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The following message was posted to: PharmPK
Dear group,
Could anyone give me advice, or indicate relevant reference about the
following issue: how to deal with CYP inactivation (mechanism-based, or just
simply time-dependent) evidenced in vitro for a compound at preclinical
stage of development, what kind of prediction of drug-drug interactions can
we make, do we have to avoid such problems and stop immediately the
development of such compound as a drug ?
Thanks very much in advance for your help.
Frederic MASSIERE, PhD
Study Director
BIOPREDIC, Rennes, France
E-mail: frederic.massiere.-a-.biopredic.com
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[Two replies - db]
=46rom: alex.macdonald.-at-.pharma.novartis.com
Date: Thu, 29 Nov 2001 09:42:15 +0100
To: david.aaa.boomer.org
Subject: Re: PharmPK CYP inactivation
The following message was posted to: PharmPK
Hi Fred=E9ric,
Below are references which I have found extremely useful for explaining
drug-drug interactions, including mechanism-based, and methods for in-vitro
in-vivo prediction.
1. Ito K, Iwatsubo T, Kanamitsu S, Ueda K, Suzuki H, Sugiyama Y.
Prediction of pharmacokinetic alterations caused by drug-drug
interactions:
metabolic interaction in the liver. Pharmacol Rev 1998;50(3):387-411.
2. Kanamitisu S, Ito K, Sugiyama Y. Quantitative prediction of in
vivo drug-drug interactions from in vitro data based on physiological
pharmacokinetics:use of maximum unbound concentration of inhibitor at
the inlet to the liver. Pharm Res 2000;17(3):336-43.
3. Kanamitsu S, Ito K, Green CE, Tyson CA, Shimada N, Sugiyama Y.
Prediction of in vivo interaction between triazolam
and erythromycin based on in vitro studies using human liver
microsomes and recombinant human CYP3A4. Pharm Res 2000;17(4):419-26.
4. Lin JH. Sense and nonsense in the prediction of drug-drug
interactions. Current Drug Metabolism 2000;1:305-31.
Best Regards
Alex
---
=46rom: "AZHER HUSSAIN"
Date: Thu, 29 Nov 2001 07:53:51 -0600
To: david.-at-.boomer.org
Subject: Re: PharmPK CYP inactivation
The following message was posted to: PharmPK
Hi Frederic,
There are couple of experiments you can do to study mechanism based
CYP inactivation. First one is investigate "Reversible"
metabolsim-dependent inhibition. In such a investigation the Test
Article is is preincubated with the drug and NADPH for 15 min to
allow the generation of metabolites that could inhibit CYP. After
preincubation, the marker substrate (Km) is added and the incubation
is continued to measure residual activity. Preincubations containing
no drug but solvent used to dissolve serves as negative control.
The second experiment you can design is to investigate the
"Irreversible" or "Quasi-Irreversible" metabolism dependent
inhibition. In This, the Test Article (concentration of which causes
less than 30% inhibtion) is preincubated with microsomes at times
concentration and ten times concentration of protein for 15 min with
NADPH. After the preincubation, an aliquot of the preincubation
mixture is diluted ten times to measure residual activity. If the
Test Article is strictly reversible inhibitor it would cause only a
30% inhibition.
We routinely do these experiments at Xenotech as Contract research. I
would suggest you to refer the following reference for a detailed
protocol for drug inhibtion studies. Ajay Madan et al. In vitro
approaches for studying the inhibition of drug-metabolizing enzymes
and indentifying the drug metabolizing enzymes responsible for the
metabolism of drugs. In Drug-Drug Interactions Ed. A. David Rorigues.
pp 217-294. Mercel Decker, Vol 116, New York.
