Regioselective 2-hydroxylation of 17β-estradiol by rat cytochrome P4501B1

Rahman, Mostafizur; Sutter, Carrie Hayes; Emmert, Gary L.; Sutter, Thomas R.

doi:10.1016/j.taap.2006.06.004

Cited by 15 publications

(21 citation statements)

References 49 publications

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“…For the initial AI screen, the same preparation of CYP1B1 microsomes was used for the inhibition studies. The measured E2 4-hydroxylase activity was 1.23 nmol/min/nmol P450, consistent with the turnover number published in our previous studies Rahman et al, 2006). The inhibition kinetics of CYP1B1 was determined in a range expected to produce 30 to 90% inhibition.…”

Section: Cyp19 Protein Expression and Purification Of Hexahistidine-tsupporting

confidence: 89%

“…Anastrozole was obtained from AstraZeneca Pharmaceuticals LP (Wilmington, DE), and vorozole was from Jannsen (Beerse, Belgium). The other materials used in this study were described previously in detail (Rahman et al, 2006). CYP19 Protein Expression and Purification of Hexahistidine-Tagged Proteins in Escherichia coli.…”

Section: Methodsmentioning

confidence: 99%

“…The E2 hydroxylation assay was performed with the addition of inhibitor and has been previously described in detail (Rahman et al, 2006). Inhibition was calculated as percentage of product formation compared with the corresponding control (enzyme-substrate reaction) without the inhibitors.…”

Section: Methodsmentioning

confidence: 99%

“…The recombinant human CYP1B1 protein was expressed in Saccharomyces cerevisiae as described previously . Specific CYP1B1 protein content was estimated and evaluated by measuring the enzymatic activity as previously described (Rahman et al, 2006). For the initial AI screen, the same preparation of CYP1B1 microsomes was used for the inhibition studies.…”

Section: Cyp19 Protein Expression and Purification Of Hexahistidine-tmentioning

confidence: 99%

“…A fixed substrate concentration and varying inhibitor concentrations were used to determine the IC 50 value at the point where 50% inhibition of the catalytic activity of the enzyme occurred. The E2 hydroxylation assay was performed with the addition of inhibitor and has been previously described in detail (Rahman et al, 2006). Inhibition was calculated as percentage of product formation compared with the corresponding control (enzyme-substrate reaction) without the inhibitors.…”

mentioning

confidence: 99%

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CYP1B1 Is Not a Major Determinant of the Disposition of Aromatase Inhibitors in Epithelial Cells of Invasive Ductal Carcinoma

