- On 28 Mar 2003 at 05:45:22, "Suresh.B.L." (suresh_bl1.at.rediffmail.com) sent the message
The following message was posted to: PharmPK
Dear all,
I am planning to determine the drug concentration in tissue at
various time points during my pharmacokinetic study along with the
plasma level.
What would be the ideal method to determine the tissue conc.
without being either underestimated or overestimated?
Some of the issues which is bothering me is:
Tissue contains the blood and if i take whole tissue- weigh,
homogenize and then extract the drug with suitable solvent? won't
the conc. is overestimated because of the presence of the blood.
I would really appreciate your comments.
Thanks,
B.L.Suresh
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- On 28 Mar 2003 at 17:54:51, "J.W. Mouton" (mouton.-at-.cwz.nl) sent the message
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hi
determination of tissue levels have , in general not much value if you
are not aware in which 'tisue compartment' the drug resides. mixing of
blood, intracellular and extracelular compounds all add together to a
mean concentations but does not tell you anything of the active
concentration where the drug is needed. See various publications of
Otto Cars and Hartmut Derendorf . the only possible exceptin are
perhaps very lipophilic drugs such as amphotericin B (at least in the
animicrobials)
for extracellular tissue concentrations the microdialysis method, if
properly applied is good. has been applied by Markus Muller in Vienna
among others.
For various methods on intracellular concentrations you could contact
tulkens in louvain
johan mouton
Johan W Mouton, MD PhD
Dept Medical Microbiology and Infectious Diseases
Canisius Wilhelmina Hospital C-70
Weg door Jonkerbos 100
6532 sz Nijmegen
The Netherlands
tel + 31 24 3657514
fax + 31 24 3657516
email mouton.aaa.cwz.nl
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- On 28 Mar 2003 at 18:24:14, (Kees.Bol.-a-.kinesis-pharma.com) sent the message
The following message was posted to: PharmPK
Dear Suresh,
If you really want to know the (free) concentration in the tissue than
you have to use microdialysis.
What you probably mean is the amount of drug per gram tissue. I know
people often call this 'concentration'. However, much of this drug is
not freely available. Therefore, be carefull with drawing conclusions
when you want to compare with (free)plasma concentrations or use these
'concentrations' as 'target' concentrations for a certain
pharmacological effect. For toxicity purposes, it is OK to use amount
of drug per gram tissue to see where you might have drug accumulation
and may expect toxicity .
Best regards,
Kees
Kees Bol, PhD
Director Clinical Pharmacokinetics
Kinesis Holding
Lage Mosten 29
4822 NK Breda
The Netherlands
tel: +31 (0) 76 54 80 621
gsm: +31 (0) 65 31 26 153
fax: +31 (0) 76 54 21 777
kees.bol.aaa.kinesis-pharma.com
www.kinesis-pharma.com
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- On 28 Mar 2003 at 09:37:54, "Pan, Weijian" (Weijian.Pan.at.elan.com) sent the message
The following message was posted to: PharmPK
Dear Sir,
Microdialysis method is a good way to probe tissue concentrations. We
have
published a paper for simultaneous sampling of blood and brain tissue
concentrations in an awake freely moving rat model:
Pan, W. J.; Hedaya, M. A.: An animal model for simultaneous
pharmacokinetic
/pharmacodynamic investigations: application to cocaine. Journal of
Pharmacological and Toxicological Methods 39:1-8, 1998.
Wei-jian
Wei-Jian Pan, Ph.D.
Associate Director of Clinical Pharmacokinetics
Clinical Pharmacology and Pharmacokinetics Group
Clinical Development Department
Lusk 2.3 - 3063T
Elan Pharmaceuticals
7475 Lusk Blvd.
San Diego, CA 92121
Tel: (858) 320-7673
Fax: (858) 558-4609
E-mail: weijian.pan.-a-.elan.com
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- On 28 Mar 2003 at 17:10:09, "Dunnington, Shelly" (DunningtonShelly.at.PRAIntl.COM) sent the message
The following message was posted to: PharmPK
Check the literature, I have seen published methods for correction of
blood
volume within certain tissues. I'm sorry, I don't have any references in
hand.
Shelly Dunnington R.Ph., Ph.D.
DunningtonShelly.-a-.praintl.com
913-577-2767
16400 College Blvd.
Lenexa, KS 66219
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- On 28 Mar 2003 at 19:52:33, Rostam Namdari (chista90.-a-.yahoo.com) sent the message
The following message was posted to: PharmPK
Dear Suresh,
There are two ways that you can correct for blood
contribution in the tissues a) use exsanguinated
animals b) use blood volume correction based on
literature values. In my experience, there is not much
difference between tissue concentrations of
exsanguinated and non-exsanguinated rats (cannot
comment on other species). So it may be better if you
use published % blood volume in the tissues/organs of
your interest to correct for the blood contribution.
