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We have been trying to measure drug concentrations in tumors. The problem we
run into is the blood vessels surrounding the tumor. If we grind the tumor
with the blood vessels, we will be measuring some composite value of the
concentration of drug in tumor and in blood. We have tried to remove the blood
vessels surrounding the tumor but have not been able to do it successfully. I
was wondering if anyone in this group has experience in measuring the
concentration of drug in tumors with a significant amount of vasculature. Is
there a way to remove the vasculature or correct for the contribution of drug
in blood to calculate the concentration in tumor? Any guidance you might have
on this topic will be appreciated. Please include any references that discuss
this issue.
Thanks,
Chetan D. Lathia
Pharmacokinetics and Drug Metabolism
Parke-Davis Pharmaceutical Research
E-mail: Lathiac.-a-.AA.WL.COM
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> was wondering if anyone in this group has experience in measuring the
> concentration of drug in tumors with a significant amount of vasculature. Is
> there a way to remove the vasculature or correct for the contribution of drug
> in blood to calculate the concentration in tumor?
This enquiry begs the question "Why do you want to measure drug concs in
tumour?". Assuming you want to know the conc at the site of action, even
if you could find a way to measure 'blood free' conc in a tumour
you still do not know the conc at the site of action but just the
average conc in all the (non-blood) tissues (which depending on the
tumour could be quite variable).
This average conc is not the 'true' conc at the
site of action but just represents binding and partition among all the
tissues in the sample which would only resemble the conc at the site of
action by very good luck.
It is my personal opinion that all attempts at so
called tissue conc measurements based on grinding up hetergenous tissue
masses are a form of fool's gold. Unless you have some good reason to
believe that there is active transport to the site of action (or active
transport out of the site) then concs at the site of action will be
proportional to unbound conc in blood at steady state - so you may as
well measure blood concs and forget messing around with the tissue.
So please let us know why you think blood free tissue concs would be
useful to you.
Note that you can get an approximate answer by injecting the animal
with a marker substance that does not cross blood vessels, measures its
conc in blood and the amount in the tissue and thus work out the volume
of blood in the tissue. If you know the blood conc of your drug then you
can figure out the amount of your drug in the blood in the tissue.
Similar principles can be used to work out the amount of extra-cellular
drug.
--
Nick Holford, Dept Pharmacology & Clinical Pharmacology
University of Auckland, Private Bag 92019, Auckland, New Zealand
email:n.holford.aaa.auckland.ac.nz tel:+64(9)373-7599x6730 fax:373-7556
http://www.phm.auckland.ac.nz/Staff/NHolford/nholford.html
Back to the Top
> We have been trying to measure drug concentrations in tumors. The problem we
> run into is the blood vessels surrounding the tumor. If we grind the tumor
> with the blood vessels, we will be measuring some composite value of the
> concentration of drug in tumor and in blood. We have tried to remove the
>blood
> vessels surrounding the tumor but have not been able to do it
>successfully. I
> was wondering if anyone in this group has experience in measuring the
> concentration of drug in tumors with a significant amount of vasculature. Is
> there a way to remove the vasculature or correct for the contribution of drug
> in blood to calculate the concentration in tumor? Any guidance you might
>have
> on this topic will be appreciated. Please include any references that
>discuss
> this issue.
We use a correction factor for blood volume to account for the amount of
material residing in the vasculature of that tissue. A detailed review on
this can be found in the book entitled "Liposome Technology", 2nd edition,
Volume 3 (Interactions of liposomes with biological milieu) edited by G.
Gregoriadis. See chapter 3, the article written by Marcel Bally, Lawrence
Mayer, Micheal Hope, and Rajiv Nayar, "Pharmacodynamics of liposomal drug
carriers: methodological considerations" (1993, CRC Press). Correction
factors (red cell volume and blood volumes) for tumor and non-tumor
bearing animals are listed here. Trust this is of help.
Rajesh Krishna
BC Cancer Agency
Vancouver, BC
http://members.tripod.com/~rkrishna/index.html (A liposome page)
Back to the Top
On Friday, 28 Feb 1997 11:51:25 -0600 Chetan D. Lathia,wrote by way of David_Bourne
To the Multiple recipients of PharmPK on the
Subject: How are tumor drug concentrations measured?
>We have been trying to measure drug concentrations in tumors. The
>problem we run into is the blood vessels surrounding the tumor. If we
>grind the tumor with the blood vessels, we will be measuring some
>composite value of the concentration of drug in tumor and in blood. We
>have tried to remove the blood vessels surrounding the tumor but have
>not been able to do it successfully. I was wondering if anyone in this
>group has experience in measuring the concentration of drug in tumors
>with a significant amount of vasculature. Is there a way to remove the
>vasculature or correct for the contribution of drug in blood to calculate
>the concentration in tumor? Any guidance you might have on this topic
>will be appreciated. Please include any references that discuss this
>issue.
The issue raised here by Dr. Lathia has quite interesting implications, and
is one that we have been concerned for many years. Originally, in the
1970's, we would express our biodistribution data in a variety of forms:
% ID [injected dose]/g of tissue, total
% ID/g of tissue, blood corrected
% ID/organ, total
% ID/organ, blood correct
% RD [retained dose]/ of tissue, total, etc, etc,
where the injected dose [ID] is the total amount of the drug administered to
the animal, and where the retained dose [RD] is the amount of the drug
remaining at time t. Each of these values may have some relevance. See for
example:
Synthesis and Distribution of 195mPt-cis-Dichlorodiammine Platinum (II).
W. Wolf and R.C. Manaka. J. Clin. Hematol. Oncol., 7, 79-85 (1977).
There are, however, many more issues to consider that an simple
correction, and assuming that the tumoral blood pool is totally "external"
to the tumor cell mass may not be correct.
