Successful engineering of a highly potent single-chain variable-fragment (scFv) bispecific antibody to target disialoganglioside (GD2) positive tumors.
Successful engineering of a highly potent single-chain variable-fragment (scFv) bispecific antibody to target disialoganglioside (GD2) positive tumors.
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DOI：
10.1080/2162402x.2016.1168557
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发表时间：
2016-06
期刊：
影响因子：
7.2
通讯作者：
Cheung NK
中科院分区：
文献类型：
--
作者：
Cheng M;Santich BH;Xu H;Ahmed M;Huse M;Cheung NK
由于单链可变片段（scFv）固有的不稳定性和与亲本免疫球蛋白形式的结合不足，从单链可变片段（scFv）中提取强效双特异性抗体仍然很困难。之前，我们描述了一种基于抗GD2小鼠5F11-scFv和抗cd3 huOKT3-scFv （5F11-scBA）的基于scfv的抗双胞脂苷（GD2）的双特异性抗体（scBA）。在本研究中，我们用亲和力更高（13倍）的hu3F8-scFv取代5F11-scFv，形成hu3F8-scBA。通过这种修饰，hu3F8-scBA重定向T细胞杀死GD2（+）癌细胞系，在细胞毒性试验中，其效力（飞摩尔EC50）比5F11-scBA（皮摩尔EC50）高5000倍，即使是针对低GD2密度的靶细胞。此外，通过Ca2+通量和细胞因子释放测量，hu3F8-scBA诱导的t细胞活化比5F11-scBA更强。此外，在体内，在神经母细胞瘤和黑色素瘤异种移植模型中，hu3F8-scBA比5F11-scBA更有效地抑制肿瘤生长和延长小鼠存活。我们得出结论，scBA的功能特性可以通过相对适度的抗原亲和力增加而大大增加。
Engineering potent bispecific antibodies from single-chain variable fragments (scFv) remains difficult due to the inherent instability and insufficient binding of scFv's compared to their parental immunoglobulin format. Previously, we described a scFv-based bispecific antibody (scBA) against disialoganglioside (GD2) based on the anti-GD2 murine 5F11-scFv and the anti-CD3 huOKT3-scFv (5F11-scBA). In this study, we substituted the 5F11-scFv with the higher affinity (13-fold) hu3F8-scFv to form hu3F8-scBA. With this modification, hu3F8-scBA redirected T cells to kill GD2(+) cancer cell lines with up to 5,000-fold higher potency (femtomolar EC50) compared with 5F11-scBA (picomolar EC50) in cytotoxicity assays, even against target cells with low GD2 densities. Furthermore, hu3F8-scBA induced stronger T-cell activation than 5F11-scBA, as measured by Ca2+ flux and cytokine release. Additionally, in vivo, hu3F8-scBA suppressed tumor growth and prolonged mice survival much more effectively than 5F11-scBA, in both neuroblastoma and melanoma xenograft models. We conclude that the functional properties of scBA's can be increased substantially by relatively modest increases in antigen affinity.