Vascular endothelial growth factor receptor 2 (VEGFR2)/KDR plays a critical role in tumor growth, diffusion, and invasion. The amino acid sequence homology of KDR between mouse and human in the VEGF ligand-binding domain was low, thus the WT mice could not be used to evaluate Abs against human KDR, and the lack of a suitable mouse model hindered both basic research and drug developments. Using the CRISPR/Cas9 technique, we successfully inserted different fragments of the human KDR coding sequence into the chromosomal mouse Kdr exon 4 locus to obtain an hKDR humanized mouse that can be used to evaluate the marketed Ab ramucirumab. In addition, the humanized mAb VEGFR-HK19 was developed, and a series of comparative assays with ramucirumab as the benchmark revealed that VEGFR-HK19 has higher affinity and superior antiproliferation activity. Moreover, VEGFR-HK19 selectively inhibited tumor growth in the hKDR mouse model but not in WT mice. The most important binding epitopes of VEGFR2-HK19 are D257, L313, and T315, located in the VEGF binding region. Therefore, the VEGFR2-HK19 Ab inhibits tumor growth by blocking VEGF-induced angiogenesis, inflammation, and promoting apoptosis. To our best knowledge, this novel humanized KDR mouse fills the gaps both in an animal model and the suitable in vivo evaluation method for developing antiangiogenesis therapies in the future, and the newly established humanized Ab is expected to be a drug candidate possibly benefitting tumor patients.

中文翻译：

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新型 hKDR 小鼠模型描述了针对血管内皮生长因子受体 2 的中和抗体的抗血管生成和促进细胞凋亡作用

血管内皮生长因子受体 2 (VEGFR2)/KDR 在肿瘤生长、扩散和侵袭中起关键作用。小鼠和人KDR在VEGF配体结合结构域的氨基酸序列同源性较低，因此WT小鼠不能用于评估抗人KDR抗体，缺乏合适的小鼠模型阻碍了基础研究和药物研究事态发展。利用CRISPR/Cas9技术，我们成功地将人类KDR编码序列的不同片段插入到小鼠染色体Kdr中外显子 4 基因座以获得 hKDR 人源化小鼠，可用于评估市售的 Ab ramucirumab。此外，还开发了人源化单克隆抗体VEGFR-HK19，以ramucirumab为基准的一系列比较试验表明VEGFR-HK19具有更高的亲和力和优异的抗增殖活性。此外，VEGFR-HK19 在 hKDR 小鼠模型中选择性抑制肿瘤生长，但在 WT 小鼠中没有。VEGFR2-HK19 最重要的结合表位是位于 VEGF 结合区的 D257、L313 和 T315。因此，VEGFR2-HK19 Ab 通过阻断 VEGF 诱导的血管生成、炎症和促进细胞凋亡来抑制肿瘤生长。据我们所知，这种新型人源化 KDR 小鼠填补了动物模型和未来开发抗血管生成疗法的合适体内评估方法的空白，