胸腺免疫抑制性五肽（TIPP）抗哮喘作用及机制研究

Allergic asthma is a complex inflammatory disease, and corticosteroids are the most effective and commonly used drugs for asthma treatment. However, severe toxic and side effects will occur in asthma patients for whom a high dosage or long term use of corticosteroids is needed. Thus there is a need for the research and development of new anti-asthma drugs which have high therapeutic effects and low adverse effects. Thymic immunosuppressive pentapeptide (TIPP) is a novel pentapeptide originally isolated by our group. Experiment results showed that TIPP had inhibitory effects on immune, inflammation and allergic responses, and it showed a specific interaction with Grb2 in our primary study. This project is designed to identify and verify the effects and and mechanisms of TIPP against asthma. The research content of the project includes: examining the effects and mechanisms of TIPP against asthma in mouse models of asthma and verifying Grb2 as the action target of TIPP; studying the binding domain of Grb2 with TIPP; evaluating the interaction specificity between TIPP and Grb2; analyzing the binding affinity and dynamics of TIPP and Grb2; verifying the action mechanism of TIPP as a Grb2 blocking agent; verifying inhibitory activity of the known Grb2 blocking peptides on allergic responses; testing the pharmacokinetics of TIPP in rats; designing and screening of TIPP derivatives with higher activity. The results of this project may find out a new mechanism for asthma treatment and lay the foundation for the deep research and development of TIPP.

过敏性哮喘是复杂的炎症性疾病,糖皮质激素是治疗该类疾病最常用药物,但高剂量或长期用药会产生严重毒副反应,因此研发疗效好、毒副作用小的抗过敏药具有重要意义。胸腺免疫抑制五肽(TIPP)是本课题组分离得到的一个具有免疫抑制、抗炎、抗I型变态反应作用的新肽，且初步发现与Grb2有特异性相互作用。本项目拟研究TIPP的抗哮喘作用及机制。主要研究内容包括：研究TIPP是否具有缓解模型小鼠哮喘的作用及对Grb2相关通路的影响；确定Grb2是否是TIPP作用的靶点；研究Grb2与之结合的结构域；评价TIPP与Grb2结合特异性；分析TIPP与Grb2的亲和力及结合动力学；对TIPP作为Grb2阻断剂发挥作用的机制进行验证；验证已知Grb2肽类阻断剂抗过敏活性；检测TIPP在大鼠体内的药代动力学参数；设计和筛选效果更佳的TIPP衍生物。本研究可能发现一种治疗哮喘的新机制,并为TIPP的深入开发奠定基础。

哮喘是一种高发病率的慢性疾病，分为2型哮喘和严重的非2型哮喘。但目前的一线用药地塞米松一方面具有较大的副作用，另一方面对非2型哮喘缺乏治疗效果。本课题研究了具有抗炎活性的胸腺免疫抑制五肽（TIPP）的抗哮喘作用和机制，并深入探讨了其作用靶点，设计研究了活性更好的衍生物。.本项目取得的主要成果有以下几个方面。. TIPP在小鼠体内药代动力学研究和定量入胞的分析：（1）首次建立TIPP在空白鼠血浆、RBL-2H3细胞裂解液、细胞培养上清液和最后一次的细胞冲洗液四种基质中的LC-MS/MS定量方法；（2）证明TIPP体内半衰期较短；（3）证明TIPP能够进入细胞内部。. TIPP抗哮喘活性研究：（1）证明TIPP在类固醇敏感的2型小鼠哮喘模型中能够减少气道高反应性，缓解炎症并抑制相关免疫细胞的活化；（2）证明TIPP可以改善类固醇不敏感的非2型小鼠哮喘模型的气道高反应性，炎症以及相关免疫细胞的表达和活化异常。. TIPP的靶点筛选和验证：（1）证明AKT1与TIPP有特异性结合；（2）证明TIPP和AKT1蛋白具有较高的亲和性；（3）证明TIPP可以通过靶向AKT1抑制脱颗粒过程中的炎症。. 较好活性的TIPP衍生物的设计与评价：（1）证实设计合成的TIPP衍生物，TIPP二联体T2、柔性肽链接TIPP二联体G-T2和二联体环肽C-T2，在体外均具有优于TIPP的抗过敏活性；（2）证明TIPP衍生肽在抗2型哮喘中表现出较好的体内活性，特别是T2和C-T2；（3）证明TIPP衍生肽比TIPP具有更长的体外血浆半衰期，特别是T2和C-T2。. 本项目的研究结果为治疗2型哮喘和类固醇抵抗的非2型哮喘疾病的新药开发奠定了基础。