PharmPK Discussion - Relative Contribution of CYP2C19 and CYP3A4 to Omeprazole 5-Hydroxylase Activities in HLM

• On 17 Mar 2008 at 17:38:53, "Sean Xiang Wu" (swu.aaa.exelixis.com) sent the message
  

  
  Hello,
  
  Omeprazole 5-hydroxylation is an acceptable in vitro experiment to
  monitor CYP2C19 activity by FDA.  When we tried to develop a CYP2C19
  inhibition assay with omeprazole as substrate and human liver
  microsome as enzyme source, we noticed that many CYP3A4 inhibitors
  showed inhibitory activities.  It=92s known that both CYP2C19 and CYP3A4
  are involved in omeprazole 5-hydroxylation.  Even though CYP2C19 may
  play a major role, but its enzyme content in HLM is much lower than
  CYP3A4 (10-60 fold).  Therefore, most of 5-hydroxyomeprazole was
  probably made through CYP3A4 pathway at our conditions.
  
  What is the relative contribution of the two enzymes to omeprazole 5-
  hydroxylation when HLM is used?  How can we optimize assay conditions
  to make it more CYP2C19 specific?
  
  Thanks for your help,
  
  Sean Wu
  Exelixis
  
  

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• On 18 Mar 2008 at 08:02:30, "Sean Xiang Wu" (swu.aaa.exelixis.com) sent the message
  

  
  The following message was posted to: PharmPK
  
  Dear Mansi,
  
  http://www.pharmgkb.org/do/serve?objId=PA152530846&objCls=Pathway
  
  Enantiomer/Enantiomer Interactions between the S- and R-Isomers of
  Omeprazole in Human Cytochrome P450 Enzymes: Major Role of CYP2C19 and
  CYP3A4, JPET 315:777-787, 2005
  
  Thanks,
  
  Sean
  
  

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• On 22 Mar 2008 at 11:14:41, "Wichittra Tassaneeyakul" (wichittra.tassaneeyakul.aaa.gmail.com) sent the message
  

  
  Dear Sean Wu,
  Omeprazole hydroxylation shows biphasic kinetic.  Suugest that at
  least two enzymes are involve in this reaction.  It has been reported
  by Andersson et al. that CYP2C19 is the high
  affinity enzyme nd CYP3A4 is the low affinity enzyme.  So it depends
  on the substrate concentration of omeprazole that you used in the
  incubation. KM for high affinity 5-omeprazole hydroxylation is around
  9 micro Molar where as Km for low afinity is around 175 micro Molar.
  Thus, you need to use low concentration of omeprazole (5-10 uM) if you
  want to measure activity of CYP2C19. You could find details in the
  article below.
  
  1. Andersson TA, Miners JO, Tassaneeyakul W, Tassaneeyakul W, Veronese
  ME, Meyer, UA. Birkett DJ.  Identification of human liver cytochrome
  P450  isoforms mediating omeprazole metabolism.  Br J  Clin Pharmacol
  1993; 36: 521-530.
  2. Tassaneeyakul W, Guo L, Fukuda K, Ohta T, Yamazoe Y. Inhibition
  selectivity of grapefruit juice components on human cytochromes P450.
  Arch Biochem Biophys 2000; 378: 356-363.
  
  Wichittra
  
  

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• On 24 Mar 2008 at 22:50:06, "Sean Xiang Wu" (swu.aaa.exelixis.com) sent the message
  

  
  The following message was posted to: PharmPK
  
  Dear Wichittra,
  
  Thanks for your explanation and references.  We did try to adjust
  omeprazole concentration to shift contribution towards CYP2C19.
  Unfortunately, we still observed good correlation between HLM and
  recombinant CYP3A4 even when omeprazole concentration was 0.1-1 uM.
  
  Sean
  
  

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