Construction of a human monoclonal antibody against bFGF for suppression of NSCLC.
Construction of a human monoclonal antibody against bFGF for suppression of NSCLC.
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DOI：
10.7150/jca.24255
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发表时间：
2018
影响因子：
3.9
通讯作者：
Wang H
中科院分区：
文献类型：
--
作者：
Wang S;Qin Y;Wang Z;Xiang J;Zhang Y;Xu M;Li B;Xia Y;Zhang P;Wang H
有令人信服的证据表明，碱性成纤维细胞生长因子（bFGF）和成纤维细胞生长因子受体1（FGFR 1）在非小细胞肺癌（NSCLC）中的过表达驱动肿瘤进展，可作为NSCLC患者的预后生物标志物或治疗靶点。但目前仍缺乏有效的bFGF治疗药物。因此，迫切需要制备抗bFGF的单克隆抗体或了解其作用机制。以前，我们使用杂交瘤技术来生产鼠抗bFGF单克隆抗体（E12）。然而，E12存在异质性和免疫原性的风险。在本工作中，我们通过取代互补决定区（CDR）中或附近的残基，产生了基于E12的三种人源化变体（H1 L1、H2 L2和H3 L3）。此外，我们彻底探索了VH/VL结构域组合，以模拟全长IgG 1抗体使用计算蛋白质设计。选择H3 L3进行进一步研究，因为它表现出最好的人源化和对bFGF的最强亲和力。H3 L3轻链和重链的人源化率分别为100%和98.89%。通过ELISA证实H3 L3的FGF 2中和作用。我们还发现H3 L3可以通过降低AKT和MAPK的磷酸化水平来有效抑制肿瘤的生长和血管生成。此外，H3 L3显着降低了裸小鼠的肿瘤大小和微血管密度。总之，我们的研究表明，H3 L3通过阻碍NSCLC的发展发挥抗肿瘤作用。
Compelling evidence implicates that overexpression of basic fibroblast growth factor (bFGF) and fibroblast growth factor receptor 1 (FGFR1) in non-small cell lung cancer (NSCLC) drives tumor progression, can serve as prognostic biomarkers or therapeutic targets for NSCLC patients. But at present, we still lack of effective drugs for bFGF. The preparation of monoclonal antibodies against bFGF or to understand its mechanism of action is urgently need. Previously, we used hybridoma technology to produce a murine anti-bFGF monoclonal antibody (E12). However, E12 carries risks of heterogeneity and immunogenicity. In the present work, we produced three humanized variants (H1L1, H2L2 and H3L3) based on E12 by substituting residues in or near the complementarity-determining region (CDR). In addition, we thoroughly explored VH/VL domain combinations to simulate full-length IgG1 antibodies using computational protein design. H3L3 was selected for further study, as it demonstrated the best humanization and strongest affinity for bFGF. Specially, humanization of H3L3's light chain and heavy chain were 100% and 98.89%, respectively. The FGF2 neutralizing effect of H3L3 were confirmed by ELISA. We also found that H3L3 can effectively suppress the growth and angiogenesis of cancer through reduce the phosphorylation of AKT and MAPK. Moreover, H3L3 dramatically reduced tumor size and micro-vessel density in nude mice. Altogether, our study demonstrates that H3L3 exerts anti-tumor effects by impeding NSCLC development.
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