1) Which is the specific tumor that is being studied? Not all tumors are the
same. Their growths may be significantly different, their
microvasculature structure may have differences, as we have begun to
show. See:
Pharmacokinetic Imaging of Animal Tumors can be Used to Evaluate
the Pathophysiological Compartments that Regulate Drug Delivery.
Alfredo R. Sancho, James A. Dowell and Walter Wolf. Abstract PPDM 8245,
presented at the 1996 Meeting of the AAPS. Pharm.Res. 13:S-454, 1996.
More of this work, where we have shown how it is possible to measure,
noninvasively, the vascular space in a given tumor mass, will be presented
at the upcoming AACR meeting.
2) We have also noted that an understanding of the tumoral microvascular
space may be quite relevant to our understanding of drug transport into
tumors, and this can be graphically seen in two posters presented at the
recent meeting on the Future of NMR in Medicine. These posters,
Magnetic Resonance Spectroscopy (MRS) allows Pharmacokinetic
Imaging (PKI) Studies in Human Tumors. Walter Wolf, Victor
Waluch, Cary A. Presant and Hyun K. Kim., and Magnetic Resonance
Imaging in Functional Studies of Human Tumors. Hyun K. Kim,
Victor Waluch, Alfredo Sancho, Cary A. Presant and Walter Wolf,
both of them presented at the first Internet Conference on the Future of
Magnetic Resonance in Medicine, Japan, January 27-February 3, 1997
may be viewed at our URL -
http://www.usc.edu/hsc/pharmacy/pkimaging/posters.html
Thus, rather than ignoring the microvascular space, we postulate that it
may, together with the tumor's interstitial space and the tumor's
intracellular space, be one of the important elements to consider in the
rather complex picture of how drugs get delivered to tumor cells.
The availability of pharmacokinetic imaging methods allows the direct
measurement of such spaces in individual animals - and in humans.
I would be glad to answer any specific questions, either directly or through
this group.
=========================================================================| Professor Walter Wolf, Ph.D. E-Mail: wwolfw.at.hsc.usc.edu |
| Director, Pharmacokinetic Imaging Program |
| Department of Pharmaceutical Sciences Telephone: 213-342-1405 |
| University of Southern California Fax: 213-342-9804 |
| 1985 Zonal Ave., Los Angeles, CA 90033 |
| |
| Center for Noninvasive Pharmacology, Los Angeles Oncologic Institute |
| MRI at St. Vincent Medical Center Telephone: 213-484-7235 |
| 2131 Third St., Los Angeles, CA 90057 Fax: 213-484-7447 |
========================================================================------------------------------
Date: Mon, 3 Mar 1997 12:24:29 -0600
From: eddyjh.at.moose.ncia.net (John Eddy)
Subject: Ideal Body Weight Calculations
>I would like to examine the various BSA formulas; we are having quite a
argument at work over this. Historically our hospital utilized the charts
or calculators found in many texts that are based on the 1916 Annals of
Internal Medicine formula of Dubors and Dubois. When pharmacy became more
involved we used the numbers in our computer software package. This formula
involves the square root..... I wish I had the facts here at home. Needless
to say, the numbers come out different; I expect if I use the formula in
this newsgroup I will get a third figure to work with. I am not a
scientist, just a pharmacist, but I would like to know alot more about the
different bsa formulas and their relationship, if any. Likewise, what
happens to the toxicity of my chemo doses if I use one over the other. I
guess I'm really looking to defend the software program and prove the non
clinicians at the hospital they should stick to their areas of expertise.
Thanks for your help
Back to the Top
Not knowing the characteristics of your tumor, I'm going out on a limb as I
make the following coments.
This is a classic dilemma faced by anyone who has tried to target
tumors. The tumor vasculature is giong to vary from tumor to tumor, how
the tumors differentiate, etc.. When I used to work on
radioimmunoconjugates, we washed the tumors as best as we could
with saline/PBS etc and then counted the tumors for radioactivity. To
show specific tumor uptake we would use a control that was
non-binding. This would give us an idea of blood pool effects. Without
belaboring much on this issue, here are a few suggestions:
1. If this is a targeted agent, then the solution is easy. Use a similar
molecule but without any binding specificity. I am assuming here that the
targeting vector is distinct from the drug. Otherwise, try the following:
2. If it is a small molecule then determine the concentration of your drug in
the tumor (before extensive washing), following washing. You might
even want to slice the tumor open, if it is a solid tumor. If you then
compare the Tumor to Blood ratios, you ought to get a feel for specific
tumor concentrations. Of course, this will only be an estimate at best :-(
There is no easy solution to your problem, unfortunately.
HTH
Vinay Desai
Nycomed Inc.
Back to the Top
I don't have any experience in measuring drug concentrations in tumors. But
we use microdialysis to measure drug concentrations in liver. People have
used microdialysis in tumors also. You implant a microdialysis probe which
consist of a dialysis membrane. The probe is perfused with a solution
isotonic to the tumor medium. The small molecular weight drug will pass
through the dialysis membrane and be carried away by the perfusion fluid
which can be collected and assayed. The samples are protein free so you
don't need any sample preparation or tissue grinding. Since you can place
the probe anywhere in the tumor, vasculature is not a problem. Following are
some references on tumors.
1. Microdialysis Sampling in Tumor and Muscle: Study of the Diposition
of 3-Amino-1,2,4-Benzotriazine-1,4-DI-N-Oxide (SR 4233) R. K. Palsmeier and
C. E. Lunte, Life Sciences YEAR: 1994 VOLUME: 55(10) PAGES: 815-825
2. Pharmacokinetic and Metabolism Studies of 3-Amino-1,2,4-
Benzotriaznine-1,4,Di-N-Oxide In Solid Tumors By In Vivo Microdialysis. R.
