancer Cell还配发了失巢凋亡领域奠基人西维吉尼亚大学Steven Frisch教授撰写的preview进行点评，Frisch教授高度评价我们的工作“创新性的证明了肿瘤细胞能够利用其他细胞调控程序获得失巢凋亡抵抗特性”。