This method also has its own limitations and sometimes
you end up getting negative numbers particularly at
early time points. For start you can look at Triplett
et al (J Pharm Sci vol 74, No. 9 Sep 1985, P. 1007-1009).>
Rostam
[i.e. Triplett, J.W., Hayden, T.L., Mc Whorter, L.K., Gautam, S.R.,
Kim, E.E., and Bourne, D.W.A. 1985. Determination of Gallium
Concentration in "Blood Free" Tissues Using a Radiolabeled Blood
Marker. J. Pharm. Sci., 74(9), 1007- 1009 :-) db]
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- On 29 Mar 2003 at 06:21:30, "Suresh.B.L." (suresh_bl1.-a-.rediffmail.com) sent the message
The following message was posted to: PharmPK
Dear all,
Thanks everyone for your input.
So if i understand correctly (please correct if i have interpreted
wrongly) the whole organ/tissue consists of Blood
component(capillaries)+Tissue component(Extracellularfluid-20%>+Intracellular).
If i take whole organ and homogenize and determine the drug
concentration, then i will get the mixture(Blood,
Extra/Intracellular) of total(free+bound)concentration of drug.
By knowing the % of blood flow through that particular organ and
do the blood volume correction, then i will get the total drug
conc. in tissue.
i.e.,amt.of drug/gram tissue which will be useful for toxicity
studies(accumulation).
By using Microdialysis technique i can determine the free drug
concentration which is present in tissue extracellular
fluid(interstitial fluid), which is responsible for the
pharmacological effect (if it is the target organ).
Thanks,
B.L.Suresh
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- On 29 Mar 2003 at 06:55:07, "Prof. Walter Wolf" (wwolfw.at.usc.edu) sent the message
Suresh:
There are a number of types of procedures one can use to study drug
biodistribution at various tissues. Let me concentrate on those that
use radiolabeling of the drug for such studies.
One set of approaches is based on the "cut-and-count" mechanism. Your
drug, radiolabeled with either 3H or 14C, is administered and after you
sacrifice the animals at times t1, t2, t3, etc, you collect all types
of tissues and you count the activity. That will give you probably the
most quantitative information on the total activity in all those
tissues and organs you have measured, but it will not discriminate
between the parent drug and its metabolites, nor will it account for
those products (active, inactive or toxic) formed from your compound
where the radiolabel has been metabolized off. But then, if you do not
use radioactivity, you will have to have quantitative assays for all
your products, and be able to measure them in tissues, taking into
account they may be sometimes bound and can not be extracted readily.
A variant in the use of 3H or 14C is to perform whole body
autoradiography. That will reveal tissue localizations that one may not
be readily aware from the cut-and-count approach, inasmuch as one does
not normally collect 100% of all tissues separately.
Finally, there is the use of 11C to perform noninvasive biodistribution
studies of the drug. This approach has significant advantages because
you can quantitate the kinetics of the drug in each individual animal -
and hence, you have multiple time points for each animal, instead of a
single one per animal as when you have to sacrifice them. You can then
also repeat the study to measure modulating or inhibiting effects in a
quantitative manner and using each animal as its own control. The
drawback in this approach is that you need to have access to a
micro-PET and to a cyclotron, and be able to make your compound in a
reasonable time frame (e.g., in less than 3 hrs), inasmuch as the t1/2
of 11C is only 20 minutes.
Naturally, if your drug contains other atoms (e.g., F), you may have a
whole series of other tools available for such studies.
Professor Walter Wolf, Ph.D.
Distinguished Professor of Pharmaceutical Sciences
Director, Pharmacokinetic Imaging Program
Department of Pharmaceutical Sciences, School of Pharmacy
University of Southern California
1985 Zonal Ave., Los Angeles, CA 90089-9121
E-Mail: wwolfw.-a-.usc.edu
Telephone: 323-442-1405
Fax: 323-442-9804
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- On 31 Mar 2003 at 06:44:35, "Walter E. Stumpf" (stumpfwe.aaa.email.unc.edu) sent the message
The following message was posted to: PharmPK
Dear Dr. Suresh,
In your quest for organ/tissue distribution during your pharmacokinetic
studies, please consider the following test to validate your conclusion
from the advice received by a number of pharmacokineticists:
Take a compound known to be localized to specific receptor sites. Test
your microdialysis technique and hypothesis as stated by you:
"Using the microdialysis technique I can determine the free drug
concentration which is present in tissue extracellular fluid
(interstitial fluid), which is responsible for the pharmacological
effect (if it is the target organ)."