K. Palsmeier and C. E. Lunte: Internat. Symp. on Microdialysis and Allied
Analytical Techniques. (In: BAS' Current Separations) YEAR: 1993 VOLUME:
12:2 PAGES: Abst# 55
You can find more information on microdialysis on our web site at
http://www.bioanalytical.com.
Chandrani Gunaratna, Ph.D.
Senior Research Chemist
Bioanalytical Systems
2701 Kent Avenue
West Lafayette, IN 47906
Phone: (317)463-4527
E-Mail: prema.at.bioanalytical.com
Back to the Top
Dr. Gunaratna:
We had thought of using microdialysis to measure antineoplastic drug
concentration in and around tumors of various sizes that had been implanted
in rats. We hesitated and did not use this method, instead we used true
noninvasive imaging methods.
My question is: How invasive is the probe? Does it perturb to a large
extent the "normal" (as normal as a tumor can be) growth and patho-histology
of the tumor? Does it cause extra leakiness?
Thank you,
Alfredo R. Sancho,
USC-PK Imaging Ctr.
Back to the Top
Nick Holford:
I was woundering if you might suggest something for the following scenario:
I agree with you that more traditional methods of measuring drug
concentration are available and upto reacently used extensively. Such
methods as blood sampling, urine output, and others are considered to be the
"traditional" pk drug measuring methods, from these measurements
"assumptions" through mathematical analysis and "data-processing" are made
to "estimate" or "approximate" drug concentrations in various tissues
-including tumors- can be done.
My question is, when working with patients and with novel drugs or
"cocktails" of antineoplastic drugs, which are highly toxic. How do you
justify pushing the limits of such drugs MTD into a patient so to sample
blood and estimate the amount of drug reaching the tumor? Whould it not be
better to somehow administer lower dosages -less toxic, less costly, less
possibilities of negative side effects- of the same drug/s and measure
noninvasively the amount of the drug in the tumor? How else can you not
perturb the system -possibly freeing tumor cells into the body- yet obtain
meaningful data? How else can you not find yourself making leap-of-faith
assumptions from calculations?
You see, after studying tumors for several years, no two tumors are alike,
even if they are of the same origin and found in the same patient and tissue.
Each tumor will grow in its own radically different style and madness.
Particularly their microvascular network. Being that the case would you not
need to measure exactly -or as precise as possible- the %ID of your
administered drug? For instance, what would happen if you make assumptions
based on blood sampling and due to the odd and unique pathophysiological
factors found in tumors -e.g. microvasculature- the drug never reaches the
tumor in any therapeutic amount? And clinicians continue the protocol for
several months. Would it not be nice to know if the drug is going to reach
the tumor, this from a single dose measurement? In that manner the
researcher/clinician group can select the most effective drug for that
patient?
Thank you for you response and time, I just thought to put my two tid-bits
into the discussion.
Alfredo R. Sancho,
USC PK-Imaging Ctr.
Back to the Top
We've used microdialysis to measure brain extracellular drug
concentrations (Scheyer, R.D., During, M.J., Hochholzer, J.M., Spencer,
D.D., Cramer,
J.A. and Mattson, R.H. Phenytoin concentrations in the human brain -
an in vivo microdialysis study. Epilepsy Res. 18:227-232,1994.,
Scheyer, R.D., During, M.J., Spencer, D.D., Cramer, J.A. and Mattson,
R.H. Measurement of carbamazepine and carbamazepine epoxide in the
human brain using in vivo microdialysis. Neurology
44:1469-1472,1994). After several hours, microdialysis
concentrations reflect the extracellular space, rather than the
vascular compartment, although there may be some residual
"leakiness" for longer periods.
Restoration of the integrity of tumor vascularity might be
different, and might vary with tumor type. Possibly of greater
importance is what you wish to measure. Microdialysis is a useful
probe of the extracellular space. While this is a region of interest
for many drugs, it may not be of particular importance for
antineoplastic compounds.
- Richard Scheyer
Richard Scheyer, M.D.
Dept. Neurology, Yale School of Med.
P.O. Box 208018, New Haven, CT 06520-8018 USA
Richard.Scheyer.-at-.yale.edu
Back to the Top
Alfredo,
Thank you for your comments and stimulating questions.
> I agree with you that more traditional methods of measuring drug
> concentration are available and upto reacently used extensively. Such
> methods as blood sampling, urine output, and others are considered to be the
> "traditional" pk drug measuring methods, from these measurements
> "assumptions" through mathematical analysis and "data-processing" are made
> to "estimate" or "approximate" drug concentrations in various tissues
> -including tumors- can be done.
These 'traditional' methods have been used successfully to understand
how the time course of the effects of anti-tumour agents are linked to
drug dose via models of drug concentration in plasma and at the site of
action.
In the clinical pharmacology setting this has been helpful in
identifying the predictable components of the dose-effect relationship
e.g. differences in clearance from individual to individual and from
occasion to occasion within the same individual.
>
> My question is, when working with patients and with novel drugs or
> "cocktails" of antineoplastic drugs, which are highly toxic. How do you
> justify pushing the limits of such drugs MTD into a patient so to sample
> blood and estimate the amount of drug reaching the tumor? Whould it not be
> better to somehow administer lower dosages -less toxic, less costly, less
> possibilities of negative side effects- of the same drug/s and measure
> noninvasively the amount of the drug in the tumor?
I did not advocate 'pushing the limits of drugs MTD...so to sample blood
and estimate the amount of durg reaching the tumor'. The kinds of
studies that I am referring to may be done during Phase I (when the MTD
is determined) but have been done and have been most informative at
doses which are thought to provide optimal clinical responses.
My question about the use of so called tumour concentrations should be
considered in the setting of giving the same dose to the same patient.
