Meng Jie, Liu Yuanyi, Gao Shuying, Lin Stephen, Gu Xinbin, Pomper Martin G, Wang Paul C, Shan Liang
a Molecular Imaging Laboratory; Department of Radiology; Howard University ; Washington , DC USA.
b Angimmune LLC ; Rockville , MD USA.
Cancer Biol Ther. 2015;16(12):1764-74. doi: 10.1080/15384047.2015.1095403.
EGFR and EGFRvIII are overexpressed in various types of cancer, serving as optimal targets for cancer therapy. Capitalizing on the high specificity of humanized antibody 806 (mAb806) to the EGFR and EGFRvIII overexpressed in cancer, we designed and generated a bivalent recombinant immunotoxin (RIT, DT390-BiscFv806) by fusing the mAb806-derived bivalent single-chain variable fragment with a diphtheria toxin fragment, DT390. In vitro, DT390-BiscFv806 efficiently internalized into the cells and exhibited high cytotoxicity against the U87 glioblastoma cells and the EGFRvIII-transfected U87 (U87-EGFRvIII) cells with a half maximal inhibition concentration (IC50) of 1.47 nM and 2.26 × 10(-4) nM, respectively. Notably, DT390-BiscFv806 was 4 orders of magnitude more potent against the U87-EGFRvIII cells than against the parent U87 cells. The cytotoxicity against a group of 6 head and neck squamous cell carcinoma cell lines were further analyzed, showing an IC50 ranging from 0.24 nM to 156 nM, depending on the expression level of EGFR/EGFRvIII. In animals, the U87-EGFRvIII tumor xenografts grew extremely faster than the parental U87, and systemic administration of DT390-BiscFv806 significantly inhibited the growth of established U87-EGFRvIII and U87 tumor xenografts, showing a growth inhibition rate of 76.3% (59.82-96.2%) and 59.4% (31.5-76.0%), respectively. In pathology, the RIT-treated tumors exhibited a low mitotic activity and a large number of degenerative tumor cells, compared with the control tumors. The results indicate that DT390-BiscFv806 is promising for treatment of various types of cancer, especially for those with high EGFR expression or with EGFR and EGFRvIII co-expression.
表皮生长因子受体（EGFR）和EGFRvIII在多种癌症中过表达，是癌症治疗的理想靶点。利用人源化抗体806（mAb806）对癌症中过表达的EGFR和EGFRvIII的高特异性，我们通过将mAb806衍生的二价单链可变片段与白喉毒素片段DT390融合，设计并构建了一种二价重组免疫毒素（RIT，DT390-BiscFv806）。在体外，DT390-BiscFv806能有效内化进入细胞，并对U87胶质母细胞瘤细胞和EGFRvIII转染的U87（U87-EGFRvIII）细胞表现出高细胞毒性，其半数最大抑制浓度（IC50）分别为1.47 nM和2.26×10(-4) nM。值得注意的是，DT390-BiscFv806对U87-EGFRvIII细胞的效力比对亲代U87细胞高4个数量级。进一步分析了其对一组6种头颈鳞状细胞癌细胞系的细胞毒性，结果显示IC50范围为0.24 nM至156 nM，具体取决于EGFR/EGFRvIII的表达水平。在动物实验中，U87-EGFRvIII肿瘤异种移植瘤的生长速度比亲代U87快得多，全身给予DT390-BiscFv806可显著抑制已建立的U87-EGFRvIII和U87肿瘤异种移植瘤的生长，生长抑制率分别为76.3%（59.82 - 96.2%）和59.4%（31.5 - 76.0%）。在病理学上，与对照肿瘤相比，经RIT治疗的肿瘤表现出低有丝分裂活性和大量退行性肿瘤细胞。结果表明，DT390-BiscFv806在治疗多种类型癌症方面具有潜力，尤其是对于那些EGFR高表达或EGFR与EGFRvIII共表达的癌症。
