Drug
Metabolism Letters
ISSN: 1872-3128
Drug Metabolism Letters
Volume 2, Number 1, January 2008
Contents
Visualization of First-Pass Drug Metabolism of
Terfenadine by MALDI-Imaging Mass Spectrometry Pp.
1-4
Jiwen Chen, Yunsheng Hsieh, Ian Knemeyer, Lee Crossman
and Walter A. Korfmacher
[Abstract]
Phenotyping of Cytochrome P450 3A Enzyme in Gujarat
Population Pp. 5-10
Ashutosh J. Jani, Shivprakash Rathnam and Anita A. Mehta
[Abstract]
Renal and Hepatic Transporter Expression in Type 2
Diabetic Rats Pp. 11-17
Michael T. Nowicki, Lauren M. Aleksunes, Sharmilee P.
Sawant, Ankur V. Dnyanmote, Harihara M. Mehendale and José
E. Manautou
[Abstract]
Allelic Variations in 5, 10-Methylenetetrahydrofolate
Reductase Gene and Susceptibility to Cervical Cancer in Indian
Women Pp. 18-22
Naveen Kumar Nandan, Saima Wajid, Shilpie Biswas, Sominder
Singh Juneja, Moshahid Rizvi, Raminder Prakash and Samar Husain
Naqvi
[Abstract]
Pharmacokinetics of Ceftriaxone in Carbontetrachloride
Induced Hepatopathic and Uranyl nitrate-Induced Nephropathic
Goats Following Single Dose Intravenous Administration
Pp. 23-28
Tapas Kumar Sar, Tapan Kumar Mandal, Shymal Kumar Das
and Animesh Kumar Chakraborty
[Abstract]
Modulation of Porcine (Sus scrofa domestica)
and Pheasant (Phasianus colchicus) Carbonyl Reducing
Enzymes by Anthelmintic Therapy with Flubendazole
Pp. 29-34
Barbora Szotáková, Milan Nobilis, Jirí
Lamka, Veronika Krížová, Michal Šavlík
and Lenka Skálová
[Abstract]
Identification of a Novel Glutathione Conjugate of
Diclofenac by LTQ-Orbitrap Pp. 35-40
Yohannes Teffera, Daniel J. Waldon, Adria E. Colletti,
Brian K. Albrecht and Zhiyang Zhao
[Abstract]
Rapid and Sensitive Characterization of the Metabolite
Formation Enzyme Kinetics of Radiolabeled Drugs Using Stop
Flow Liquid Radiochromatography Pp. 41-46
Weiping Zhao, Lifei Wang, Donglu Zhang and Mingshe Zhu
[Abstract]
Time and Dose Dependent Study of Doxorubicin Induced
DU 145 Cytotoxicity Pp. 47-50
Saeed S. Al-Ghamdi
[Abstract]
An Examination of IC50 and IC50-Shift Experiments
in Assessing Time-Dependent Inhibition of CYP3A4, CYP2D6 and
CYP2C9 in Human Liver Microsomes Pp. 51-59
Loren M. Berry and Zhiyang Zhao
[Abstract]
Induction of Cytochrome P450 3A by the Ginkgo
biloba Extract and Bilobalides in Human and Rat Primary
Hepatocytes Pp. 60-66
Ying Deng, Hui-chang Bi, Li-zi Zhao, Xue-ding Wang, Jie
Chen, Zhi-min Ou, Liang Ding, Le-jia Xu, Su Guan, Xiao Chen,
Shu Feng Zhou and Min Huang
[Abstract]
Investigations into the Drug–Drug Interaction
Potential of Tapentadol in Human Liver Microsomes and Fresh
Human Hepatocytes Pp. 67-75
Christa Kneip, Rolf Terlinden, Horst Beier and Genfu Chen
[Abstract]
Abstracts
[Back to top]
Visualization of First-Pass Drug Metabolism of
Terfenadine by MALDI-Imaging Mass Spectrometry
Jiwen Chen, Yunsheng Hsieh, Ian Knemeyer, Lee Crossman
and Walter A. Korfmacher
The aim of this article is to focus on the implementation
and the application of matrix-assisted laser desorption/ionization-imaging
mass spectrometric system (MALDI-IMS) to determine the disposition
or biotransformation pathway of terfenadine and its active
metabolite, fexofenadine in mouse and rat whole-body tissue
sections. Whole-body MALDI-IMS data showed that the poor oral
bioavailability of terfenadine was largely due to high first-pass
metabolism in the intestines and the liver before the compound
reached systemic circulation.