In that case, what could be learned from 1) blood concentrations 2)
invasive tumour concentrations (as proposed originally in this thread)
3) non-invasive imaging? Which would help understand the effects of the
drug?
I would think that a combination of 1) and with the others would be most
informative but the specificity of the average tumour concs or the
imaging pictures as markers of concs at the site of action would
determine their relative contributory merits. Approaches which use 1)
plus some marker of drug effect and a suitable model have been very
helpful (see below).
> How else can you not
> perturb the system -possibly freeing tumor cells into the body- yet obtain
> meaningful data?
A combined approach should be considered, of course.
> How else can you not find yourself making leap-of-faith assumptions
> from calculations?
Most of the assumptions I need to make are based on strong biological
and chemical a priori knowledge. I mentioned one such assumption in my
original posting i.e. equal free water concs in blood and at the site of
action when at steady state.
What leap-of-faith assumptions are you thinking of?
> You see, after studying tumors for several years, no two tumors are alike,
> even if they are of the same origin and found in the same patient and tissue.
> Each tumor will grow in its own radically different style and madness.
> Particularly their microvascular network. Being that the case would you not
> need to measure exactly -or as precise as possible- the %ID of your
> administered drug? For instance, what would happen if you make assumptions
> based on blood sampling and due to the odd and unique pathophysiological
> factors found in tumors -e.g. microvasculature- the drug never reaches the
> tumor in any therapeutic amount? And clinicians continue the protocol for
> several months. Would it not be nice to know if the drug is going to reach
> the tumor, this from a single dose measurement? In that manner the
> researcher/clinician group can select the most effective drug for that
> patient?
The issues of variability, from patient to patient (or tumour to tumour
if you wish), are indeed very important. They have been the focus of
substantial efforts among the clinical pharmacology PKPD community.
Large fractions of variability are currently without explanation so any
methods that offer insights into this are very welcome. It should be
noted however that PKPD studies have been critical in understanding that
the variability in drug disposition and response from one treatment
occasion to another can be so large that the first treatment response
only predicts a very small part of the second treatment.
e.g.
Karlsson MO. Port RE. Ratain MJ. Sheiner LB. A population model for the
leukopenic effect of etoposide. Clinical Pharmacology & Therapeutics.
57(3):325-34, 1995 Mar.
reported occasion to occasion variability in EC50 was so large that the
same average conc should be targeted for all patients but the variation
in clearance was small enough that use could be made of plasma concs
from the first course to predict the dose needed for the second.
An analysis based only on tumour site sampling could only have addressed the PD
part (eg very variable EC50) while a combined approach of PK and PD
(using leukopenia as the effect) revealed the relative important of PK
as a way to optimise response.
So please understand that I am not saying tumour concs are a waste of
time. But neither do they offer the gold standard for understanding how
to use anti-cancer agents. I would still be interested in hearing from
the originator of the thread about what was hoped for by estimating blood
free concentrations in a tumour.
It is also important to appreciate that different approaches are needed
at different stages of understanding the pharmacology of a drug. During
drug discovery tumour concs may be a very helpful screening tool for
certain compounds while in phase III clinical trials blood concs may be
critical in understanding what the optimal dose might be.
--
Nick Holford, Dept Pharmacology & Clinical Pharmacology
University of Auckland, Private Bag 92019, Auckland, New Zealand
email:n.holford.-a-.auckland.ac.nz tel:+64(9)373-7599x6730 fax:373-7556
http://www.phm.auckland.ac.nz/Staff/NHolford/nholford.html
Back to the Top
Nick:
I am glad that the vagueness is cleared. Moreover I agree with you that a
multidisciplinary method or approach is needed to truly and fully understand
what happens in and around the tumour site.
Let me throw this question in the pile:
1) If clearance is a way of calculating the amount of drug left in the blood
compartment, hence how much is left in the tumor... what model are you using
to compare the tumor compartment? There is no tissue in the body that has
the same infrastructure as that of tumours. We could not even compare it to
the tissue in which the tumour is in. Therefore, either assumptions are made
or invasive measurements are needed.
Now if we agree that there is great patient intervariability and that
measurements of clearance can be used as an indicator for tumor trapping of
the drug. How accurate is this? Is is not contradictory? In the clinical
scenario, would a one "common" or "average" dose -statistically calculated
from previous patients- and noninvasive measurements be in order? That is
the first does measured at the tumor site itself, might be a indicator -if
not a predictor- of tumor response.
You see I put myself in the patients position... do I want to go through an
entire protocol of chemotherapy without know if it is effective or not. Based
solely on "statistical analysis" of previous patient tumor response data. Or
do I want to take a single dose ("average" dose) and measure its prescence or
lack of it in the tumor itself. This would tell the clinician that the drug
is not effective for that tumor, in that tissue, with those
histo-pathological characteristics. Which in turn would save money, time and
pain, leading to possibly finding the drug that does have the desired
therapeutic effect.
Now that this has been said, I understand the origin of this thread as
someone coming from this point. That is, how can we measure the
antineoplastic in the microvasculature of the tumor. For how much reaches
the tumour's microvasculature may be indicative of how effective is the drug.
Without having to see a PD response from the tumor.
At least that is how I understood the origin of the thread. Maybe they
should provide the discussion group their reasons for seeking such data?
Cheers,
Alfredo R. Sancho,
USC PK-Imaging Ctr.
Back to the Top
Would the microdialysis suffer of "clogging", at least in the case of
tumours? That is, due to the rapid and chaotic growth of tumour cells. I
would hate to introduce the probe into the tumor, which already is leaky in
nature. But I would hate to implant the tissue or cell around the probe and
have it grow around it!
Any suggestions?