Azher Hussain
Post Doctoral Researcher-Drug Inhibition
Xenotech,LLC
Kansas City, KS
USA
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[Two more replies - db]
=46rom: uceph hijazi
Date: Thu, 29 Nov 2001 21:30:12 +0100 (CET)
To: david.-at-.boomer.org
Subject: Re: PharmPK CYP inactivation
The following message was posted to: PharmPK
Dear Fred=E9ric,
I can indicate to you some references that seems to be
useful in your investigation
Becquemont L, Le Bot MA, Riche C, Funck-Brentano C,
Jaillon P and Beaune P (1998) Use of heterologously
expressed human cytochrome P450 1A2 to predict
tacrine-fuvoxamine drug interaction in man.
Pharmacogenetics 8 : 101-108.
Bourrie M,. Meunier V,. Berger Y, and. Fabre G (1996)
Cytochrome P450 isoform inhibitors as a tool for the
investigation of metabolic reactions catalyzed by
human liver microsomes. J. Pharmacol. Exp. Ther. 277 :
321-332 .
Halpert J R, Guengerich F P, Bend J R, and. Correia M
A (1994) Selective inhibitors of cytochromes P450.
Toxicol. Appl. Pharmacol. 125 :, 163-175.
Iwatsubo T, Hirota N, Ooie T, Suzuki H, Shimada N,
Chiba K, Ishizaki T, Green CE, Tyson CA and Sugiyama Y
(1997) Prediction of in vivo drug metabolism in the
human liver from in vitro metabolism data. Pharmacol
Ther 73: 147-171.
Youssef Hijazi
H=F4pital neurologique et neurochirurgicale
UNITE DE PHARMACOLOGIE CLINIQUE
Service Pharmaceutique
d=E9partement de dosage de m=E9dicaments et de Pharmacocin=E9tique
B.P. Lyon Montchat 69394 Lyon Cedex 03-FRANCE
Tel: 04 72 35 72 45
---
=46rom: Walt Woltosz
Date: Thu, 29 Nov 2001 12:52:33 -0800
To: david.aaa.boomer.org
Subject: Re: PharmPK CYP inactivation
The following message was posted to: PharmPK At 08:38 PM 11/28/01, you wrote=
:
>Could anyone give me advice, or indicate relevant reference about the
>following issue: how to deal with CYP inactivation (mechanism-based, or jus=
t
>simply time-dependent) evidenced in vitro for a compound at preclinical
>stage of development, what kind of prediction of drug-drug interactions can
>we make, do we have to avoid such problems and stop immediately the
>development of such compound as a drug ?
>
There is no need to stop development of a drug just because of CYP
inactivation. There are many drugs now on the market that are
metabolized by CYP 3A4, for example, the bioavailability of which are
influenced by the inactivation of gut 3A4 produced by grapefruit
juice. These drugs are in use every day, and are making money for
their manufacturers. It is "simply" a matter of understanding the
mechanisms and knowing how to deal with them - of course, not really
so "simple" in practice.
In our GastroPlus(tm) software, we simulate the effects of these
enzymes and transporters using Michaelis-Menten kinetics. To simulate
these effects properly, you need to know the Vmax and Km for each
enzyme or transporter in a reference environment - such as in the
experiments your company produces. Those values can then be scaled to
the environments in different regions in the human intestine, as well
as in the liver.
We have demonstrated that with in vitro inputs, good in vivo
predictions can be obtained that include all of the interactions
among nonlinear pharmacokinetic effects, dissolution effects produced
by low (regionally dependent) solubility, and regional variations in
permeability. This has been done with complex drugs that have
multiple metabolic pathways, each saturating at different
concentrations [1].
=46or example, inactivation of CYP3A4 because of food effects or other
drugs can be understood through simulation. In the case of our
analysis of the effects of grapefruit juice on midazolam and
saquinavir, we reduced the Vmax for gut metabolism by an amount equal
to the observed reduction in gut metabolism for each drug. With only
this one change to the inputs, and using pharmacokinetic parameters
derived from literature IV data, predicted plasma concentration-time
was found to closely match observations both with and without
grapefruit juice.
