Preparation and properties of the immunoconjugate composed of anti-human colon cancer monoclonal antibody and mitomycin C-dextran conjugate.
Monoclonal antibody (mAb) A7, produced against human colon cancer, was conjugated with a polymeric prodrug of mitomycin C (MMC), the MMC-dextran conjugate with an anionic charge (MMCDan) and a molecular weight of 70,000. The amino groups were introduced into the MMCDan by reacting ethylenediamine with the carboxyl group in the spacer arm of the dextran bridge by a carbodiimide-catalyzed reaction. The coupling to mAb A7 was performed using SPDP. A 15 M excess of ethylenediamine produced an optimal MMCDan with amino groups, which resulted in a homogenous conjugate (A7-MMCD) with minimal formation of high-molecular-weight aggregates in about a 30% yield of both IgG and MMC. The molar binding ratio of IgG:dextran:MMC in A7-MMCD was estimated to be 1:1.2:40. A7-MMCD, having MMC prodrug properties, released active MMC with a half-life of 29.1 h and had an almost neutral electric charge under physiological conditions. A competitive binding assay using 125I-labeled A7 revealed that the A7-MMCD almost fully retained its antibody-binding activity. The cytotoxicity of A7-MMCD was assayed by determining the degree of inhibition of [3H]-thymidine in corporation in two different ways using the human colon cancer cell line SW1116. A 48-h continuous exposure test revealed that the pharmacological activity of MMC in A7-MMCD was completely preserved. In addition, A7-MMCD exhibited about a 14-fold greater cytotoxicity than MMCDan when the IC50 values determined using a 2-h pretreatment exposure system were compared. These results suggest that A7-MMCD could be useful in immunotargeting chemotherapy for colorectal cancer.
Noguchi
A
A
Department of Surgery, Kyoto Prefectural University of Medicine, Japan.
Takahashi
T
T
Yamaguchi
T
T
Kitamura
K
K
Takakura
Y
Y
Hashida
M
M
Sezaki
H
H
D000911:Antibodies, Monoclonal / Q000737:chemistry*; D002470:Cell Survival / Q000187:drug effects; D003110:Colonic Neoplasms / Q000276:immunology* / Q000473:pathology; D003911:Dextrans / Q000737:chemistry*; D004337:Drug Carriers; D006801:Humans; D016685:Mitomycin / Q000737:chemistry / Q000494:pharmacology*; D008970:Molecular Weight; D011355:Prodrugs / Q000737:chemistry / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D003911:Dextrans; D004337:Drug Carriers; D011355:Prodrugs; D016685:Mitomycin