Construction and chemotherapeutic potential of carboxypeptidase-A/monoclonal antibody conjugate.
Carboxypeptidase-A and a monoclonal antibody (KS1/4) directed against a human lung carcinoma cell line (UCLA-P3) were derivatized by treatment with succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate and N-succinimidyl 3-(2-pyridyldithio)propionate, respectively. Admixture of these entities produced a stable conjugate containing 4 to 5 enzyme molecules per molecule of antibody. The conjugate (Mr approximately equal to 300 kDa) was purified to homogeneity by HPLC gel filtration and HPLC ion-exchange chromatography. Neither the catalytic activity of the enzyme nor the antigen-binding capacity of the monoclonal antibody was impaired in the conjugate. UCLA-P3 cells that had been exposed to the conjugate and then washed thoroughly were extremely sensitive to methotrexate alpha-alanine (MTX-Ala), a prodrug form of MTX. At 10(-5) M, MTX-Ala was almost as effective as free MTX in blocking the replication of conjugate-treated cells. These results demonstrate the chemotherapeutic potential of enzyme-monoclonal antibody conjugates used in conjunction with prodrugs.
Esswein
A
A
Department of Molecular and Experimental Medicine, Research Institute of Scripps Clinic, La Jolla, CA 92037.
Hänseler
E
E
Montejano
Y
Y
Vitols
K S
KS
Huennekens
F M
FM
D000911:Antibodies, Monoclonal; D002268:Carboxypeptidases / Q000494:pharmacology*; D043422:Carboxypeptidases A; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D006801:Humans; D000922:Immunotoxins / Q000138:chemical synthesis* / Q000302:isolation & purification / Q000494:pharmacology; D008175:Lung Neoplasms; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000494:pharmacology; D011355:Prodrugs / Q000494:pharmacology*
D000911:Antibodies, Monoclonal; D000922:Immunotoxins; D011355:Prodrugs; C066904:methotrexate-alpha-alanine; D002268:Carboxypeptidases; D043422:Carboxypeptidases A; D008727:Methotrexate