Changing Vmax is one approach, and is based on observed changes in
gut metabolism, but such data will probably not be available in a
preclinical setting. Another approach is to alter the Km of Compound
A by an inhibition constant for the presence of Compound B, so that
Km' =3D Km (1 + Ki)
In GastroPlus, the (1 + Ki) can be input as a Km scale factor. In
vitro measurements of the Vmax and Km for Compound A, and the Ki for
the effect of Compound B on Compound A, can be used to estimate in
vivo effects.
In short, there are ways to understand CYP inhibition (or induction),
as well as transporter inhibition (or induction) through
Michaelis-Menten kinetics. They are not perfect (Michaelis-Menten
itself incorporates some simplifying assumptions), but they seem to
work very well in all cases we've investigated to date. So I would
suggest that, unless you have a good backup compound that is not
affected by CYP, do not stop development of your compound; rather,
seek to understand it through a combination of a few more experiments
and state-of-the-art modeling/simulation.
Walt Woltosz
Chairman & CEO
Simulations Plus, Inc. (SIMU)
1220 W. Avenue J
Lancaster, CA 93534-2902
U.S.A.
http://www.simulations-plus.com
Phone: (661) 723-7723
=46AX: (661) 723-5524
E-mail: walt.aaa.simulations-plus.com
[1] Agoram B, Woltosz WS, and Bolger MB: (2001) Predicting the impact
of physiological and biochemical processes on oral drug
bioavailability. Adv. Drug. Deliv. Rev. 50:S41-S67
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Dear Frederic,
there are two issues to be kept in mind for chemicals undergoing
mechanism-based inhibition of CYPs. One is the potential for
drug-drug interactions due to irreversible inactivation of the
enzyme. The decision whether this has an impact on your development
program should be done on a case by case basis depending on the
extent of inhibition. However, I agree with other participants of
this discussion group that mechanism-based inhibition per se should
not be considered as a reason to discontinue a preclinical
development program.
The second implication of mechanism-based inhibition is the formation
of a reactive metabolite which might either bind to the CYP itself
or, depending on its intrinsic chemical reactivity, leave the CYP
active site and bind to other cell macromolecules. Formation of these
covalently modified enzyme-metabolite complexes is suspected as being
the initial step triggering a cascade of events finally leading to an
immune-mediated idiosyncratic drug toxicity (hepatic failure,
agranulocytosis, systemic lupus erythematosus, hypersensitivity).
However, this affects usually only 1 out of 10-30000 patients and the
understanding of the mechanisms leading to these rare toxicities
(e.g. genetical predisposition, habit, co-medications) are poorly
understood. The issue was extensively discussed during a conference
about two years ago and the general consensus was that most drug
companies do not consider covalent binding as an a priori reason to
stop a program but that care should be taken in the clinical
development program to identify any signs of immune-mediated toxicity.
Best regards
Alex
Back to the Top
Dear Frederic,
there are two issues to be kept in mind for chemicals undergoing
mechanism-based inhibition of CYPs. One is the potential for
drug-drug interactions due to irreversible inactivation of the
enzyme. The decision whether this has an impact on your development
program should be done on a case by case basis depending on the
extent of inhibition. However, I agree with other participants of
this discussion group that mechanism-based inhibition per se should
not be considered as a reason to discontinue a preclinical
development program.
The second implication of mechanism-based inhibition is the formation
of a reactive metabolite which might either bind to the CYP itself
or, depending on its intrinsic chemical reactivity, leave the CYP
active site and bind to other cell macromolecules. Formation of these
covalently modified enzyme-metabolite complexes is suspected as being
the initial step triggering a cascade of events finally leading to an
immune-mediated idiosyncratic drug toxicity (hepatic failure,
agranulocytosis, systemic lupus erythematosus, hypersensitivity).
However, this affects usually only 1 out of 10-30000 patients and the
understanding of the mechanisms leading to these rare toxicities
(e.g. genetical predisposition, habit, co-medications) are poorly
understood. The issue was extensively discussed during a conference
about two years ago and the general consensus was that most drug
companies do not consider covalent binding as an a priori reason to
stop a program but that care should be taken in the clinical
development program to identify any signs of immune-mediated toxicity.
Best regards
Alex
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