Development of a human lymphoblastoid cell line constitutively expressing human CYP1B1 cDNA: substrate specificity with model substrates and promutagens

Abstract

An AHH-1 TK+/- cell derivative was developed that stably expresses human cytochrome P4501B1 (CYP1B1) cDNA in an extrachromosomal vector which confers resistance to 1-histidinol and co-expresses NADPH cytochrome P450 oxidoreductase (OR). The CYP1B1-expressing cell line was designated h1B1/OR. Microsomes prepared from CYP1B1 cDNA expressing cells exhibit elevated levels of 7-ethoxy-resorufin deethylase (EROD), 7-ethoxy-4-trifluoromethyl-coumarin deethylase (EFCD), benzo(alpha)pyrene hydroxylase (BPH), bufuralol 1'-hydroxylase, testosterone hydroxylase activities and spectrally quantifiable cytochrome P450. CYP1B1-containing microsomes did not contain detectable coumarin 7-hydroxylase, p-nitrophenol hydroxylase, lauric acid hydroxylase, (S)-mephenytoin 4'-hydroxylase or diclofenac 4'-hydroxylase activities. Kinetic parameters for selected substrates were compared among CYP1B1 and the two additional members of the CYP1 family, CYP1A1 and CYP1A2. For BPH and EFCD, the rank order of rates of substrate metabolism were CYP1A1 > CYP1B1 > CYP1A2. For EROD, the rank order of substrate metabolism was CYP1A1 > CYP1A2 > CYP1B1. For both EROD and EFCD the apparent K(m) values for CYP1B1 were more similar to CYP1A1 than to CYP1A2. In order to begin to characterize the promutagen activating ability of CYP1B1, the mutagenicity of selected chemicals was examined in h1B1/OR cells; there was increased sensitivity (CYP1B1-expressing relative to control cells) to the mutagenicity of benzo(a)pyrene, cyclopenta(c,d)pyrene, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone and aflatoxin B1 (AFB). CYP1B1, expressed in this system, appears to be particularly efficient at activating AFB.

Transcript profiling of enzymes involved in detoxification of xenobiotics and reactive oxygen in human normal and simian virus 40 T antigen‐immortalized oral keratinocytes

“…Indeed, the results in SVpgC2a indicate that expression of CYP1B1 may increase at an early stage of malignant transformation. Furthermore, CYP1B1 efficiently activates aflatoxin B1, 36 and thus, the present results indicate that this CYP may have substantially contributed to the metabolic activation of this carcinogen in SVpgC2a. 6 In SVpgC2a vs. NOK, the comparatively higher expression of GSTM1, 2, 4, 5, GSTT1 and SOD2 and lower expression of UGT2B and MGST1, may to some extent be compensatory in nature.…”

The cancer preventative agent resveratrol is converted to the anticancer agent piceatannol by the cytochrome P450 enzyme CYP1B1

“…The metabolism of CYP1B1 was carried out using a microsomal preparation of the human CYP1B1 enzyme (Crespi et al, 1997). Using HPLC analysis with fluorescence detection we observed the formation of two major metabolites (M1 and M2) and one minor metabolite M3 (Figure 2A).…”

Immunohistochemical Localization of Cytochrome P450 CYP1B1 in Breast Cancer with Monoclonal Antibodies Specific for CYP1B1

“…CYP1B1 is also capable of metabolizing a variety of putative human carcinogens, including polycyclic aromatic hydrocarbons and heterocyclic amines (Shimada et al 1996;Crespi et al 1997). Therefore, CYP1B1 appears to have potentially important roles in tumor development and progression, as a potential target for anti-cancer drugs, and as a tumor biomarker.…”