Back to the Top
On Thursday, 6 Mar 1997 11:06:29 -0600
"Richard Scheyer"wrote:
>Restoration of the integrity of tumor vascularity might be
>different, and might vary with tumor type. Possibly of greater
>importance is what you wish to measure. Microdialysis is a useful
>probe of the extracellular space. While this is a region of interest
>for many drugs, it may not be of particular importance for
>antineoplastic compounds.
The above statement is not correct. The extracellular space [interstitial
fluid space, IF + microvascular space] is definitively of
very major importance in drug transport into tumoral cells. We had recognized
this on mechanistic grounds when we were studying the mechanism of gallium
localization to tumors, where we had suggested a pH gradient in the
interstitial fluid space due to lactic acidosis. See for example:
The Mechanism of Tumor Localization of Gallium-67 Citrate: Role of
Transferrin Binding and Effect of Tumor pH. S.R. Vallabhajosula, J.F. Harwig
and W. Wolf. Intl. J. Nucl. Med., 8, 363-370, 1981.
The importance of the IF space became apparent again when we were trying to
model the tumoral pharmacokinetics of two anticancer drugs, 5-fluorouracil
(5FU) and cisplatin. We had to incorporate another compartment between the
vascular and the intracellular spaces to account for the trapping of 5-FU in
tumors. See:
Pharmacokinetic Imaging of 5-Fluorouracil in Tumors using 19F-NMR Spectroscopy.
Walter Wolf, Victor Waluch, Cary A. Presant, Hyun K. Kim and Alfredo R.
Sancho. Eurospin Annual 1995-1996, p. 99-103. and
Noninvasive 19F-NMRS of 5-Fluorouracil in Pharmacokinetics and Pharmacodynamic
Studies. Walter Wolf, Victor Waluch and Cary A. Presant. NMR in Biomedicine.
In Press.
Perhaps the major difference between studies in the CNS, as reported by
Dr. Scheyer, and in tumors, is reflected in what had been stated previously
in that message:
>After several hours, microdialysis concentrations reflect the extracellular
^^^^^
>space, rather than the vascular compartment, although there may be some
>residual "leakiness" for longer periods.
The time frame in tumors is much, much shorter. 5-FU, cisplatin, and presumably
many other antitumor drugs distribute by what is essentially a first passage
process. That is one of the major reasons we felt that the introduction of
another process whose kinetics were likely to be slow (equilibration between
the interstitial fluid space and that of the microdialysis chamber) would
not be conducive of measuring the fast kinetics of transfer between the 3
spaces that need to be considered when using PK to understand transport of
drugs into tumor cells: the vascular space, the interstitial fluid space
and the intracellular space.
This difference in the time frames (seconds or minutes, rather than hours)
is one of the main reasons why we believe that the use of noninvasive methods
is essential for capturing the kinetics of drugs in tumors. Another major
reason is that each tumor mass is different - in structure, in vascularization,
in the ratios of the various spaces - and therefore, in the transport of
drugs into tumor cells.
=========================================================================| Professor Walter Wolf, Ph.D. E-Mail: wwolfw.aaa.hsc.usc.edu |
| Director, Pharmacokinetic Imaging Program |
| Department of Pharmaceutical Sciences Telephone: 213-342-1405 |
| University of Southern California Fax: 213-342-9804 |
| 1985 Zonal Ave., Los Angeles, CA 90033 |
| |
| Center for Noninvasive Pharmacology, Los Angeles Oncologic Institute |
| MRI at St. Vincent Medical Center Telephone: 213-484-7235 |
| 2131 Third St., Los Angeles, CA 90057 Fax: 213-484-7447 |
========================================================================------------------------------
Date: Fri, 7 Mar 1997 10:03:48 -0600
From: Chandrani Gunaratna
Subject: Re: How are tumor drug concentrations measured?
At 11:39 AM 3/5/97 -0600, you wrote:
>PharmPK - Discussions about Pharmacokinetics
> Pharmacodynamics and related topics
>
>Dr. Gunaratna:
>
>We had thought of using microdialysis to measure antineoplastic drug
>concentration in and around tumors of various sizes that had been implanted
>in rats. We hesitated and did not use this method, instead we used true
>noninvasive imaging methods.
>
>My question is: How invasive is the probe? Does it perturb to a large
>extent the "normal" (as normal as a tumor can be) growth and patho-histology
>of the tumor? Does it cause extra leakiness?
>
>Thank you,
>
>Alfredo R. Sancho,
>USC-PK Imaging Ctr.
>
Microdialysis probe is totally invasive. They need to be implanted
surgically. The linear probes are very small in dimensions so they cause
minimal tissue damage. I do not recommend implanting them in humans at all
because they can release the tumor cells. Histological studies in liver
tissues done by Prof. Craig Lunte's group at University of Kansas have
shown that there is some necrosis at the implantation site in implantations
of longer than 12 hours.
I don't know what you mean by "leakiness"? Is it the ultrafiltration
property of the dialysis membrane or leaking of tumor components? I am not
very familiar with tumor physiology. Amount of ultrafiltration depends on
the membrane length, flow rate etc. Ultrafiltration is not so significant in
vivo.
You can get more information on this subject from Dr. Craig
Lunte(C-LUNTE.-a-.ukans.edu) or Dr. Malonne Davies
(mdavies.aaa.smissman.hbc.ukans.edu) at the University of Kansas.
Chandrani Gunaratna, Ph.D.
Senior Research Chemist
Bioanalytical Systems
2701 Kent Avenue
West Lafayette, IN 47906
Phone: (317)463-4527
E-Mail: prema.aaa.bioanalytical.com
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I'm in complete agreement that the interstitial space is vital in
modelling the kinetics. My point was only regarding the presumed
site of action. - Richard Scheyer
> Date: Fri, 07 Mar 1997 10:03:00 -0600
> From: Walter Wolf
> Subject: Re: How are tumor drug concentrations measured?
