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Key Event Title
CYP7B activity, inhibition
Key Event Components
Key Event Overview
AOPs Including This Key Event
|AOP Name||Role of event in AOP||Point of Contact||Author Status||OECD Status|
|Inhibition of CYP7B leads to decreased locomotor activity||MolecularInitiatingEvent||Florence Pagé-Larivière (send email)||Not under active development|
|Inhibition of CYP7B activity leads to decreased sexual behavior||MolecularInitiatingEvent||Florence Pagé-Larivière (send email)||Not under active development|
|All life stages|
Key Event Description
Site of action:
CYP7B is expressed in different organs including liver, prostate and brain.
How does it work :
CYP7B is a member of the cytochrome P450 family of enzymes. It is involved in steroidogenic pathways as well as in the synthesis of bile acids. In the brain, it is involved in neurosteroids synthesis.
In the brain, the reactions catalyzed by CYP7B are :
- Probably in all vertebrates: Pregnenolone into 7α-hydroxypregnenolone and its stereoisomer 7β-hydroxypregnenolone (bird only) (R08943) (Matsunaga et al., 2004; Rose et al., 1997; Tsutsui et al., 2008)
- Proven in mouse and human: Dehydroepiandrosterone (DHEA) to 7α-hydroxy-DHEA and its stereoisomer 7β-hydroxy-DHEA (Martin et al., 2004; Weihua et al., 2002).
In the human and mouse liver, CYP7B is responsible for (Toll et al., 1994):
- 5-cholesten-3-beta, 25(S)-diol into Cholest-5-ene-3 beta-7 alpha, 25-thiol (R07209 R08723),
- Cholest-5-ene-3 beta, 26-diol into 7 alpha, 27-dihydroxycholesterol (R07372 R08724),
- 3 beta-hydroxy-5-cholestenoate into 3 beta, 7 alpha-dihydroxy-5-cholestenoate (R08727 R08728).
- It is expressed in the chicken liver and is probably involved in the same reactions (Handschin et al., 2005).
In the prostate:
- Proven for human and rat: Dehydroepiandrosterone (DHEA) to 7α-hydroxy-DHEA and 7β-hydroxy-DHEA (Martin et al., 2001; Martin et al., 2004).
Inhibitors prevent the metabolism of pregnenolone into 7-alpha-hydroxypregnenolone, thereby decreasing the concentration of the neurosteroid.
How It Is Measured or Detected
To measure CYP7B activity in vitro, different experiments based on HPLC and GS-MS analysis can be performed.
- An assay in liver microsome followed by HPLC analysis of the metabolites (Souidi et al., 2000).
- Labeled steroid conversion in vitro with cell or tissue extract in presence of NADPH followed by GS-MS analysis (Rose et al., 1997; Tsutsui et al., 2008).
- CYP7B can be cloned in bacteria to produce an active protein in vitro. In presence of adequate precursor and cofactors, the enzymatic activity of the protein can be measured and analyzed using HPLC.
- CYP7B can be transfected in a cell line unable to synthesize 7α-hydroxypregnenolone in order to measure with HPLC the ability of the protein to catalyze the enzymatic reaction in presence of the appropriate substrate and cofactor (Tsutsui et al., 2008)
Experiments may include knock-out of mice (followed by RNA, protein blotting and enzymatic activity to confirm knock-out) (Li-Hawkins et al., 2000) followed by the measurement of substrate and metabolites of CYP7B in plasma and tissues (Rose., 2001).
Domain of Applicability
CYP7B is known to be conserved in chimpanzee, Rhesus monkey, dog, cow, mouse, rat, chicken, zebrafish, and frog. https://www.ncbi.nlm.nih.gov/homologene/3544
Evidence for Perturbation by Stressor
Overview for Molecular Initiating Event
The binding of inhibitors to CYP7B is demonstrated in vitro with purified recombinant protein in presence of the inhibitor. Ligand-induced spectral changes is analyzed using spectrophotometric titration as a shift of the heme (Yantsevich et al., 2014).
Ketoconazole and other conazole are known to bind to CYPs preventing its enzymatic activity.
- CYP7B inhibitor (ketoconazole, 10-4 M) decreased the synthesis of 7α-hydroxypregnenolone
- CYP7B inhibitor (intracerebroventricular injection of ketoconazole) decreased the synthesis of 7α-hydroxypregnenolone in the male quail and newt brain, in vivo (Matsunaga et al., 2004; Rose et al., 1997; Tsutsui et al., 2008).
- The heme prosthetic group (catalytic site) of human recombinant CYP7B thightly bound to various imidazole- and triazole-based drugs in an in vitro spectrometric titration assay. The drugs with the highest affinities were the industrial pesticides tebuconazole (0.11 μm), propiconazole (0.13 μm) and the antifungal drugs tioconazole (0.15 μm) and miconazole (0.23 μm). Voriconazole and metyrapone (non-azole compound) also interacted with CYP7B (Yantsevich et al., 2014).
It is clearly demonstrated that ketoconazole directly inhibits CYP7B (Matsunaga et al., 2004). It is expected for the other members of the conazole family to have the same effect.
Some other azoles such as clotrimazole can also inhibit CYP7B activity (Liu et al., 2011; Rose et al., 1997).
In vitro, tebuconazole was shown to bind to the catalytic site of the human recombinant CYP7B and to inhibit its catalytic activity (Yantsevich et al., 2014).
In vitro, propiconazole was shown to bind to the catalytic site of the human recombinant CYP7B and to inhibit its activity (Yantsevich et al., 2014).
In vitro, tioconazole was shown to bind to the catalytic site of the human recombinant CYP7B and to inhibit its activity (Yantsevich et al., 2014).
