Antitumor activities of a cephalosporin prodrug in combination with monoclonal antibody-beta-lactamase conjugates.
7-(Phenylacetamido)cephalosporin mustard (CM) and 7-(4-carboxybutanamido)cephalosporin mustard (CCM) were developed as anticancer prodrugs that could be activated site selectively by monoclonal antibody-beta-lactamase conjugates targeted to antigens present on tumor cell surfaces. Both CM and CCM were hydrolyzed by purified beta-lactamases from Escherichia coli (EC beta L), Bacillus cereus (BC beta L), and Enterobacter cloacae (ECl beta L). This resulted in the release of phenylenediamine mustard (PDM), a potent cytotoxic drug. The Km and kcat values of the reactions were determined, and it was found that ECl beta L effected the hydrolysis of CM and CCM more rapidly than the other enzymes. Conjugates of ECl beta L were prepared by reacting maleimide-substituted F(ab')2 fragments of the monoclonal antibodies L6 and P1.17 to ECl beta L that had been modified with sulfhydryl groups. In vitro experiments indicated that CCM (IC50 = 25-45 microM) was less toxic than PDM (IC50 = 1.5 microM) to H2981 lung adenocarcinoma cells (L6 antigen positive, P1.17 antigen negative) and that immunologically specific prodrug activation took place when the cells were treated with L6-ECl beta L. In vivo experiments in nude mice demonstrated that CCM was less toxic than CM, and that both prodrugs were much less toxic than PDM. Neither CCM nor PDM exerted antitumor activity on subcutaneous H2981 tumors in vivo. However, a significant antitumor effect was obtained in mice that received L6-ECl beta L 96 h prior to the administration of CCM. The effect was immunologically specific (P < 0.05), since a smaller degree of antitumor activity was obtained in mice that received the nonbinding control conjugate P1.17-ECl beta L prior to CCM.(ABSTRACT TRUNCATED AT 250 WORDS)
Vrudhula
V M
VM
Bristol-Myers Squibb Pharmaceutical Research Institute, Seattle, Washington 98121.
Svensson
H P
HP
Kennedy
K A
KA
Senter
P D
PD
Wallace
P M
PM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D002511:Cephalosporins / Q000138:chemical synthesis* / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D004337:Drug Carriers; D005260:Female; D006868:Hydrolysis; D051379:Mice; D008819:Mice, Nude; D009588:Nitrogen Mustard Compounds / Q000138:chemical synthesis* / Q000494:pharmacology; D011355:Prodrugs / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014407:Tumor Cells, Cultured; D001618:beta-Lactamases / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000970:Antineoplastic Agents; D002511:Cephalosporins; D004337:Drug Carriers; D009588:Nitrogen Mustard Compounds; D011355:Prodrugs; C076378:cephalosporin mustard; C085091:7-(4-carboxybutanamido)cephalosporin mustard; D001618:beta-Lactamases