> To: Multiple recipients of PharmPK - Sent by
> Reply-to: PharmPK.-at-.pharm.cpb.uokhsc.edu
> PharmPK - Discussions about Pharmacokinetics
> Pharmacodynamics and related topics
>
> On Thursday, 6 Mar 1997 11:06:29 -0600
> "Richard Scheyer"wrote:
>
> >Restoration of the integrity of tumor vascularity might be
> >different, and might vary with tumor type. Possibly of greater
> >importance is what you wish to measure. Microdialysis is a useful
> >probe of the extracellular space. While this is a region of interest
> >for many drugs, it may not be of particular importance for
> >antineoplastic compounds.
>
> The above statement is not correct. The extracellular space [interstitial
> fluid space, IF + microvascular space] is definitively of
> very major importance in drug transport into tumoral cells. We had recognized
> this on mechanistic grounds when we were studying the mechanism of gallium
> localization to tumors, where we had suggested a pH gradient in the
> interstitial fluid space due to lactic acidosis.
Richard Scheyer, M.D.
Dept. Neurology, Yale School of Med.
P.O. Box 208018, New Haven, CT 06520-8018 USA
Richard.Scheyer.-a-.yale.edu
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Alfredo,
> 1) If clearance is a way of calculating the amount of drug left in the blood
> compartment, hence how much is left in the tumor... what model are you using
> to compare the tumor compartment?
I assume "compare the tumor compartment" means "predict the tumor
compartment amount (or conc)".
n> There is no tissue in the body that has
> the same infrastructure as that of tumours. We could not even compare it to
> the tissue in which the tumour is in. Therefore, either assumptions are made
> or invasive measurements are needed.
I agree with you that purely PK guided methods based on plasma concs
cannot predict tumour concs. Early attempts (1970s) to explain delayed drug
effects (digoxin and LSD were the classical examples) nevertheless had
some success in *describing* the time course of drug *effect* using 2
compartment PK models based only on plasma concs of drug.
In 1979 Lew Sheiner formulated what is known as the effect compartment
model which uses the time course of drug effect and assumes a
first-order process for equilibration of active drug substance at the
site of drug effect. Often a parametric model for the relationship
between conc and effect is also assumed but this is not required.
Another important class of models used to explain delayed drug effects
are based on a physiological mechanism which is modified by the drug.
Warfarin is the classic example here but these so called indirect effect
models have been used in lots of places where the drug action can be
understood in terms of an influence on the kinetics of a physiological
substance that mediates the observed drug response.
Some assmptions are involved but especially with the indirect effect
models these are usually strongly supported by the known physiology/
biochemistry.
I personally dont have an aversion to making assumptions like these.
They are not leap-of-faith as you have suggested in an earlier posting
(or at least they are dont seem so to me and you have not given me any
examples yet of such LOF assumptions).
> Now if we agree that there is great patient intervariability and that
> measurements of clearance can be used as an indicator for tumor trapping of
> the drug. How accurate is this?
I dont think accuracy is really the issue here. It depends on what you mean.
> Is is not contradictory?
I dont see any contradiction (yet).
> In the clinical
> scenario, would a one "common" or "average" dose -statistically calculated
> from previous patients- and noninvasive measurements be in order?
Sure. When planning the first dose for a patient you cannot do anything
else but rely on prior knowledge.
> That is
> the first does measured at the tumor site itself, might be a indicator -if
> not a predictor- of tumor response.
I quite agree. Once you have given a dose you can observe effects (and
concs) and start fine tuning how to achieve your therapeutic target (see
Holford NHG The target concentration approach to clinical drug
development. Clin Pharmacokin 1995; 29:287-291 for details of such an
algorithm).
Of course,just how good a predictor the observed concs or effects are for the
response to the next dose depends a lot on the drug. The etoposide
example I cited in my previous posting predicted that concs used
to individualise clearance would be of more value than attempting to
individualise EC50 based on measured effects.
> You see I put myself in the patients position... do I want to go through an
> entire protocol of chemotherapy without know if it is effective or not. Based
> solely on "statistical analysis" of previous patient tumor response data. Or
> do I want to take a single dose ("average" dose) and measure its prescence or
> lack of it in the tumor itself. This would tell the clinician that the drug
> is not effective for that tumor, in that tissue, with those
> histo-pathological characteristics. Which in turn would save money, time and
> pain, leading to possibly finding the drug that does have the desired
> therapeutic effect.
The example you give is rather simplistic. If there was such a drug and
such a non-invasive test that could, on the basis of the a measurement
made after the first dose, predict that the drug would not be effective then
I am sure it would be very helpful. But I do not know of any real
example of such a drug and test. Maybe a drug which exhibits multi-drug
resistance (eg via P-glycoprotein transporter action) could show such an
all or none penetration into a tumour.
As you belong to the USC imaging group do you have any published
examples of such phenomena?
It also seems unlikely that knowing the kinetics of the drug in the
tumour would be all you would need. The sensitivity (or resistance if
you want to think of it like that) of the tumour to the drug as a
function of the tumour conc would also need to be understood. So you
need a measure of drug effect as well as drug kinetics in tumour.
Now, if you have the time course of the drug effect on the tumour and
you have the time course of plasma concs a model connecting the two sets
of observations can be built that can identify and distinguish the
kinetic eg. CL, and dynamic eg. EC50, components of the response and
the next dose can be predicted on the basis of this improved
understanding of what determines the response in that patient.
>
> Now that this has been said, I understand the origin of this thread as
> someone coming from this point. That is, how can we measure the
> antineoplastic in the microvasculature of the tumor. For how much reaches
> the tumour's microvasculature may be indicative of how effective is the drug.