In vitro, miconazole was shown to bind to the catalytic site of the human recombinant CYP7B and to inhibit its activity (Yantsevich et al., 2014).
In vitro, fluconazole was shown to bind to the catalytic site of the human recombinant CYP7B and to inhibit its activity (Yantsevich et al., 2014).
In vitro, voriconazole was shown to bind to the catalytic site of the human recombinant CYP7B and to inhibit its activity (Yantsevich et al., 2014).
Clotrimazoles can inhibit CYP7B activity (Liu et al., 2011; Rose et al., 1997).
Dulos, J., van der Vleuten, M.A., Kavelaars, A., Heijnen, C.J., and Boots, A.M. (2005). CYP7B expression and activity in fibroblast-like synoviocytes from patients with rheumatoid arthritis: regulation by proinflammatory cytokines. Arthritis Rheum 52, 770-778.
Handschin C., Gnerre C., Fraser DJ., Martinez-Jimenez C., Jover R., Mever UA., (2005) Species-specific mechanisms for cholesterol 7α-hydroxylase (CYP7A1) regulation by drugs and bile acids, Archives of Biochemistry and Biophysics, Vol 434-1, pp75-85
Haraguchi, S., Koyama, T., Hasunuma, I., Okuyama, S., Ubuka, T., Kikuyama, S., Do Rego, J.L., Vaudry, H., and Tsutsui, K. (2012). Acute stress increases the synthesis of 7alpha-hydroxypregnenolone, a new key neurosteroid stimulating locomotor activity, through corticosterone action in newts. Endocrinology 153, 794-805.
Haraguchi, S., Yamamoto, Y., Suzuki, Y., Hyung Chang, J., Koyama, T., Sato, M., Mita, M., Ueda, H., and Tsutsui, K. (2015). 7alpha-Hydroxypregnenolone, a key neuronal modulator of locomotion, stimulates upstream migration by means of the dopaminergic system in salmon. Sci Rep 5, 12546.
Li-Hawkins, J., Lund, E.G., Turley, S.D., and Russell, D.W. (2000). Disruption of the oxysterol 7alpha-hydroxylase gene in mice. J Biol Chem 275, 16536-16542.
Liu, C., Yang, X.V., Wu, J., Kuei, C., Mani, N.S., Zhang, L., Yu, J., Sutton, S.W., Qin, N., Banie, H., et al. (2011). Oxysterols direct B-cell migration through EBI2. Nature 475, 519-523.
Martin, C., Bean, R., Rose, K., Habib, F., and Seckl, J. (2001). cyp7b1 catalyses the 7alpha-hydroxylation of dehydroepiandrosterone and 25-hydroxycholesterol in rat prostate. Biochem J 355, 509-515.
Martin, C., Ross, M., Chapman, K.E., Andrew, R., Bollina, P., Seckl, J.R., and Habib, F.K. (2004). CYP7B generates a selective estrogen receptor beta agonist in human prostate. J Clin Endocrinol Metab 89, 2928-2935.
Matsunaga, M., Ukena, K., Baulieu, E.E., and Tsutsui, K. (2004). 7alpha-Hydroxypregnenolone acts as a neuronal activator to stimulate locomotor activity of breeding newts by means of the dopaminergic system. Proc Natl Acad Sci U S A 101, 17282-17287.
Rose, K., Allan, A., Gauldie, S., Stapleton, G., Dobbie, L., Dott, K., Martin, C., Wang, L., Hedlund, E., Seckl, J.R., et al. (2001). Neurosteroid hydroxylase CYP7B: vivid reporter activity in dentate gyrus of gene-targeted mice and abolition of a widespread pathway of steroid and oxysterol hydroxylation. J Biol Chem 276, 23937-23944.
Rose, K.A., Stapleton, G., Dott, K., Kieny, M.P., Best, R., Schwarz, M., Russell, D.W., Bjorkhem, I., Seckl, J., and Lathe, R. (1997). Cyp7b, a novel brain cytochrome P450, catalyzes the synthesis of neurosteroids 7alpha-hydroxy dehydroepiandrosterone and 7alpha-hydroxy pregnenolone. Proc Natl Acad Sci U S A 94, 4925-4930.
Souidi, M., Parquet, M., Dubrac, S., Audas, O., Becue, T., and Lutton, C. (2000). Assay of microsomal oxysterol 7alpha-hydroxylase activity in the hamster liver by a sensitive method: in vitro modulation by oxysterols. Biochim Biophys Acta 1487, 74-81.
Toll, A., Wikvall, K., Sudjana-Sugiaman, E., Kondo, K.H., and Bjorkhem, I. (1994). 7 alpha hydroxylation of 25-hydroxycholesterol in liver microsomes. Evidence that the enzyme involved is different from cholesterol 7 alpha-hydroxylase. Eur J Biochem 224, 309-316.
Tsutsui, K., Inoue, K., Miyabara, H., Suzuki, S., Ogura, Y., and Haraguchi, S. (2008). 7Alpha-hydroxypregnenolone mediates melatonin action underlying diurnal locomotor rhythms. J Neurosci 28, 2158-2167.
Weihua, Z., Lathe, R., Warner, M., and Gustafsson, J.A. (2002). An endocrine pathway in the prostate, ERbeta, AR, 5alpha-androstane-3beta,17beta-diol, and CYP7B1, regulates prostate growth. Proc Natl Acad Sci U S A 99, 13589-13594.
Yantsevich, A.V., Dichenko, Y.V., Mackenzie, F., Mukha, D.V., Baranovsky, A.V., Gilep, A.A., Usanov, S.A., and Strushkevich, N.V. (2014). Human steroid and oxysterol 7alpha-hydroxylase CYP7B1: substrate specificity, azole binding and misfolding of clinically relevant mutants. FEBS J 281, 1700-1713.