TY - JOUR
T1 - Substrate specificity of bleomycin hydrolase
AU - Sebti, Said M.
AU - Deleon, John C.
AU - Ma, Ling Tai
AU - Hecht, Sidney M.
AU - Lazo, John S.
N1 - Funding Information:
* This work was supported by USPHS Grants CA-43917 and CA-27603 and by American Cancer Society Grant CH-316. J.S.L. is a USPHS Research Career Development Awardee (CA-01012). $ Present address: Department of Pharmacology, University of Pittsburgh, School of Medicine, 518 Scaife Hall, Pittsburgh, PA 15261. I Send reprint requests to: Dr. Said M. Sebti, Department of Pharmacology, University of Pittsburgh, School of Medicine, 518 Scaife Hall, Pittsburgh, PA 15261. 1 Abbreviations: BLM, bleomycin; TLM, tallysomycin; PEP, peplomycin; BAPP, butylamino-3-propylamino-3-propylamine bleomycin; dgBLM Ar, deglycobleomycin Ar; HSLC, high speed liquid chromatography.
PY - 1989/1/1
Y1 - 1989/1/1
N2 - Bleomycin (BLM) hydrolase is believed to protect both malignant and normal tissue from the toxicity of the antitumor drug BLM. Little is known about the substrate specificity of BLM hydrolase. Thus, we developed ion-paired reverse phase high speed liquid chromatography systems to assay for the metabolism of several BLM analogs. We found that BLM A2, BLM B2, tallysomycin S10b (TLM S10b) peplomycin (PEP), butylamino-3-propylamino-3-propylamine bleomycin (BAPP), deglyco bleomycin A2 (dgBLM A2) and bleomycinic acid were each metabolized by rabbit lung BLM hydrolase to a single metabolite. When compared to their corresponding parent compounds, these metabolites were 6- to 35-fold less potent in their ability to inhibit the proliferation of A-253 human head and neck squamous carcinoma cells in culture. Furthermore, we found that substitutions in various regions of the BLM molecule greatly affected the kinetic parameters of BLM hydrolase. For example, the Km with BLM B2 (0.056 ± 0.005 mM) was 15-fold lower than that seen with BLM A2 (0.83 ± 0.11 mM). In contrast, the Vmax was not affected markedly by these terminal amine substitutions but was influenced greatly by deletion of the carbohydrate groups of BLM. For example, a 4-fold higher Vmax was observed with dgBLM A2 compared to BLM A2. Thus, these results demonstrate that BLM hydrolase can recognize and metabolize a broad spectrum of BLM analogs regardless of their structural features. This enzymatic conversion resulted in the inactivation of the BLMs as demonstrated by a substantial decrease in their cytotoxidty. Furthermore, the terminal amine and carbohydrate regions, respectively, dictate the apparent affinity and the rate of metabolism of BLM hydrolase substrates.
AB - Bleomycin (BLM) hydrolase is believed to protect both malignant and normal tissue from the toxicity of the antitumor drug BLM. Little is known about the substrate specificity of BLM hydrolase. Thus, we developed ion-paired reverse phase high speed liquid chromatography systems to assay for the metabolism of several BLM analogs. We found that BLM A2, BLM B2, tallysomycin S10b (TLM S10b) peplomycin (PEP), butylamino-3-propylamino-3-propylamine bleomycin (BAPP), deglyco bleomycin A2 (dgBLM A2) and bleomycinic acid were each metabolized by rabbit lung BLM hydrolase to a single metabolite. When compared to their corresponding parent compounds, these metabolites were 6- to 35-fold less potent in their ability to inhibit the proliferation of A-253 human head and neck squamous carcinoma cells in culture. Furthermore, we found that substitutions in various regions of the BLM molecule greatly affected the kinetic parameters of BLM hydrolase. For example, the Km with BLM B2 (0.056 ± 0.005 mM) was 15-fold lower than that seen with BLM A2 (0.83 ± 0.11 mM). In contrast, the Vmax was not affected markedly by these terminal amine substitutions but was influenced greatly by deletion of the carbohydrate groups of BLM. For example, a 4-fold higher Vmax was observed with dgBLM A2 compared to BLM A2. Thus, these results demonstrate that BLM hydrolase can recognize and metabolize a broad spectrum of BLM analogs regardless of their structural features. This enzymatic conversion resulted in the inactivation of the BLMs as demonstrated by a substantial decrease in their cytotoxidty. Furthermore, the terminal amine and carbohydrate regions, respectively, dictate the apparent affinity and the rate of metabolism of BLM hydrolase substrates.
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U2 - 10.1016/0006-2952(89)90160-3
DO - 10.1016/0006-2952(89)90160-3
M3 - Article
C2 - 2462878
AN - SCOPUS:0024497864
SN - 0006-2952
VL - 38
SP - 141
EP - 147
JO - Biochemical Pharmacology
JF - Biochemical Pharmacology
IS - 1
ER -