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Hi Dear All:
In my previeous monkey toxicokinetic study, the exposure of a test
article (a small molecular) was decreased after 30 days repeated oral
dose in low dose level(10mg/kg/day), the AUC values on Day 30 were
nearly half of that on Day 1, but in middle(30mg/kg/day) and high
level(90mg/kg/day). Did this reasonable? I'm sure the formulation
preparation, dose and sample analysis were all correct. No special
emzyme induce or inhibition were found in in vitro studies.
Did anybody have the similar experience, or can you please give me
some suggestion or explanation?
Regards
Ning
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Dear Ning,
Enzyme inhibition is often a direct effect, however it may sometimes
take much longer for induction (days to weeks) to occur and not all in-
vitro assays are suitable to pick up slow induction. For example:
incubation of hepatocytes in suspension should not be longer than 4
hours. If you know which what metabolites are formed by which enzymes,
quantification of these metabolites in collected plasma, faeces and/or
urine could give you an indication of what's happening.
Sincerely,
Rob ter Heine
--
Rob ter Heine, MSc, PharmD
Department of Pharmacology, Slotervaart Hospital
Amsterdam, The Netherlands
E: rob.terheine.aaa.slz.nl
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Dear leining: 1) You may be missing some punctuation which makes
assistance
difficult.
You say... the AUC values on Day 30 were
nearly half of that on Day 1, but in middle (30mg/kg/day) and high
level (90mg/kg/day)... In middle and high dose what did you see?
Were the
decreases greater or lesser than the low dose? Were all the doses
consistent- same time of day, pre or post prandial?
You also say that you are sure the formulation, dosing and sample
analysis
were correct. But do you have analyses to support this? Did you test
the
formulation on each day of use/preparation? Do you have stability on the
material in formulation and in the biological sample? Did a second
person
observe the formulation, dosing and sample analyses? Was the analytical
method validated or were controls run along with the samples? Did the
controls perform within acceptance?
If you rule out misformulation, stability and analytical issues, you
may be
observing the induction of enzymes or transporters which accelerate the
clearance.
It is also not impossible for a small molecule to induce antibodies,
particularly if glass syringes or vessels were involved, if the
vehicle or
carrier or both were irritants. It is more likely with Sc injections
but
oral is not impossible.
What was the vehicle?
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Ning,
Please provide more details on the issues. As Ed noted, you will need
to provide as much clarity for these to be addressed properly (of
course without divulging any proprietary information that could get
you into trouble).
What was the formulation that you were using? Was it shown to have
full stability over the course of the study.
Were the formulations form a single batch used daily or reconstituted
fresh each day? Did the dose anlysis show stability and consistency
for concentraton?
Indicated solubility within the formulations matrix, was the compound
a solution of a suspension?
What is the therapeutic indication (if you can answer this) for the
compound and could it be altering the local gut environment such that
prolonged exposure reduced the fraction absorbed?
What induction assays were used to rule out enzyme induction
phenomena? Were they in vitro, in vivo or post mortem liver evaluation
for enzyme content and activity?
Do you have any supprting hepatocyte induction data for looking at
inducible elements in monkey heptocytes (or any otherspecies for that
matter)/
It is an interesting issue that has mnay components that some of us
have seen before on this board, so you're in luck - in a sense.
Sanjeev Thohan, PhD
SARXconsulting
SARXconsult.at.Gmal.com
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Dear Ed,
you wrote:
"It is also not impossible for a small molecule to induce antibodies,
particularly if glass syringes or vessels were involved...".
Could you please elaborate on this intruiging comment?
Best regards,
Frederik Pruijn
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Thanks for all guys answered my previous question, here I'd like to
give some details.
1, the Test article was suspended in 0.5%CMC-Na, the stability were
over 2 weeks at RT, we prepared it each week, and analysis (HPLC)each
batch to make sure the Homogeneity and concentration were within the
acceptance(bias 90%-110%).
2, I reviewed all the raw data and preparation procedure, and I'm sure
that everything is fully followed the protocol and SOP, so this lower
phenomenon was not due to anything wrong occurred during the
formulation, dosing, sample collection.
3, The method was validated before, and within all analysis runs,
Standard curves and QCs were passed. So, the bioanalysis is OK.
4, the in vitro studies were not performed in our facility, but the
people involved these told me no enzyme inhibition and induce were
found.
5, In middle and high dose levels, the AUC values increased less
proportionally, no gender difference was observed.
6, In rats TK study, in all dose levels, the AUC increased less
proportionally.
6, I'm afraid the points should be found in gastrointestinal
absorption, maybe the repeated dose changed some factors such as
bacterial growth, enzyme activities and physicochemical environment,
etc,.
Any further suggestion? Yes, this is a interesting issue, maybe I'm
lucky. ^_^
Regards
Ning
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Did you examine stomach contents?
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Ning,
A couple of comments:
1. I won't rule out variability as the simple cause for the 50% drop
in low dose AUC, especially the low dose may be close to the
inflection point of the sigmoidal exposure-dose curve, since you have
suggested there may be a saturation process during the absorption.
I'd check the historical data from all SD/MD PK/TK studies to see what
the typical inter-animal CV values were for all comparable doses,
including other species.
2. Were liver tissues collected from the animals per study protocol?
It's a 28-day study so I assume all animals were to be sac'ed. Take a
look at the CYP level in the liver for treatment groups vs. control
and you may rule out induction. This is more reliable than in vitro
assay as some have already pointed out.
Also, is it possible the compound can induce Pgp but not CYP? At
higher doses, saturation may overcome increased drug efflux.
Jack
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Hi Ning,
I have seen this before but, unlike your case, was dose related and
rather explainable with enzyme induction in vitro.
How does each individual monkey data on Day 1 and Day 30 match/compare
in low dose group?
What was the %CV? If you remove potential outlier(s) still see 2-fold
reduction in AUC?
Was any emesis (vomiting) reported for low dose group on Day 30?
Any of your tox findings e.g., clinical observation shows a trend
(e.g., improvement/reduction in frequency or severity) that is
consistent with reduce exposure over time?
Rostam
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Hi dear all;
The %CV of AUC was 57% for male and 25% for female, however, It seemed
that the Cmax of Female animals decreased more, compared with that of
male. Anyway, no vomiting was observed in low level group and no
obvious change was found by histological pathologist.
That's really confused me.
Regards
Ning
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Hi Ning,
You have correctly mentioned that the decreased in AUC could be due to
decrease in GIT absorption,provided that there is no role of metabolic
enzyme induction.
Physicochemical property of the drug itself as well as its effect on GIT
motility can also lead to decrease absorption.
Repeated dosing with the drug sometime tends to damage the intestinal
lining leading to decrease absorption.
For this, you can check for the food consumption and weights of the rat.
Regards
Anasuya
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Hi,
Another, simpler thought is that the rats might have taken in the
compound over a longer period of time, so decreasing Cmax and
increasing the first pass effect and so reducing AUC. If the compound
was bitter for example, that might have made the animals learn to take
in the food more gradually.
Andrew Sutton
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You then did not find concretions or bezoars? Were the fecal pellets
the
same number, size, weight and appearance between groups? Was the fluid
balance the same between groups?
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