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Hi all
I am wondering about an experimental design to determine whether a
compound is P-gp substrate. Our lab is not equipped for in vitro cell
culture techniques, I would like to do this in vivo. Thanks in advance
for your suggestions.
satheesh anand
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Dear Satheeshanand,
Various publications reported the use of excised intestinal segments
(everted sac and ussing chambers) of rat/mice for carriying out P-gp
substrate screening. Further, in situ studies were also reported to be
more effective and correlate well to in vivo relevence of P-gp in
limiting drug transport (Adachi etal.). The refence given below used
all these techniques in demonstrating P-gp mediated transport of
paclitaxel.
Varma MV, Panchagnula R. Enhanced oral paclitaxel absorption with
vitamin E-TPGS: Effect on solubility and permeability in vitro, in situ
and in vivo.
Eur J Pharm Sci. 2005.
Best wishes
Varma Manthena
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Dear Satheeshanad,
In theory; you can assess P-gp substrates with a microdialysis
approach, whereby you investigate brain concentrations by means of
intracerebral microdialysis. The use of P-gp blockers such as e.g.
verapamil, GF120918, PSC833 or others can help do distuingish
substrates from non substrates. But as you probably know, the use of
these is limited due to toxicity problem and also non P-gp related
transport. So you will not be able to have an absolute answer.
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The following message was posted to: PharmPK
Dear Dennis,
Could you provide me any literature on this P-gp and microdialysis
approach.
Verapamil may be a more suitable inhibitor (depending upon its KI/IC50
as
its pharm/tox effects are well known.
Also, may I take this opportunity to mention about the forthcoming 2-day
workshop on microdialysis on Nov 4-5, 2005 in Nashville, TN immediately
preceding the AAPS Annual Meeting.
Regards,
Chandra Chaurasia
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The following message was posted to: PharmPK
Dear Satheesh,
If you could get your hands on mdr1 null mice (mice deficient of the
Mdr1 Pgp; from Taconic 273 Hover Avenue Germantown, NY 12526-9608. Tel:
518-537-6208 or (888) TACONIC FAX: 518-537-7287 Email:
custserv.-a-.taconic.com Internet URL: http:// www.taconic.com) and the
appropriate wild-type control mice) you may be able to do a few studies
that could give you a good idea. (Have a look at this reference Huisman
MT et al., Mol Pharmacol. 2001 Apr;59(4):806-13, it could help too.)
Compare both PK and brain penetration (so always try to get the brain to
plasma ratio as well) of your drug (preferably IV)in presence or absence
of e.g. G918 (say 25 or 50 mg/kg PO, maybe 1 to 2 hours before dosing of
the test drug). This may give you an idea whether CNS penetration of
your compound may depend on Pgp function.
Hope it helps.
Regards
Hans Smit
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E.g.
Br J Pharmacol. 1999 Oct;128(3):563-8. The role of P-glycoprotein in
blood-brain barrier transport of morphine: transcortical
microdialysis studies in mdr1a (-/-) and mdr1a (+/+) mice. Xie R,
Hammarlund-Udenaes M, de Boer AG, de Lange EC.
Currently we are writing an article about morphine in vivo MD studies
with and without GF120918; and loperamide in the same model.
Verapamil is a suitable compound, although its inhibitory properties
are not as good as the P-gp inhbitors as GF120918 or PSC833. There
are plenty of sources avaibalble about this.
Goodluck,
Dennis
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Dear Sateesh,
The in-vivo experiments to evaluate the P-
glycoprotein substrate specificity of your compound can be designed
as follows:
1.Establish the oral pharmacokinetic properties in P-gp knockout
(mdr1a (-/-) mice to confirm whether your compound is a substrate or
not (or)
2. Co-administer your compound orally either with verapamil or any
other specific p-gp inhibitor
(Research papers have already been published on those lines for your
reference) or
3.Determine the oral PK of any known p-gp substrate viz., Etoposide,
doxorubicin etc., in presence and absence of your test compound and
observe the PK parameters.
Insight in to the literature reveals that the hydrogen bond acceptor
or electron donor is an important chemical feature associated with p-
gp substrate activity.Interestingly, the p-gp substrates are observed
to have high molecular weights, lower log Kow values, and greater
hydrogen-bonding potential due to hydrogen bond donors. Cautiously
you may use in-silico tools viz., ADME boxes from algorithms or
similar ADMET tools to predict the possiblity.
Good Luck.
Regards,
Syed Mustafa,
Dept of Pharmacokinetics and Metabolism,
Orchid RND centre,
Plot No:476/14,
Old Mahabalipuram Road,
Sholinganallur,
Chennai: 600119
syedmustafa.at.orchidpharma.com
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The following message was posted to: PharmPK
Bonjour
Some years ago we published a very simple, quick and cheap method to
check for Pgp activity in the intestine. Put in your substrate plus /
minus verapamil. It will also tell you about the potential oral
availability.
Ref: "The improved everetd gut sac: a simple method to study
intestinal P-glycoprotein"
Barthe, L , Bessouet, M., Woodley, J.F. and Houin, G.
Int. J. Pharm. 173 (1998) 255-258
John Woodley
John Woodley
Laboratoire: Cinetique des Xenobiotiques
Faculte des Sciences Pharmaceutiques
35 chemin des Maraichers
31062 Toulouse, France Tel/Fax: + 33 (0)562256885
woodley.at.cict.fr
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