title: Construction of an immunotoxin, HN3-mPE24, targeting glypican-3 for liver cancer therapy.
abstract: Glypican-3 (GPC3) is overexpressed in hepatocellular carcinoma (HCC). We constructed a recombinant immunotoxin, HN3-mPE24, which contains a truncated form of Pseudomonas exotoxin A. The toxin portion lacks most of domain II and has seven point mutations in domain III to remove the B-cell epitopes thought to be responsible for causing off-target side effects and immunogenicity. We also fused a bivalent HN3 to mPE24. We tested these two molecules for GPC3 binding and cytotoxicity in HCC cell models. The KD values of HN3-mPE24 and HN3-HN3-mPE24 for GPC3-expressing tumor cells were 12 nM and 1.4 nM, respectively. The IC50 values of HN3-mPE24 and HN3-HN3-mPE24 for HCC cells were 0.2 nM and 0.4 nM, respectively. We also evaluated their toxicity and anti-tumor efficacy in mice. The maximum tolerated doses of HN3-mPE24 and HN3-HN3-mPE24 were 7 mg kg-1 and 3.6 mg kg-1, respectively. We treated mice with 5 mg kg-1 of HN3-mPE24 intravenously every other day for ten injections. The alpha-fetoprotein level of HN3-mPE24 treated group was approximately 700 fold less than that of the untreated group (1.1 μg ml-1 vs. 692.1 μg ml-1). In addition, 25% of the mice treated with HN3-mPE24 survived to the end of this study, which was 105 days after HCC tumor implantation. In conclusion, the HN3-mPE24 immunotoxin caused liver tumor regressions and extended survival with no significant side effects in mice. It is a promising candidate for the treatment of liver cancer that may be readily translated to humans.
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.|Department of Thoracic Surgery, Second Hospital of Jilin University, Changchun, 130041, China.
authors.forename: Chunguang
authors.initials: C
authors.lastname: Wang
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.|Key Laboratory of Human Functional Genomics of Jiangsu Province, Department of Cell Biology, School of Basic Medical Science, Nanjing Medical University, Nanjing, Jiangsu, 210029, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Gao
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Mingqian
authors.initials: M
authors.lastname: Feng
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Ira
authors.initials: I
authors.lastname: Pastan
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Mitchell
authors.initials: M
authors.lastname: Ho
mesh_terms: D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000473:pathology / Q000628:therapy*; D018929:Cell Culture Techniques; D019610:Cytoprotection; D004195:Disease Models, Animal; D005260:Female; D053673:Glypicans / Q000378:metabolism*; D006801:Humans; D000922:Immunotoxins / Q000378:metabolism*; D008113:Liver Neoplasms / Q000473:pathology / Q000628:therapy*; D051379:Mice; D008819:Mice, Nude; D023041:Xenograft Model Antitumor Assays
chemical_list: C105937:GPC3 protein, human; D053673:Glypicans; D000922:Immunotoxins
keywords: glypican-3 or GPC3; hepatocellular carcinoma; mouse xenograft model; recombinant immunotoxin; single-domain antibody fragment