整合素αvβ3介导组织激肽释放酶结合蛋白抗肿瘤起始细胞的作用机制

Tumor initiating cells(TIC) ,also known as cancer stem cells or tumor-propagating cells, as contributors to tumor dormancy, metastasis, and relapse, are highly aggressive and often display drug resistance. TIC represent a subpopulation of highly tumorigenic cancer cells that are capable of anchorage-independence, self-renewal, and multi-lineage differentiation, properties which render these cells particularly resistant to therapy. Integrin αvβ3 is the key molecules that induces TIC producing drug resistance and tumor stemness. Chemotherapy drugs induce cell producing drug resistance, tumor stemness and induce integrin αvβ3 expression highly. Our previous studies have found that tissue kallikrein-binding protein (kallistatin) can combine with integrin αvβ3, therefore we speculate that kallistatin effects TIC through integrin αvβ3 signaling pathway possibly. The project will using Pull down, immunoprecipitation, yeast two hybrid methods to verify that kallistatin combined with TIC’s integrin αvβ3, from the molecular and cellular level; using molecular biology techniques and flow cytometry technique, from molecular, cellular and animal models to study whether integrin αvβ3 mediated kallistatin anti-tumor effects, in vivo and in vitro; then, we will study whether kallistatin intervene TIC proliferation, migration, apoptosis and other processes through the signal transduction pathways mediated by intetgrin αvβ3. Our research hopes to provide a new idea for drug-resistant tumors therapy.

肿瘤起始细胞（TIC）具有高度的侵袭性和干性，是导致肿瘤耐药性的主要细胞，在肿瘤耐药性和自我更新过程中非常重要。整合素αvβ3不仅在TIC高表达，还是化疗过程中肿瘤细胞产生耐药性和干性的关键分子。我们的前期研究发现，组织激肽释放酶结合蛋白（kallistatin）能够与整合素αvβ3相互结合，因此推测kallistatin可能通过整合素αvβ3信号通路对TIC发挥作用。本项目将利用Pull down、免疫共沉淀、酵母双杂交等方法，从分子和细胞水平，验证kallistatin和肿瘤起始细胞的整合素αvβ3相互结合；利用分子生物学、流式细胞分选等技术，从分子、细胞和模型动物三个水平，探讨kallistatin是否通过整合素αvβ3介导的相关信号转导通路，干预TIC的生长、增殖、迁移、凋亡等过程，希望能为抗药性肿瘤的治疗提供新思路。

肿瘤起始细胞是与肿瘤休眠、转移和复发相关的一类重要癌细胞亚群，具有高度致癌性、独立贴壁、自我更新、分化多谱系等特点。TIC 是产生肿瘤耐药性的主要细胞，也是影响erlotinib、lapatinib 等酪氨酸激酶抑制剂类抗癌药物疗效的主要原因。因此，TIC 细胞已经成为抗肿瘤药物首选的靶细胞。TIC 具有肿瘤干细胞的属性，与肿瘤的发生发展、转移和耐药性相关。而整合素αvβ3不仅在TIC高表达，还是在化疗过程中诱导耐药性和干性的关键分子。我们利用肺癌的TIC细胞通过pull down、SRP、免疫共沉淀等技术发现，组织激肽释放酶结合蛋白（Kallistatin）能够与整合素αvβ3相互结合；利用siRNA、抗体封闭等技术将细胞膜表面整合素αvβ3沉默，体内外实验证实整合素αvβ3介导kallistatin抗TIC作用； kallistatin与整合素αvβ3结合从而抑制整合素αvβ3的磷酸化，进而抑制其下游的KRAS与RaIB的激活。另外本文得到kallistatin的另外一个受体核仁素，以血管内皮细胞为模型探讨了核仁素介导kalistatin抗血管生成作用。 本项目为kallisatin抗TIC细胞活性及其具体分子机制奠定了基础，为kallistatin成为抗药性肿瘤的治疗提供新思路。