miR-185调控PYK2-STAT5A通路在弥漫性大B细胞淋巴瘤CAM-DR中的作用及机制

Cell adhesion-mediated drug resistance (CAM-DR) is thought to be one of the major causes of chemotherapy failure in patients with diffuse large B-cell lymphoma (DLBCL). Dysregulation of apoptosis is a major mechanism responsible for the CAM-DR phenotype. Previous studies have reported that proline-rich tyrosine kinase 2 (PYK2) and signal transducer and activator of transcription 5A (STAT5A) play crucial roles in the regulation of cell apoptosis. Our preliminary experiment demonstrated that miR-185, PYK2 and STAT5A were all significantly correlated with the CAM-DR phenotype of DLBCL cells. Moreover, miR-185 could modulate the chemotherapeutic sensitivity of DLBCL cells by targeting PYK2. Furthermore, PYK2 was found to interact with STAT5A. Based on these, we infer that cell adhesion mediated-miR-185 downregulation may enhance STAT5A activation and its downstream apoptotic-related genes expression via augmenting PYK2 expression, thereby promoting CAM-DR phenotype. To verify this assumption, we first intend to investigate the effect of PYK2-STAT5A interaction on CAM-DR. Subsequently, we will analyze the effect of miR-185 on PYK2 expression and CAM-DR. Lastly, the clinical tissues and xenograft nude mouse model will be utilized to further evaluate the effect of miR-185 on PYK2-STAT5A axis and the CAM-DR phenotype. Our study may provide insights into the mechanism of CAM-DR in DLBCL.

细胞粘附介导的耐药（CAM-DR）是弥漫性大B细胞淋巴瘤（DLBCL）化疗失败的重要原因。细胞凋亡调节失控是CAM-DR的重要促成因素。富含脯氨酸的酪氨酸激酶2（PYK2）及信号转导与转录激活因子5A（STAT5A）在调节肿瘤细胞凋亡中有重要作用。课题组预实验发现miR-185、PYK2、STAT5A均与CAM-DR有关，miR-185可靶向抑制PYK2，且PYK2可与STAT5A相互作用。据此推测：细胞粘附引起miR-185下调可能通过增强PYK2的表达促进STAT5A活化，继而影响下游凋亡相关蛋白表达，促进CAM-DR进程。课题组拟首先分析PYK2与STAT5A相互作用对CAM-DR的影响；接着分析miR-185调节PYK2对CAM-DR的影响；最后结合临床样本及裸鼠模型综合分析miR-185调节PYK2-STAT5A信号轴对CAM-DR的影响。本研究将进一步阐明CAM-DR分子机制。

细胞凋亡调节失控是细胞粘附介导的耐药（CAM-DR）产生的重要原因。根据前期工作基础，我们提出假说：细胞粘附引起miR-185下调可能通过增强PYK2的表达促进STAT5A活化，继而影响下游凋亡相关蛋白表达，促进CAM-DR进程。首先，我们证实淋巴瘤细胞OCI-Ly8及OCI-Ly10粘附到纤连蛋白(FN)可引起miR-185下调以及PYK2的活化。接着，我们发现，miR-185可直接靶向调控PYK2，细胞粘附介导的miR-185下调可通过诱导PYK2活化促进CAM-DR。而后我们发现FAK/PYK2抑制剂PF-562271可部分消除miR-185下调介导的CAM-DR。此外，我们还发现，淋巴瘤细胞粘附到FN可显著增强STAT5A总蛋白表达水平，并促进STAT5A第694位酪氨酸磷酸化；过表达STAT5A可促进CAM-DR。PYK2可通过与STAT5A相互作用增强STAT5A的表达，并促进STAT5A第694位酪氨酸磷酸化。项目的研究成果将为进一步揭示CAM-DR发生发展的分子机制提供实验数据。