[Back to top]
Phenotyping of Cytochrome P450 3A Enzyme in Gujarat
Population
Ashutosh J. Jani, Shivprakash Rathnam and Anita A. Mehta
CYP3A isoforms account for about 30% of total hepatic P450s.
Most statins have been accepted probes for CYP3A4. The metabolic
ratio of atorvastatin/ortho-hydroxyatorvastatin showed a bimodal
distribution with respect to metabolism of atorvastatin. These
observations showed that the frequency of occurrence of the
poor metabolizer phenotype is 2.4 % in the Gujarat subjects.
[Back to top]
Renal and Hepatic Transporter Expression in Type 2
Diabetic Rats
Michael T. Nowicki, Lauren M. Aleksunes, Sharmilee P.
Sawant, Ankur V. Dnyanmote, Harihara M. Mehendale and José
E. Manautou
Membrane transporters are critical for the uptake as well
as elimination of chemicals and by-products of metabolism
from the liver and kidneys. Since these proteins are important
determinants of chemical disposition, changes in their expression
in different disease states can modulate drug pharmacokinetics.
The present study investigated alterations in the renal and
hepatic expression of organic anion and cation transporters
(Oats/Octs), multidrug resistance-associated proteins (Mrps),
breast cancer resistance protein (Bcrp), P-glycoprotein (Pgp),
and hepatic Na+-taurocholate
cotransporting polypeptide (Ntcp) in type 2 diabetic rats.
For this purpose, type 2 diabetes was induced by feeding male
Sprague-Dawley rats a high fat diet followed by a single dose
of streptozotocin (45 mg/kg, i.p., in 0.01 M citrate buffer
pH 4.3) on day 14. Controls received normal diet and vehicle.
Kidney and liver samples were collected on day 24 for generation
of crude plasma membrane fractions and Western blot analysis
of Oat, Oct, Mrp, Bcrp, Pgp, and Ntcp proteins. With regards
to renal uptake transporters, type 2 diabetes increased levels
of Oat2 (2.3-fold) and decreased levels of Oct2 to 50% of
control kidneys. Conversely, efflux transporters Mrp2, Mrp4,
and Bcrp were increased 5.4-fold, 2-fold, and 1.6-fold, respectively
in type 2 diabetic kidneys with no change in levels of Mrp1,
Mrp5, or Pgp. Studies of hepatic transporters in type 2 diabetic
rats reveal that the protein level of Mrp5 was reduced to
4% of control livers with no change in levels of Bcrp, Mrp1,
Mrp2, Mrp4, Ntcp, or Pgp. The changes reported in this study
may have implications in type 2 diabetic patients.
[Back to top]
Allelic Variations in 5, 10-Methylenetetrahydrofolate
Reductase Gene and Susceptibility to Cervical Cancer in Indian
Women
Naveen Kumar Nandan, Saima Wajid, Shilpie Biswas, Sominder
Singh Juneja, Moshahid Rizvi, Raminder Prakash and Samar Husain
Naqvi
Methylenetetrahydrofolate reductase (MTHFR)
gene located on chromosome 1p36.3 catalyses the conversion
of 5,10-methylenetetrahydrofolate to 5,methyltetrahydrofolate,
the major methyl donor for the conversion of homocysteine
to methionine. Two common polymorphisms in the MTHFR
gene have been identified, 677C>T in exon 4, leading to
substitution of alanine by valine and 1298A>C in exon 7
which leads to the replacement of glutamic acid by alanine
re-sulting into reduced enzyme activity. The potential influence
of MTHFR activity on DNA methylation and on the availib-lity
of uridylates and thymidylates for DNA synthesis and repair
makes MTHFR an attractive candidate for cancer predis-posing
gene. In order to elucidate the role of MTHFR polymorphism
in cervical cancer, both the exons for 677C>T and 1298A>C
mutations were analyzed among 219 females, including 77 females
with normal cervical cytology, 80 with cer-vical dysplasia
and 62 with squamous cell carcinoma of uterine cervix. Females
with mutant allele at 677 position (CT/TT genotypes) were
found to be almost three times the risk of cervical dysplasia
than females with CC genotype [OR, 2.9; (CI, 1.5-5.7)], but
were less likely to develop squamous cell carcinoma [OR, 1.5
(CI, 0.7-3.2)]. Similar findings were ob-served for mutation
at 1298 position, females with AC/CC genotypes were almost
four times the risk of cervical dysplasia [OR, 4.3 (CI, 2.1-9.0)],
as compared to AA genotype. Our study lends further support
to the hypothesis that the MTHFR polymorphism (677C>T
or 1298A>C) is involved in susceptibility to cervical dysplasia.
