基于PGRP单克隆抗体的稳定人源化单链抗体改造及其功能研究

小细胞肺癌（Samll Cell Lung Cancer, SCLC）是一种恶性度较高的肿瘤。前胃泌素释放肽（Pro－Gastrin Releasing Peptide, PGRP）是SCLC肿瘤标志物，抗-PGRP人源化单链抗体是SCLC实现抗体靶向诊断和治疗的关键技术。我们前期已完成PGRP重组抗原及其单克隆抗体的研制，发现该抗体靶向药物显著抑制了SCLC的增殖和迁移。本项目拟在此基础上深入分析抗-PGRP单克隆抗体可变区抗原互补决定区结合位点、序列和结构改变等对其稳定性和亲和力的影响，建立抗-PGRP人源化单链抗体基因稳定性表达技术；利用计算机模拟和定点突变技术定向改造单链抗体，获得与131I、99mTc相结合的稳定放射性药物；通过 SCLC小鼠模型分析该靶向放射性药物的功能。研究为开发新型SCLC诊断和治疗药物提供科学数据，同时为人源化单链抗体改造的技术突破提供新的思路。

Small Cell Lung Cancer (SCLC) is characterized by relatively aggressive spread and poorer prognosis. SCLC has been failed for drug development and showed extremely challenging malignancy to treat. Several new therapeutic targets and strategies have shown encouraging results in pre-clinical and early clinical trials, but their utility in clinical practice needs to be determined in randomized trials. Pro-gastrin-releasing Peptide (PGRP) is produced by SCLC in an autocrine fashion. It is a serum precursor peptide of GRP and stable in serum and may be used as a possible tumor marker of SCLC. PGRP was as a marker for monitoring the disease and for the detection of recurrences. Previously, we have prepared the PGRP and obtained monoclonal antibodies against PGRP. The anti-PGRP monoclonal antibody inhibited the proliferation and migration of SCLC cells. In the study, we will clone the gene encoding anti-PGRP monoclonal antibody, analyze the complementarity-determining region (CDR) of anti-PGRP monoclonal antibody, and establish technology for expressing humanized anti-PGRP monoclonal antibodies. The single chain antibody (scFv) will be further optimized by computer simulation and site directed mutation. The radioisotope 131I and 99mTc -conjugated humanized scFv will be created and optimized. The tumor bearing model of nude mice will be prepared. The stability and efficacy of anti-PGRP humanized scFv will be evaluated in vivo. The study will lay the foundation for the development of single-chain radioactive clinical drugs of SCLC detection and diagnosis.