活化B细胞（ABC）型弥漫大B细胞淋巴瘤中NF-κB/YY1/miR-181a/b反馈通路研究

Diffuse large B-cell lymphoma (DLBCL) remains an incurable disease,despite of the recent developments in pathological classification and clinical treatment field. Multiple mechanisms are involved in apoptosis-resistance of active B cell (ABC) like DLBCL cells, among which YY1 is an important regulatory factor involved in NF-κB signaling pathway. YY1 plays a crucial role in regulating NF-κB activity and involved in survival and drug-resistance of DLBCL. Our preliminary experiment displayed abnormal expression of miRNA-181a/b and their precursor may be potential targets of NF-κB/YY1 in DLBCL cells , transfection of miR-181a/b mimics will increase the sensitivity to immunochemotherpy of DLBCL cells in vitro, which indicated that there is potential feedback regulation involving NF-κB/YY1/miR-181a/b pathway regulating biologic characteristics of DLBCL. The purpose of our study is to verify the hypothesis: ①To determine miR-181a/b and YY1 expression in large-scale samples of DLBCL patients and their prognostic value; ② To confirm the interaction between YY1 and miR-181a/b and explore the exact function of NF-κB/YY1/miR-181a/b pathway in DLBCL cells; ③ To predict direct interaction between miR-181a/b and DLBCL associated targets through bioinformatics database, and further verify it by employing dual-luciferase reporter assay system; ④To investigate potential targeted therapy basing on NF-κB/YY1/miR-181a/b feedback signaling pathway.We expect to confirm and elucidate the exact role of NF-κB/YY1/miR-181a/b feedback circuitry in the development of DLBCL,which will provide a novel direction of DLBCL diagnosis and treatment paradigm.

活化B细胞（ABC）型弥漫大B细胞淋巴瘤（DLBCL）常伴NF-κB/YY1激活,进而介导细胞增殖耐药，治疗反应差。申请人前期研究发现ABC型DLBCL存在miR-181a/b低表达，其前体可能是NF-κB/YY1的转录抑制靶标，同过表达miR-181a/b可能通过抑制YY1、CARD11等关键基因反向调节NF-κB通路，提高DLBCL细胞对免疫化疗的敏感性。本研究拟针对此假说开展以下研究：①研究DLBCL中miR-181a/b及YY1表达及预后意义；②明确ABC型DLBCL中NF-κB/YY1/miR-181a/b相互作用方式；③鉴定与ABC型DLBCL发生发展密切相关的miR-181a/b靶基因；④在细胞水平和模式生物水平进行NF-κB/YY1/miR-181a/b通路及功能研究。通过研究阐明NF-κB/YY1/miR-181a/b通路在ABC型DLBCL发生发展及耐药中的机理。

活化B细胞（ABC）型弥漫大B细胞淋巴瘤（DLBCL）常伴NF-κB激活，进而介导细胞增殖耐药，治疗反应差。我们前期研究发现ABC型DLBCL存在miR-181a/b低表达，其前体可能是NF-κB/YY1的转录抑制靶标，同过表达miR-181a/b可能通过抑制YY1、CARD11等关键基因反向调节NF-κB通路，提高DLBCL细胞对免疫化疗的敏感性。本研究针对此假说开展以下研究：①研究DLBCL中miR-181a/b及YY1表达及预后意义；②明确ABC型DLBCL中NF-κB/YY1/miR-181a/b相互作用方式；③鉴定与ABC型DLBCL发生发展密切相关的miR-181a/b靶基因；④在细胞水平和模式生物水平进行NF-κB/YY1/miR-181a/b通路及功能研究。通过研究阐明NF-κB/YY1/miR-181a/b通路在ABC型DLBCL发生发展及耐药中的机理。更针对性地进行靶向治疗以提高临床疗效。经过3年的研究工作，我们发现：DLBCL患者miR-181a表达较正常人有不同程度下降，miR-181a表达与患者PFS、OS相关。双荧光素酶报告实验及western实验显示miR-181a/b对CARD11均有明显的抑制作用，流式Ａnnexin/PI双染法显示miR-181a过表达组早期凋亡和晚期凋亡率均高于miR-181a+CARD11转染组及转染对照序列组，裸鼠成瘤实验显示注射miR-181a后较注射对照或注射miR-181a+CARD11的肿瘤显著缩小。miR-181a 可通过负性调节CARD11调控NF-kB通路，靶向抑制miR-181-CARD11可显著抑制DLBCL肿瘤形成，经过本项目研究，阐明了miR-181-CARD11通路在DLBCL中的作用机制，并在细胞和模式动物水平明确了靶向抑制miR-181-CARD11可显著抑制DLBCL肿瘤形成，从而为完善DLBCL发病机理和潜在的分子靶向治疗提供依据。