双特异性抗体mAb04-MICA对人白血病细胞K562的体内外抗肿瘤活性

探讨双特异性抗体mAb04-MICA对人白血病细胞K562的体内外抗肿瘤活性。前期研究显示，mAb04-MICA具有抗血管生成和重塑肿瘤微环境中的免疫监视作用，体外对K562细胞具有良好的抗肿瘤作用。通过ELISA鉴定mAb04-MICA 对抗原VEFGR2和受体NKG2D的亲和力；CCK8法分析mAb04-MICA对K562的体外增殖抑制作用；采用流式细胞术分析mAb04-MICA对鼠源VEGFR2的交叉反应能力；建立K562皮下荷瘤BALB/c裸鼠模型，通过测定瘤体积、称量瘤重、观测荷瘤鼠生存期等方法检测目的抗体的体内抗肿瘤活性；免疫组化方法测定其对肿瘤组织本身及组织内新生血管生成的影响。结果显示:mAb04-MICA对VEGFR2和NKG2D都具有良好的亲和力，并且在体外能够特异性地抑制VEGF诱导的K562细胞的增殖；mAb04-MICA与鼠源VEGFR2具有较高的结合能力；相同给药剂量下，双特异性抗体抑瘤作用显著优于母体单抗，且能延长荷瘤裸鼠生存期，其中Ki-67、p-VEGFR2、VEGF及CD34的表达量显著降低，提示mAb04-MICA可通过特异性地阻碍瘤组织VEGFR2的磷酸化而控制瘤体新生血管的生成，在治疗白血病方面具有潜在的应用价值。

Antitumor efficacy of bispecific antibody mAb04-MICA to human leukemia cell K562 in vitro and in vivo

This study aimed to investigate the efficacy of a bispecific antibody mAb04-MICA on human leukemia cell K562 both in vitro and vivo. mAb04-MICA was previously found to posses excellent anti-angiogenic activity, and had the ability to recruit immune surveillance in tumor microenvironment. In this study, the affinity of mAb04-MICA to VEGFR2 and NKG2D was identified by ELISA. CCK8 was used to detect the effect of mAb04-MICA on K562 proliferation. The cross reactivity of mAb04-MICA to murine VEGFR2 was determined by flow cytometry assay. To evaluate the antitumor activity of mAb04-MICA, tumor volume, tumor weight and the survival of K562 tumor-bearing nude mice were analyzed. The anti-angiogenic activity was determined by immunohistochemistry. The results indicated that mAb04-MICA could target to VEGFR2 and NKG2D, and inhibit K562 proliferation specifically. Besides, mAb04-MICA showed high binding capacity to murine VEGFR2. The bispecific antibody exhibited superior antitumor efficacy to the maternal monoclonal antibody and prolonged the survival of tumor-bearing mice. The expression of Ki-67, p-VEGFR2, VEGF and CD34 in mAb04-MICA treated group was significantly reduced. The results indicated that mAb04-MICA could attenuate the phosphorylation of VEGFR2 and impair angiogenesis of the tumor microenviroment. Therefore, mAb04-MICA could be further developed as a potential tumor targeted immunotherapeutic agent for leukemia.