Principal drug-metabolizing enzyme systems in L1210 leukemia sensitive or resistant to BCNU in vivo.
1,3-Bis(2-chloroethyl)-1-nitrosourea (BCNU) resistance has been mostly studied in vitro. In an attempt to better understand BCNU resistance in the in vivo situation, we compared the principal drug-metabolizing enzyme systems in two L1210 leukemia lines, one sensitive and one resistant to BCNU (L1210/BCNU), passaged in vivo in mice. The following enzymes were assayed by immunoblotting: cytochromes P-450 (1A1/1A2, 2B1/2B2, 2C8-10, 2E1, 3A), epoxide hydrolase (EH) and glutathione S-transferase (GST-alpha, -mu and -pi). The following enzymes and cofactors were assayed fluorometrically or spectrophotometrically: 1-chloro-2-4 dinitrobenzene-GST (CDNB-GST), total glutathione (GSH), UDP-glucuronosyltransferase, beta-glucuronidase, sulfatase and sulfotransferase. Results showed that cytochrome P-450 1A1/1A2 was the only isoenzyme detected in both L1210 and L1210/BCNU. CDNB-GST activity was significantly higher in L1210/BCNU compared with L1210. The isoenzyme GST-alpha was more abundant in L1210/BCNU compared with L1210, whereas GST-pi was expressed less in the BCNU-resistant leukemia line. GST-mu was not detected in either L1210 leukemia lines. GSH levels were similar in the two L1210 lines. No significant difference was observed between the two leukemia lines for the conjugative enzymes UDP-glucuronosyltransferase and sulfotransferase, whereas their corresponding hydrolytic enzymes beta-glucuronidase and sulfatase were about two-fold lower in the BCNU-resistant leukemia line. Epoxide hydrolase was 1.3-fold higher in L1210/BCNU compared with L1210 and this level was about three-fold higher than in mouse liver. In conclusion, these studies showed the presence of cytochrome P-450 1A1/1A2 in the two L1210 leukemia lines studied, and indicated noteworthy differences between the two leukemia lines for many enzyme systems such as GST, beta-glucuronidase, sulfatase and epoxide hydrolase. These data are of importance to better understand the mechanisms of drug resistance to nitrosoureas in vivo.
Ribrag
V
V
Département de Pharmacotoxicologie et de Pharmacogénétique, CNRS URA 147, Institut Gustave-Roussy, Villejuif, France.
Massaad
L
L
Janot
F
F
Bissery
M C
MC
Parise
O
O
Gouyette
A
A
Chabot
G G
GG
D000818:Animals; D002330:Carmustine / Q000378:metabolism / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004351:Drug Resistance; D004851:Epoxide Hydrolases / Q000378:metabolism; D005260:Female; D005966:Glucuronidase / Q000378:metabolism; D014453:Glucuronosyltransferase / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D007939:Leukemia L1210 / Q000201:enzymology*; D051379:Mice; D008815:Mice, Inbred Strains; D013429:Sulfatases / Q000378:metabolism; D015238:Sulfotransferases / Q000378:metabolism
D003577:Cytochrome P-450 Enzyme System; D014453:Glucuronosyltransferase; D005982:Glutathione Transferase; D015238:Sulfotransferases; D013429:Sulfatases; D005966:Glucuronidase; D004851:Epoxide Hydrolases; D005978:Glutathione; D002330:Carmustine