Generation and antitumor effects of an engineered and energized fusion protein VL-LDP-AE composed of single-domain antibody and lidamycin.
Type IV collagenase plays a pivotal role in invasion, metastasis and angiogenesis of tumor. Single domain antibodies are attractive as tumor-targeting vehicle because of their much smaller size compared with antibody molecules produced by conventional methods. Lidamycin (LDM) is a potent enediyne-containing antitumor antibiotic. In this study an engineered and energized fusion protein VL-LDP-AE composed of lidamycin and VL domain of mAb 3G11 directed against type IV collagenase was prepared using a novel two-step method. First a VL-LDP fusion protein was constructed by DNA recombination. Secondly VL-LDP-AE was obtained by molecular reconstitution. In MTT assay, VL-LDP-AE showed potent cytotoxicity to HT-1080 cells and KB cells with IC(50) values of 8.55 x 10(-12) and 1.70 x 10(-11) mol/L, respectively. VL-LDP-AE showed antiangiogenic activity in chick chrorioallantoic membrane (CAM) assay and tube formation assay. In in vivo experiments, VL-LDP-AE was proved to be more effective than free LDM against the growth of subcutaneously transplanted hepatoma 22 in mice. Drugs were given intravenously on day 3 and 10 after tumor transplantation. Compared in terms of maximal tolerated doses, VL-LDP-AE at 0.25 mg/kg suppressed the tumor growth by 89.5%, LDM at 0.05 mg/kg by 69.9%, and mitomycin at 1 mg/kg by 35%. Having a molecular weight of 25.2 kDa, VL-LDP-AE was much smaller than other reported antibody-based drugs. The results suggested that VL-LDP-AE would be a promising candidate for tumor targeting therapy. And the 2-step approach could serve as a new technology platform for making a series of highly potent engineered antibody-based drugs for a variety of cancers.
Miao
QingFang
Q
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College Beijing 100050, China.
Shang
BoYang
B
Ouyang
ZhiGang
Z
Liu
XiaoYun
X
Zhen
YongSu
Y
D000617:Aminoglycosides / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000096:biosynthesis / Q000494:pharmacology*; D000912:Antibodies, Neoplasm / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D001483:Base Sequence; D045744:Cell Line, Tumor; D002642:Chick Embryo; D017364:Collagenases / Q000276:immunology; D053281:Enediynes / Q000494:pharmacology*; D006801:Humans; D008114:Liver Neoplasms, Experimental / Q000473:pathology / Q000628:therapy; D051379:Mice; D008970:Molecular Weight; D018919:Neovascularization, Physiologic / Q000187:drug effects; D010957:Plasmids / Q000235:genetics; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology
D000617:Aminoglycosides; D000903:Antibiotics, Antineoplastic; D000912:Antibodies, Neoplasm; D053281:Enediynes; D011993:Recombinant Fusion Proteins; C058024:C 1027; D017364:Collagenases