Abstract

This is the first revision of a competitive continuation. The work is divided into four specific aims: SA1. To produce and purify a battery of Mab reagents directed against rat P450 3A1, 3A2 and 3Ax, and the related human P450s 3A3, 3A4, 3A5 and 3A7. It is proposed that members of the desired product set will be competent in ELISA assays, western blotting, inhibitory towards P450 activity in microsomal extracts, and that several antibodies binding to distinct epitopes will be found for each P450, to facilitate interspecies comparison of P450 homologues, for membrane topology analysis and immunocytochemical analysis. SA2. To study the age and sex specific regulation of 3A1 and 3A2 in the rat at the mRNA and protein level. SA3. To clone and characterize the cDNA for a novel 3A isotope (3Ax), recently identified by the principal investigator, and to study its expression as a function of age, sex and tissue type, as described in SA2. SA4. To use the inhibitory Mab s obtained in SA1, to selectively inhibit P450 expression in rat and human microsomal extracts, so as to establish the function of the various isotopes in the metabolism of selected substrates (CsA, progesterone, and others).

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM044982-07
Application #
2734677
Study Section
Chemical Pathology Study Section (CPA)
Project Start
1991-07-01
Project End
2000-06-30
Budget Start
1998-07-01
Budget End
1999-06-30
Support Year
7
Fiscal Year
1998
Total Cost
Indirect Cost

  Institution

Name
Rutgers University
Department
Biology
Type
Schools of Pharmacy
DUNS #
038633251
City
New Brunswick
State
NJ
Country
United States
Zip Code
08901

  Publications

Jan, Yi-Hua; Mishin, Vladimir; Busch, Christine M et al. (2006) Generation of specific antibodies and their use to characterize sex differences in four rat P450 3A enzymes following vehicle and pregnenolone 16alpha-carbonitrile treatment. Arch Biochem Biophys 446:101-10
Howard, Carolyn B; Samuel, Jacqueline; Henderson, Shalonda B et al. (2005) Effects of pristane on cytochrome P450 isozyme expression in rat tissues. Int J Environ Res Public Health 2:138-46
Turan, V K; Sanchez, R I; Li, J J et al. (2004) The effects of steroidal estrogens in ACI rat mammary carcinogenesis: 17beta-estradiol, 2-hydroxyestradiol, 4-hydroxyestradiol, 16alpha-hydroxyestradiol, and 4-hydroxyestrone. J Endocrinol 183:91-9
Parimoo, Bhama; Mishin, Vladimir M; Busch, Christine M et al. (2003) Identification of epitopes on cytochrome P450 3A4/5 recognized by monoclonal antibodies. Arch Biochem Biophys 414:244-54
Gunasekera, Nilhan; Musier-Forsyth, Karin; Arriaga, Edgar (2002) Electrophoretic behavior of individual nuclear species as determined by capillary electrophoresis with laser-induced fluorescence detection. Electrophoresis 23:2110-6
Turan, V K; Mishin, V M; Thomas, P E (2001) Clotrimazole is a selective and potent inhibitor of rat cytochrome P450 3A subfamily-related testosterone metabolism. Drug Metab Dispos 29:837-42
Parimoo, B; Thomas, P E (2000) Epitope mapping of rat cytochrome P450 2B1 using glutathione S-transferase-2B1 fusion constructs. Arch Biochem Biophys 380:117-25
Nims, R W; Prough, R A; Jones, C R et al. (1997) In vivo induction and in vitro inhibition of hepatic cytochrome P450 activity by the benzodiazepine anticonvulsants clonazepam and diazepam. Drug Metab Dispos 25:750-6
Ghosal, A; Sadrieh, N; Reik, L et al. (1996) Induction of the male-specific cytochrome P450 3A2 in female rats by phenytoin. Arch Biochem Biophys 332:153-62
Suchar, L A; Chang, R L; Thomas, P E et al. (1996) Effects of phenobarbital, dexamethasone, and 3-methylcholanthrene administration on the metabolism of 17 beta-estradiol by liver microsomes from female rats. Endocrinology 137:663-76

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