[Back to top]
Pharmacokinetics of Ceftriaxone in Carbontetrachloride
Induced Hepatopathic and Uranyl nitrate-Induced Nephropathic
Goats Following Single Dose Intravenous Administration
Tapas Kumar Sar, Tapan Kumar Mandal, Shymal Kumar Das
and Animesh Kumar Chakraborty
The pharmacokinetic profile of ceftriaxone was studied in
female healthy goats, induced hepatopathic and nephropathic
goats after a single intravenous dose at 50 mg kg-1.
Ceftriaxone persisted for 2 h in plasma of hepatopathic goats
compared to 1 h of healthy goats, but the kinetic behaviour
followed ‘one-compartment open model’ in both
healthy and hepatopathic goats. Mean value of t½β
(0.32 ± 0.008 h) was significantly higher in hepatopathic
goats compared to healthy goats (0.19 ± 0.002 h). Ceftriaxone
was recovered at 24 h in urine of hepatopathic goats but it
could not be detected in urine of healthy goats. However,
its metabolite ceftizoxime was present in urine of healthy
goats but not in urine of hepatopathic goats. On the other
hand ceftriaxone persisted for 2 h in plasma of kidney damaged
goats with significant higher concentration compared to healthy
goats but kinetic behaviour followed ‘one Compartment
open model’. Ceftizoxime was identified with an adequate
plasma concentration from 8 h to 12 h post dosing in nephropathic
goats. Elimination halflife (t½β)
of Elimination ceftriaxone (0.38 ± 0.01 h) in nephropathic
goats increased significantly compared to healthy goats (0.19
± 0.002 h). Ceftriaxone, not the metabolite ceftizoxime
was recovered at 24 h and 48 h post dosing in urine of nephropathic
goats, while only ceftizoxime not ceftriaxone was detected
in urine of healthy goats.
[Back to top]
Modulation of Porcine (Sus scrofa domestica)
and Pheasant (Phasianus colchicus) Carbonyl Reducing
Enzymes by Anthelmintic Therapy with Flubendazole
Barbora Szotáková, Milan Nobilis, Jirí
Lamka, Veronika Krížová, Michal Šavlík
and Lenka Skálová
Flubendazole (FLU) is a widely administered benzimidazole
anthelmintic indicated for the control of parasitic diseases
in farm animals including pigs and pheasants. This study was
designed to test the biotransformation of FLU in control animals
and animals treated with FLU in recommended therapeutic doses.
The activities of several pheasant and porcine hepatic and
intestinal carbonyl reducing enzymes and their modulation
by FLU were also studied. Twelve adult pheasant hens, approximately
1 year old, were divided into two groups and treated for 7
days with placebo or 6 mg of FLU/kg of body weight. Eight
male hog weaners, approximately 3 month old, were divided
into two groups and treated for 5 days with placebo or 1.57
mg of FLU/kg of body weight. Subcellular fractions, prepared
from livers and small intestines of control and FLU treated
animals, were incubated with FLU. In vitro formation
of two main FLU metabolites, reduced FLU, and hydrolyzed FLU
were analyzed using HPLC. While FLU was reduced significantly
more intensively in FLU-treated pheasants than in control
animals, no differences were observed in pigs. These results
were confirmed by measuring the enzyme activities: carbonyl
reducing enzyme activities were increased in pheasants treated
by FLU, whereas FLU did not affect these enzymes in pigs.
[Back to top]
Identification of a Novel Glutathione Conjugate of
Diclofenac by LTQ-Orbitrap
Yohannes Teffera, Daniel J. Waldon, Adria E. Colletti,
Brian K. Albrecht and Zhiyang Zhao
High resolution accurate MS with an LTQ-Orbitrap identified
two novel metabolites of diclofenac in rat bile and rat and
human hepatocyte incubations: a benzyl-S-glutathione conjugate
and 2-(2,6-dichlorophenylamino) benzoic acid. A mechanism
for the bioactivation of diclofenac involving decarboxylation
is proposed.
[Back to top]
Rapid and Sensitive Characterization of the Metabolite
Formation Enzyme Kinetics of Radiolabeled Drugs Using Stop
Flow Liquid Radiochromatography
Weiping Zhao, Lifei Wang, Donglu Zhang and Mingshe Zhu
This study evaluated the reproducibility, sensitivity, accuracy,
and precision of stop-flow liquid radiochromatographic detection
(RFD). Stop-flow RFD was about 10-fold more sensitive compared
with traditional RFD and had a good reproducibility for quantification
and HPLC retention time. Stop-flow RFD was applied to determine
enzyme kinetics of radiolabeled drugs. The enzyme kinetic
parameters of muraglitazar glucuronidation determined by stop-flow
RFD were comparable with those determined by microplate scintillation
counting (MSC).