Rahman

Lax²,

Sutter³

et al. 2008

Drug Metab Dispos

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ABSTRACT:CYP1B1 and CYP19 (aromatase) have been shown to be expressed in breast tumors. Both enzymes are efficient estrogen hydroxylases, indicating the potential for overlapping substrate and inhibitor specificity. We measured the inhibition properties of aromatase inhibitors (AIs) against CYP1B1-catalyzed hydroxylation of 17␤-estradiol (E2) to determine whether CYP1B1 affects the disposition of AIs. In addition, we estimated the frequency of coexpression of these enzymes in breast tumor epithelium. Immunohistochemical analyses of CYP19 and CYP1B1 in a panel of 29 cases of invasive ductal carcinoma of the breast showed epithelial cell staining for CYP19 in 76% and for CYP1B1 in 97% of the samples. Statistical analysis showed no significant correlation (0.33) for positive expression of CYP19 and CYP1B1 (p > 0.07). CYP1B1 inhibition was determined for two steroidal inhibitors:formestane and exemestane and five nonsteroidal inhibitors: aminoglutethimide, fadrozole, anastrozole, letrozole, and vorozole. Of the seven compounds tested, only vorozole exhibited inhibition of CYP1B1 activity with IC 50 values of 17 and 21 M for 4-hydroxy estradiol and 2-hydroxy estradiol, respectively. The estimated K i values of vorozole for E2 4-and 2-hydroxylation were 7.26 and 6.84 M, respectively. Spectrophotometric studies showed that vorozole was a type II inhibitor of CYP1B1. This study shows that with the exception of vorozole, the aromatase inhibitors are selective for CYP19 relative to CYP1B1. Thus, although both CYP19 and CYP1B1 are expressed in a high percentage of breast cancers, CYP1B1 is not a major determinant of the disposition of AIs.CYP19 (aromatase) catalyzes the formation of the phenolic A ring of estrogens, converting androstenedione to estrone and testosterone to estradiol (Johnston and Dowsett, 2003). CYP19 is found in glandular and nonglandular tissues, including adrenal glands, ovaries, placenta, testes, adipose tissue, muscle, brain, normal breast, and breast cancer tissue. Important to this work, studies have shown that substantial levels of estrogens arise from aromatase activity in breast tissue (Miller, 1991;Bulun et al., 1993). CYP19 protein and activity have been detected in both the epithelial cells and surrounding stroma and adipose tissue of breast tumors (Esteban et al., 1992;Lu et al., 1996;Brodie et al., 1997;Oliveira et al., 2006;Miki et al., 2007). A recent study showed that although significant CYP19 immunoreactivity occurred in each of these cellular compartments of breast tumors, a significant positive correlation between biochemical activity and immunostaining was detected only for the epithelium (Sasano et al., 2005).Aromatase inhibitors (AIs) are drugs that inhibit CYP19, preventing the formation of estrogens. The AIs are subdivided into steroidal (type I) and nonsteroidal (type II) agents (Johnston and Dowsett, 2003;Miller, 2006). Type 1 agents are analogs of androgens that bind to CYP19 through either reversible or irreversible mechanisms, competing with the natural substrates. Type II i...

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Section: Cyp19 Protein Expression and Purification Of Hexahistidine-tsupporting

confidence: 89%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Cyp19 Protein Expression and Purification Of Hexahistidine-tmentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

CYP1B1 Is Not a Major Determinant of the Disposition of Aromatase Inhibitors in Epithelial Cells of Invasive Ductal Carcinoma

Rahman

Lax²,

Sutter³

et al. 2008

Drug Metab Dispos

View full text Add to dashboard Cite

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Cytochrome P450 1B1 expression in rat esophageal tumorigenesis promoted by gastric and duodenal reflux

Devlin

McIlroy

McKeen

et al. 2008

Molecular Carcinogenesis

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Cytochrome P450 1B1 (CYP1B1) mRNA is constitutively expressed in most normal extra-hepatic tissues; however the protein is not detectable in these tissues but is expressed in a wide variety of tumors. CYP1B1 is responsible for the activation of a number of carcinogens present in tobacco smoke and food. A surgical model of rat esophageal tumorigenesis, promoted by gastric or duodenal reflux was used to determine CYP1B1 expression in premalignant esophageal tissue. Immunohistochemistry was performed using a modified amplified fluorescein tyramide protocol. CYP1B1 was not observed in normal esophageal mucosa, submucosa, or muscularis mucosa. Animals exposed to gastric reflux developed mild hyperplasia. Varying degrees of hyperplasia were observed in the duodenal reflux group. All regions of hyperplasia showed moderate or strong CYP1B1 immunoreactivity. Duodenal reflux induced a small number of premalignant changes: immunoreactivity was absent from the epithelium of squamous dysplasia (0/10), Barrett's esophagus (0/7), and majority of dysplastic Barrett's esophagus (1/4). Moderate or strong immunoreactivity was observed in the majority (7/8) of squamous cell carcinomas (SCCs) in situ. Immunoreactivity was also observed in the lamina propria and submucosa in association with inflammation, regardless of the severity of inflammation. The expression of CYP1B1 in hyperplasia, SCCs in situ, or in association with inflammation may increase the production of carcinogenic metabolites, which may promote esophageal tumorigenesis.

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