> Without having to see a PD response from the tumor.
Not having the tumour response only gives you part of the picture (see
above).
>
> At least that is how I understood the origin of the thread. Maybe they
> should provide the discussion group their reasons for seeking such data?
I agree that it would be useful if the person who started this thread
(from Parke-Davis I seem to recall) would join us again to offer some
justification for wanting to determine blood-free tumour drug concs.
If anyone else is doing this type of thing it would be nice to hear from
you too. My impression has been that we have had technical suggestions
for applying methodology but no examples showing that knowing tumour
concs makes any difference to drug development or patient care. I have
given an example which predicts direct patient care consequences from
understanding anti-tumour drug effects based on *blood* concs and *effects*.
So lets hear from you tissue drug measurement guys!
--
Nick Holford, Dept Pharmacology & Clinical Pharmacology
University of Auckland, Private Bag 92019, Auckland, New Zealand
email:n.holford.at.auckland.ac.nz tel:+64(9)373-7599x6730 fax:373-7556
http://www.phm.auckland.ac.nz/Staff/NHolford/nholford.html
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Nick:
At 10:47 PM 3/8/97 -0600, you wrote,
Re: value of measuring tumor drug concentrations.
>PharmPK - Discussions about Pharmacokinetics
> Pharmacodynamics and related topics
>If anyone else is doing this type of thing it would be nice to hear from
>you too. My impression has been that we have had technical suggestions
>for applying methodology but no examples showing that knowing tumour
>concs makes any difference to drug development or patient care. I have
>given an example which predicts direct patient care consequences from
>understanding anti-tumour drug effects based on *blood* concs and *effects*.
>So lets hear from you tissue drug measurement guys!
Although I am not a "tissue drug measurement guy", these
tissue-distribution studies are extremely valuable, in that they identify
non-linearities in drug uptake or drug efflux from normal tissues and
tumors (i.e. as may be possible with p-glycoprotein mediated efflux, as you
mention). A well-described example of non-linear tissue uptake is provided
by Dedrick et al., who constructed physiologic pk models of methotrexate
disposition.
I do not know of a single example of a PK/PD model utilizing tissue
concentrations to predict efficacy or toxicity; however, this is not
'proof' that such models would not be valuable (or should not be pursued)!
The development of rational therapeutic optimization strategies, based on
PK/PD, requires robust (if not realistic) models. Investigation of possible
areas of non-linear disposition is essential for the development of robust,
useful models.
Having said this, it is worthwhile to note that any approach investigating
tissue v. plasma concentration relationships should attempt to make said
determinations at a range of doses (or if possible, concentrations).
Evaluation of tumor distribution at a single, low dose (as suggested by
someone along this string) *may* provide an inaccurate assessment of tumor
distribution resulting from therapeutic doses.
------------
Joseph P. Balthasar, Ph.D. Phone: 801-585-5958
Assistant Professor FAX: 801-585-3614
Department of Pharmaceutics and email: jbalthasar.-at-.pharm.utah.edu
Pharmaceutical Chemistry
421 Wakara Way, Room 316
University of Utah
Salt Lake City, Utah 84108
------------
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why not float a cath into the tumar itself and biopsy from within?
On Fri, 28 Feb 1997, LATHIAC wrote:
> PharmPK - Discussions about Pharmacokinetics
> Pharmacodynamics and related topics
>
> We have been trying to measure drug concentrations in tumors. The problem we
> run into is the blood vessels surrounding the tumor. If we grind the tumor
> with the blood vessels, we will be measuring some composite value of the
> concentration of drug in tumor and in blood. We have tried to remove the
>blood
> vessels surrounding the tumor but have not been able to do it
>successfully. I
> was wondering if anyone in this group has experience in measuring the
> concentration of drug in tumors with a significant amount of vasculature. Is
> there a way to remove the vasculature or correct for the contribution of drug
> in blood to calculate the concentration in tumor? Any guidance you might
>have
> on this topic will be appreciated. Please include any references that
>discuss
> this issue.
>
> Thanks,
> Chetan D. Lathia
> Pharmacokinetics and Drug Metabolism
> Parke-Davis Pharmaceutical Research
> E-mail: Lathiac.-at-.AA.WL.COM
>
Back to the Top
> >So lets hear from you tissue drug measurement guys!
Thank you. I note however that you are an indirect tissue measurement
guy and not really a representative of the wet tissue group I was hoping
would contribute at this point. However, many of your remarks have
relevance to the study of PK on an organ or regional basis.
> Perhaps the most cogent
> reasons why we believe that relying on noninvasive methods for generating
> data are:
>
> A) The living system is, by its very nature, a dynamic system. Precise
> knowledge of the kinetic parameters is probably as important - and
> sometimes likely to be even more important - than knowledge of the
> thermodynamic values.
Thermodynamic values in this context means the same as pharmacodynamics
to me. My earlier example of etoposide PKPD (Karlsson et al).
illustrated exactly this point. PK (clearance) was a more useful
predictor of individual response than PD (EC50).
> Kinetic data, however, must be generated under
> conditions where the system to be studied is not perturbed - e.g., that these
> kinetic events are not modified during the process of attempting to measure
> them.
The latter constraint is only a problem if your kinetic model does
not include knowledge about "system perturbations". So called
physiological PK models attempt to do this (at least for blood flow and
organ size perturbations).
>
> B) Noninvasiveness does, by its very nature, not perturb the system from
> which one is trying to generate data.
I am not familiar with your technology but assuming it is along the
lines of PET scanning etc I would say that the while not very invasive
(in contrast to direct brain sampling) it is quite interventional
(scanner etc). Activities such as lying down e.g. to be scanned, can
change systemic PK.
