Huang Sheng-Cih, Wang Yu-Kuo, Huang Wan-Ting, Kuo Tsam-Ming, Yip Bak-Sau, Li Tien-Hsiung Thomas, Wu Tung-Kung
Department of Biological Science and Technology, National Chiao Tung University, Hsin-Chu, Taiwan.
Cancer Sci. 2015 Apr;106(4):447-54. doi: 10.1111/cas.12623. Epub 2015 Mar 10.
We report on the preparation of a new type of immunotoxin by conjugation of an epidermal growth factor receptor (EGFR)-binding peptide and an R46E mutation of thermostable direct hemolysin from Grimontia hollisae, (Gh-TDH(R) (46E) /EB). The hybrid immunotoxin was purified to homogeneity and showed a single band with slight slower mobility than that of Gh-TDH(R) (46E) . Cytotoxicity assay of Gh-TDH(R) (46E) /EB on EGFR highly, moderately, low, and non-expressed cells, A431, MDA-MB-231, HeLa, and HEK293 cells, respectively, showed apparent cytotoxicity on A431 and MDA-MB-231 cells but not on HeLa or HEK293 cells. In contrast, no cytotoxicity was observed for these cells treated with either Gh-TDH(R) (46E) or EB alone, indicating enhanced cytotoxic efficacy of Gh-TDH(R) (46E) by the EGFR binding moiety. Further antitumor activity assay of Gh-TDH(R) (46E) /EB in a xenograft model of athymic nude mice showed obvious shrinkage of tumor size and degeneration, necrosis, and lesions of tumor tissues compared to the normal tissues. Therefore, the combination of Gh-TDH(R) (46E) with target affinity agents opens new possibilities for pharmacological treatment of cancers and potentiates the anticancer drug's effect.
我们报道了一种新型免疫毒素的制备方法，该免疫毒素通过将表皮生长因子受体（EGFR）结合肽与来自霍利斯格氏菌的耐热直接溶血素的R46E突变体（Gh-TDH(R)(46E)/EB）偶联而成。该杂合免疫毒素被纯化至同质，显示出一条迁移率略慢于Gh-TDH(R)(46E)的单一条带。分别对EGFR高表达、中度表达、低表达和不表达的细胞A431、MDA-MB-231、HeLa和HEK293细胞进行Gh-TDH(R)(46E)/EB的细胞毒性测定，结果显示其对A431和MDA-MB-231细胞具有明显的细胞毒性，但对HeLa或HEK293细胞无细胞毒性。相比之下，单独用Gh-TDH(R)(46E)或EB处理这些细胞均未观察到细胞毒性，这表明EGFR结合部分增强了Gh-TDH(R)(46E)的细胞毒性效力。在无胸腺裸鼠异种移植模型中对Gh-TDH(R)(46E)/EB进行的进一步抗肿瘤活性测定显示，与正常组织相比，肿瘤大小明显缩小，肿瘤组织出现变性、坏死和病变。因此，Gh-TDH(R)(46E)与靶向亲和剂的结合为癌症的药物治疗开辟了新的可能性，并增强了抗癌药物的效果。
