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Hi,
I have to perform in vitro study (human hepatocytes) on oral
contraceptives
metabolism induction by the compound of interest. Most critical
contraceptive is ethinyl estradiol (EE2)
1.What is the major metabolic pathway for EE2 in humans: is it
sulphation,
glucoronidation or hydroxylation?
2.What would be preferable experimental set up: looking for induction
on EE2
directly (hepatocytes, test cmpd. and EE2) or looking for inducibility
of
metabolism by specific CYP's (hepatocytes, specific CYP substrate and
test
cmpd.)?
3. Use cryopreserved or fresh human hepatocytes?
I would appreciate answer on any of questions.
Best regards
Sinisa Stipanicic, MS Chem.
PLIVA Research Institute Ltd.
Drug Metabolism and Pharmacokinetics
Prilaz baruna Filipovica 25
10000 Zagreb, CROATIA
Phone: +385 1 372 2572
Fax: +385 1 372 1570
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Hi,
For the induction study, obviously the best choice will be fresh
hepatocytes.
EE2 metabolism essentially produces 2-hydroxy-EE2, 2-methoxy-EE2, and
EE2
sulphate and glucuronide conjugates if S9 or microsomes used for the
incubations. However, if one uses human hepatocytes the main
metabolites,
you will observe only EE2 glucuronide and sulphate conjugates.
My guess is that looking for directly EE2 glucuronide and sulphate
conjugates could be one of ways to look for the induction. However, the
most
convenient way would be to use specific substrate marker and hepatocytes
pre-treated with TA or solvent control to investigate whether EE2 causes
P450, phase II enzymes or transporter induction.
I hope this helps.
Thanks,
Kishore
Kishore K. Khan, Ph. D.
CellzDirect
8609 Cross Park Drive
Austin, TX 78754
Tel: (512) 615 2332
Fax: (512) 834 7767
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Thanks Kishore,
I agree with you. I'm wondering what would be the inducer and specific
substrate for positive control for phase II enzymes.
Many thanks
Sinisa
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Hi Sinisa
You can use estradiol or 7-hydroxycoumarin as a marker substrate and
rifampin and/or
3-methylcholanthrene as control inducer for UGT induction.
Thanks,
Kishore
Kishore K. Khan, Ph. D.
CellzDirect
8609 Cross Park Drive
Austin, TX 78754
Tel: (512) 615 2332
Fax: (512) 834 7767
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