> C) The collection of data from a single individual, and especially,
> measuring in that same individual changes induced by various
> perturbations, yields information that is often lost when analyzing pooled
> data. This is because such pooled data have all the interindividual
> variabilities superimposed on them. This topic was well documented in the
> PhD dissertation of R. Ricardo Brechner (1986), and published in J. Pharm.
> Sci. 53:873-877, 1986.
The disadvantages of naive pooled data approach have been discussed at
length in the PKPD literature and popln approaches to understand
responses in individuals are widely used e.g. see the book "Variability
in Drug Therapy" edited by Rowland, Sheiner and Steimer. Raven Press
1985 (based on a meeting in Rome in May 1984). At the second
COST B1 program meeting on "Measuring and managing variability in
response, concentration and dose" was held in Geneva, Feb 1997, and
included a review by Karlsson on advances in the anti-cancer area.
> D) Inasmuch as tumors, by their very nature, do not obey precise rules of
> growth, development, etc., it is critical to understand the degree of
>exposure
> of each tumor and or metastatic site to develop proper chemotherapeutic
> planning.
It may be that it is feasible on a tumour specific basis to plan
chemotherapy using tumour localised PK data. I acknowledge your
P<0.000001 statement below but this is not the same as prospectively
demonstrating the value of the intervention as the history of
therapeutic drug monitoring has shown.
> We now have a letter appearing in this month's Annals of Oncology that
> states that response of a tumor to 5-Fluorouracil requires that there are 3
> independent conditions that have to be met simultaneously for this drug to
> be active. These 3 conditions are:
> 1) There must be trapping of 5-FU in the tumor.
> 2) The tumor must have adequate perfusion to make such trapping
> possible.
> 3) The tumor type must be of a responsive type to make such trapping
> effective.
Fine. But note that condition 1 is rather general. Its not an all or
none world. The relation between extent of trapping and tumour response
needs to be established (e.g. in terms of Emax, EC50) and in particular
what is the variability in response given the same trapping from one
treatment occasion to the next. Unless this variability is sufficiently
small there will be no benefit from attempting to individualise doses
for a specific patient.
> > My impression has been that we have had technical suggestions for
> > applying methodology but no examples showing that knowing tumour
> > concs makes any difference to drug development or patient care.
> We do have examples that show that drug trapping in
> tumors makes a significant difference on patient care. We have shown [The
> Lancet, 343:1184-1187, 1994] that there is a very strong association
> between the trapping of 5-FU in a variety of human tumors and response
> [p<.000001]. Our clinicians have been making use of whether a patient
> traps 5-FU to make decisions on patient treatment, as well as to assess
> whether a given modulator (e.g., methotrexate, interferon, etc.) is worth
> using in a particular patient.
Apart from the statistical correlation what can you say about the
prospective benefit of making decisions about 5-FU? As noted above is
this reproducible for a specific patient in settings where repeat
treatments are usually employed? If single shot treatments are used then
of course it is hard to understand how any individualisation based on
studying the drug in that tumour can be of use (apart from the use of
tracer doses which may be misleading for non-linear disposition
processes).
>
> Our studies showing that the targeting of cisplatin [studied using 195mPt-
> cisplatin] to brain tumors is a function of both the method of drug
> administration [see Cancer Research 49:1877-1881, 1989] and modifiers of
> osmolality [Proc. Am. Assoc. Canc. Res. 36:360(2142), 1995] are being
> utilized by clinicians to achieve significant success in the treatment of
> patients with high grade astrocytoma.
Thank you for the info.
This thread has revealed that different groups within the
overal field of pharmacology are often unaware of the advances made by other
disciplines. I certainly admit to such ignorance and I am delighted to
have learned from the contributions to this thread.
--
Nick Holford, Dept Pharmacology & Clinical Pharmacology
University of Auckland, Private Bag 92019, Auckland, New Zealand
email:n.holford.at.auckland.ac.nz tel:+64(9)373-7599x6730 fax:373-7556
http://www.phm.auckland.ac.nz/Staff/NHolford/nholford.html
Back to the Top
> Although I am not a "tissue drug measurement guy", these
> tissue-distribution studies are extremely valuable, in that they identify
> non-linearities in drug uptake or drug efflux from normal tissues and
> tumors (i.e. as may be possible with p-glycoprotein mediated efflux, as you
> mention). A well-described example of non-linear tissue uptake is provided
> by Dedrick et al., who constructed physiologic pk models of methotrexate
> disposition.
This point is well taken. I wonder if this is the main reason why the
"tissue measurement guys" do it? What successes have they had?
>
> I do not know of a single example of a PK/PD model utilizing tissue
> concentrations to predict efficacy or toxicity; however, this is not
> 'proof' that such models would not be valuable (or should not be pursued)!
I accept the lack of proof concept. But on the other hand people have
been at this tissue PK game for decades (Dedrick was doing his stuff in
the '60s amd '70s). I am not aware that even the Dedrick MTX example had
a clinical spinoff but I would be happy to be corrected on that.
So just how has tissue sampling helped in understanding human
clinical pharmacology?
> Having said this, it is worthwhile to note that any approach investigating
> tissue v. plasma concentration relationships should attempt to make said
> determinations at a range of doses (or if possible, concentrations).
> Evaluation of tumor distribution at a single, low dose (as suggested by
> someone along this string) *may* provide an inaccurate assessment of tumor
> distribution resulting from therapeutic doses.
Agreed.
--
Nick Holford, Dept Pharmacology & Clinical Pharmacology
University of Auckland, Private Bag 92019, Auckland, New Zealand
email:n.holford.-at-.auckland.ac.nz tel:+64(9)373-7599x6730 fax:373-7556
http://www.phm.auckland.ac.nz/Staff/NHolford/nholford.html
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