[Back to top]
Time and Dose Dependent Study of Doxorubicin Induced
DU 145 Cytotoxicity
Saeed S. Al-Ghamdi
Doxorubicin is one of the most effective anti-tumour agent.
To clarify whether a single dose produces its effects soon
after dosing or only after hours or days and whether the effects
are brief or prolonged, time-dose relationships were explored
by calculating lethal time (LT50)
values which is a statistical estimate of the time from dosage
to death of 50% of the organisms/ or cells in a very large
population subjected to a toxicant under specific conditions.
This was achieved by the log-time log-dose curve which may
be used to predict the proper doses to be used in long-term
studies. Drug chemosensitivity using DU-145 cell lines have
been investigated for this purpose. The results shows that
the effects of doxorubicin started only after 24hrs; however,
resistance was developed, 40 hrs was the time required to
kill 50% of cells , and post-incubation with fresh media (F.M.)
exhibited more cell damages. It is concluded that doxorubicin
is effective only after 24 hrs with resistance developed and
post-incubation with F.M after treating cells with doxorubicin
causes more damage than continuous incubation with the drug.
[Back to top]
An Examination of IC50 and IC50-Shift Experiments
in Assessing Time-Dependent Inhibition of CYP3A4, CYP2D6 and
CYP2C9 in Human Liver Microsomes
Loren M. Berry and Zhiyang Zhao
The relationship between time-dependent inactivation (TDI)
and IC50 is examined using a consolidated method for evaluating
CYP450 inhibition during drug discovery. An IC50
fold-shift of >1.5 indicated significant TDI potency. Further,
the “shifted IC50” could be used to estimate,
the Kl
and TDI potency ratio kinact/Kl
to within 2-fold in most cases.
[Back to top]
Induction of Cytochrome P450 3A by the Ginkgo
biloba Extract and Bilobalides in Human and Rat Primary
Hepatocytes
Ying Deng, Hui-chang Bi, Li-zi Zhao, Xue-ding Wang, Jie
Chen, Zhi-min Ou, Liang Ding, Le-jia Xu, Su Guan, Xiao Chen,
Shu Feng Zhou and Min Huang
Ginkgo biloba is one of the most popular herbal medicines
in the world, due to its purported pharmacological effects,
including memory-enhancing, cognition-improving, and antiplatelet
effects. The study aimed to investigate the activity and expression
of cytochrome P450 (CYP) 3A in human and rat primary hepatocytes
treated with standardized G. biloba extract (100,
500, and 2500 ng/ml) for 72 hr, and to measure the protein
expression of CYP3A in human and rat primary hepatocytes treated
with bilobalide (2, 10, and 50 ng/ml) and ginkgolides B (2,
10, and 50 ng/ml). The activity of CYP3A was measured by the
quantification of dehydronifedipine formation using a validated
tandem liquid chromatography mass spectrometry (LC/MS/MS)
method. The levels of mRNA and protein of CYP3A were determined
by reverse transcription-polymerase chain reaction (RT-PCR)
and Western-blotting analysis, respectively. The G. biloba
extract at 100-2,500 ng/ml significantly induced the activity,
protein and mRNA expression of CYP3A in a dose-dependent manner
in human and rat primary hepatocytes. Bilobalide at 2-50 ng/ml
significantly increased CYP3A protein expression in a dose-dependent
manner in human and rat primary hepatocytes. However, ginkgolide
B did not affect CYP3A protein expression in vitro.
The results indicate that G. biloba extract pretreatment
significantly induced the expression of CYP3A protein and
mRNA and increased CYP3A activity, and there was no significant
species difference between human and rat. G. biloba may cause
potential interactions with substrate drugs of CYP3A. Bilobalide
might play a key role in the enzyme-inducing effects of G.
biloba extract. Further study is needed to identify the substances
in GBE that induce CYPs in vivo, and elucidate the
molecular mechanism of CYP3A induction by GBE and bilobalides.
[Back to top]
Investigations into the Drug–Drug Interaction
Potential of Tapentadol in Human Liver Microsomes and Fresh
Human Hepatocytes
Christa Kneip, Rolf Terlinden, Horst Beier and Genfu Chen
The new analgesic tapentadol was evaluated for induction and
inhibition of several cytochrome P450 enzymes in vitro, and
protein binding was assessed. It was concluded that no clinically
relevant drug–drug interactions are likely to occur
through either mechanism.
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