For instance, take one of the steroid hormones with well-known receptor
distribution, like 1,25-dihydroxyvitamin D3. There exist target tissues
in the brain, spinal cord, heart, skin, stomach and many other organs -
clearly demonstrated and documented with receptor micro autoradiography
(Stumpf WE, Histochemistry and Cell Biology (1995)104:417-437). Find
out, whether the "free drug concentration" in these target tissues can
be determined and whether the evidence thus obtained is "responsible
for the pharmacological effect."
The subject matter of target tissue pharmacokinetics has been discussed
at boomer.org in January 2001. From the answers obtained at that time I
concluded: (in vivo) pharmacokinetics of target tissues is not
determined in the common ADME studies. In vivo pharmacokinetics cannot
be determined (with some exceptions) by radioassays-HPLC, whole body
autoradiography, in vivo scanning, and biochemical homogenization.
Methods are required with high resolution and high sensitivity that
leave the tissue topography intact and retain the compounds at their in
vivo receptor sites.
In vivo tissue localization of drugs is an old and recurrent problem.
Lloyd Roth at the University of Chicago, after listening at scientific
meetings to pharmacologists that presented data on drug distribution
with disregard for tissue composition, walked up to them and handed his
business card which read on its back: "Don't homogenize the brain, the
brain you are homogenizing may be your own."
This topic is addressed in a forthcoming book (Stumpf WE: Localization
of Drugs in Tissues and Cells, IDDC-Press).
Walter E. Stumpf
Chapel Hill, NC
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- On 31 Mar 2003 at 08:48:07, "Tata, Prasad N" (Prasad.Tata.-at-.TycoHealthcare.com) sent the message
The following message was posted to: PharmPK
Dear All:
Text Book: Casarett and Doull's Toxicology - The Basic Science of
poisons
Ed. Curtis D. Klaassen Publisher McGraw Hill discuss these issues in
detail.
Prasad Tata
Mallinckrodt, Inc.
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- On 31 Mar 2003 at 11:14:39, "Prof. Walter Wolf" (wwolfw.at.usc.edu) sent the message
Walter:
While I agree with most of your comments [and I especially relished
your mention of Lloyd Roth. He was a true pioneer], I have one serious
disagreement.
In your statement: "In vivo pharmacokinetics cannot be determined (with
some exceptions) by radioassays-HPLC, whole body autoradiography, in
vivo scanning, and biochemical homogenization" you include in-vivo
scanning as one that, in your opinion, does not allow for the
determination of in-vivo pharmacokinetics. I disagree strongly with
this statement. Indeed, the reason you can ONLY get good data about
in-vivo PK is when you do noninvasive measurements that do NOT perturb
the system. And such noninvasive imaging measurements include nuclear
imaging (scanning) and MRI and MRS measurements. Perhaps optical, if
and when you can get sufficient penetration.
Thus, noninvasive measurements can and do provide very pertinent and
reliable in-vivo PK data.
Professor Walter Wolf, Ph.D.
Distinguished Professor of Pharmaceutical Sciences
Director, Pharmacokinetic Imaging Program
Department of Pharmaceutical Sciences, School of Pharmacy
University of Southern California
1985 Zonal Ave., Los Angeles, CA 90089-9121
E-Mail: wwolfw.-a-.usc.edu
Telephone: 323-442-1405
Fax: 323-442-9804
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- On 31 Mar 2003 at 14:32:18, Walter E Stumpf (stumpfwe.-a-.email.unc.edu) sent the message
The following message was posted to: PharmPK
Walter:
yes, non-invasive procedures would be optimal.
But do current non-invasive procedure provide the necessary sensitivity
and resolution??
Please, test your system and show estradiol receptor binding in the
hippocampus, the bed nucleus of the stria terminalis, thymus reticular
cells, atrial cardiomyocytes. These are the places, where we find the
receptor binding that can be followed. Or: Show vitamin D localization
in
thyrotropes of the pituitary.
If you can do that, then you can apply your procedure to tissue
pharmacokinetics, if not - and
that is where we stand - then you have to use a microscopic technique,
the one that I developed together with Lloyd Roth and developed further
during that past 30 years. The many discoveries made with receptor
micro autoradiography are proof of its utility.
With best wishes, Walter E. Stumpf
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- On 31 Mar 2003 at 18:08:10, stumpfwe.at.email.unc.edu ("Walter E. Stumpf") sent the message
The following message was posted to: PharmPK
To Walter Wolf
To clarify and not to be misunderstood: With non-invasive scanning you
can do
pharmacokinetic studies, of course. The subject matter related to Dr.
Suresh's
question, however, is pharmacokinetics of target tissue drug binding.
And for that
the resolution for detecting low capacity - high specificity sites must
be
demonstrated. Not just any pharmacokinetic data that are not directly
related to
specific receptor binding, i.e., sites of action. Here is the problem!
Walter E. Stumpf
2612 Damascus Church Rd.
Chapel Hill, NC 27516
Tel/Fax (919) 942-8646
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