Structural design of disialoganglioside GD2 and CD3-bispecific antibodies to redirect T cells for tumor therapy.
Structural design of disialoganglioside GD2 and CD3-bispecific antibodies to redirect T cells for tumor therapy.
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DOI：
10.1002/ijc.29007
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发表时间：
2015-01-15
影响因子：
6.4
通讯作者：
Cheung NK
中科院分区：
文献类型：
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作者：
Cheng M;Ahmed M;Xu H;Cheung NK
基于抗体的免疫疗法已被证明对高危神经母细胞瘤患者有效。然而，尽管T细胞是最有效的肿瘤杀伤效应物，但由于它们缺乏Fc受体，因此未得到充分利用。利用单价单链片段（ScFv）平台，我们设计了串联ScFv双特异性抗体（BsAbs），特异性靶向肿瘤细胞上的双对话甘脂苷（GD2）和T细胞上的CD3。根据与GD2的结合以及诱导T细胞介导的肿瘤细胞毒性的能力，构建了bsab的结构变异并对其进行了排序。体外热稳定性和结合测量用于表征每个结构，并使用硅分子模型来显示抗gd2 ScFv的可变区重链（VH）和轻链（VL）链的取向如何改变负责高亲和力结合的关键残基的构象。我们发现，当与亲和成熟突变结合时，VH-VL取向、（GGGGS）3连接体、scFv的二硫键稳定提供了最有效的BsAb来指导T细胞裂解GD2阳性肿瘤细胞。在体内，优化后的BsAb能有效抑制黑色素瘤和神经母细胞瘤异种移植物的生长。这些发现提供了一种基于结构的方法的临床前验证，以帮助设计用于t细胞介导治疗的BsAb。
Antibody based immunotherapy has proven efficacy for patients with high risk neuroblastoma. However, despite being the most efficient tumoricidal effectors, T cells are underutilized because they lack Fc receptors. Using a monovalent single chain fragment (ScFv) platform, we engineered tandem scFv bispecific antibodies (BsAbs) that specifically target disialoganglioside (GD2) on tumor cells and CD3 on T cells. Structural variants of BsAbs were constructed and ranked based on binding to GD2, and on competency in inducing T cell mediated tumor cytotoxicity. In vitro thermal stability and binding measurements were used to characterize each of the constructs, and in silico molecular modeling was used to show how the orientation of the variable region heavy (VH) and light (VL) chains of the anti-GD2 ScFv could alter the conformations of key residues responsible for high affinity binding. We showed that the VH-VL orientation, the (GGGGS)3 linker, disulfide bond stabilization of scFv, when combined with an affinity matured mutation provided the most efficient BsAb to direct T cells to lyse GD2 positive tumor cells. In vivo, the optimized BsAb could efficiently inhibit melanoma and neuroblastoma xenograft growth. These findings provide preclinical validation of a structure-based method to assist in designing BsAb for T-cell mediated therapy.