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Reproductive Toxicology xxx (2015) xxxxxx

Contents lists available at ScienceDirect

Reproductive Toxicology
journal homepage: www.elsevier.com/locate/reprotox

Evaluation of early fetal exposure to vaginally-administered

metronidazole in pregnant cynomolgus monkeys
Kary E. Thompson a, , Deanna L. Newcomb b , Graeme J. Moffat c , Julie Zalikowski c,1 ,
Gary J. Chellman b , Mary Ellen McNerney a
a
Reproductive Toxicology, Drug Safety Evaluation, Research & Development, Bristol-Myers Squibb Company, New Brunswick, NJ 08903, United States
b
Preclinical Services, Charles River Laboratories, Reno, NV 89511, United States
c
Amgen Inc., Thousand Oaks, CA 91320, United States

a r t i c l e i n f o a b s t r a c t

Article history: Given concern about potential embryo-fetal harm following seminal exposure to drugs with teratogenic
Received 11 August 2015 potential, pharmaceutical companies use theoretical calculations to estimate seminal concentrations,
Received in revised form maternal exposure, and distribution across the placenta to the embryo-fetal compartment for risk assess-
17 September 2015
ment. However, it is plausible that there are additional mechanisms whereby the conceptus is exposed. In
Accepted 24 October 2015
order to determine if theoretical calculations are sufciently conservative to predict embryo-fetal expo-
Available online xxx
sure from drugs in semen, pregnant cynomolgus monkeys were given a vaginal dose of metronidazole
during the early fetal period and cesarean-sectioned. Maternal, fetal, and amniotic uid samples were
Keywords:
Semen
analyzed for metronidazole and 2-hydroxymetronidazole. Exposure to metronidazole and its metabolite
Small molecule drug were comparable in all matrices. These data demonstrated no preferential transfer mechanism to concep-
Metronidazole tus following intravaginal administration of a small molecule drug; and therefore, suggest that traditional
Risk assessment modeling for embryo-fetal exposure to drugs in semen in support of risk assessment for pharmaceutical
Intravaginal administration agents is sufciently conservative.
Fetus 2015 Elsevier Inc. All rights reserved.

1. Introduction
In traditional pharmaceutical development, ICH Harmonized
Tripartite Guideline embryo-fetal development studies in animals
are conducted to support the entry of women of childbearing poten-
tial into clinical trials, and provide guidance for the women taking
pharmaceutical products during pregnancy (both intentionally or
non-intentionally, in the event that the pregnancy was not planned
or recognized) [1,2]. Based on the developmental toxicity prole
generated by nonclinical evaluation of treated maternal animals
and their conceptuses, pregnancy labels generally focus on poten-
tial implications to the embryo-fetus related to use of drugs in
women.
However, exposures to the conceptus are possible in preg-
nant partners of men who are treated with drugs. Small molecule
medicines are known to distribute to semen, and in some limited
cases accumulate in this compartment. As reviewed by Pichini et al.
Corresponding author at: Bristol-Myers Squibb Company, 1 Squibb Drive, New
Brunswick, NJ 08903, United States. Fax: +1 732 227 3218.
E-mail address: kary.thompson@bms.com (K.E. Thompson).
1
Present address: Accium Biosciences, Seattle, WA 98122, United States.
[3], Kashuba et al. [4], and Klemmt and Scialli [5], the semen-to-
blood ratio of over 50 drugs was typically less than or equivalent
to 1, i.e., concentrations in seminal uid did not exceed those in
the systemic circulation. However, there were instances whereby
higher ratios were identied (typically associated with antimicro-
bials), with accumulation in semen up to 11.3 that in circulation
[6]. As such, there is potential concern that pregnant partners of
men treated with an agent with teratogenic liabilities identied
at exposures less than or equivalent to clinical levels, or associated
with mechanism of actions related to development, could adversely
affect the developing conceptus, due to exposure to drug in semen.
To investigate the likely manifestation of these theoretical con-
cerns for paternally-mediated developmental toxicity, typically
companies conduct a theoretical risk assessment using a series of
assumptions to predict a worst case exposure to a developing con-
ceptus and make risk assessments. In this approach, using the upper
range of typical human semen volume, an accumulation of drug in
semen of 10-fold, complete absorption from the vaginal compart-
ment into maternal circulation, and then nally distribution of the
drug throughout the maternal depots, including the conceptus, a
predicted maternal systemic exposure is estimated. However, due
to the relatively small seminal volume and dilution of the seminal
dose in maternal circulation, the likely maternal exposure to drugs

http://dx.doi.org/10.1016/j.reprotox.2015.10.013
0890-6238/ 2015 Elsevier Inc. All rights reserved.

Please cite this article in press as: K.E. Thompson, et al., Evaluation of early fetal exposure to vaginally-administered metronidazole in
pregnant cynomolgus monkeys, Reprod Toxicol (2015), http://dx.doi.org/10.1016/j.reprotox.2015.10.013
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in semen is several orders of magnitude less than those in male
patients directly taking the drug, limiting the concern for adverse
paternally-mediated effects to only the most potent of teratogens
[5].
While these conservative assumptions indicate the risk is low,
it is recognized there are data gaps for potentially unmodeled
exposure to the conceptus because typically, nonclinical animal
models are not receptive to mating following onset of pregnancy. As
described by Klemmt and Scialli [5], there are hypothetical mecha-
nisms for potential local delivery of drugs in semen to the conceptus
including: (1) countercurrent transfer from vaginal veins to the
uterine arteries, (2) trans-cervical transport, and (3) adsorption of
the drug to sperm (the last relevant to the time of fertilization only).
In order to derive animal data to evaluate the potential for delivery
of drugs in semen to a conceptus at levels beyond those modeled
by theoretical assumptions, this study was designed and conducted
to evaluate the exposures to monkey fetuses in early gestation fol-
lowing maternal vaginal administration of the antimicrobial agent,
metronidazole. Additionally, this study was designed to contribute
to the body of science as part of the Health and Environmental Sci-
ences Institute (HESI) Developmental and Reproductive Toxicology
(DART) Drugs in Semen Consortium [7].
2. Materials and methods
2.1. Animals and animal care
Care and use of the animals was in conformity with the American
Association for Laboratory Animal Science Policy on the Humane
Care and Use of Laboratory Animals. All procedures performed
on the animals were reviewed and approved by the Institu-
tional Animal Care and Use Committee (IACUC) at Charles River
Laboratories. Sexually mature and experimentally-nave Chinese
cynomolgus monkeys (Macaca fascicularis) were used for this study.
Females were naturally-mated for 3 days with a breeder male based
on anticipated timing of ovulation; pregnancies were conrmed
by ultrasound periodically, and on the morning of experimen-
tal treatment. Animals were individually housed in stainless-steel
cages with environmental enrichment and grooming bar access to
adjacent cages. Rooms were environmentally controlled with an
approximate 12-h light/dark cycle and maintained at 64 to 84 F
and 3070% humidity. Monkeys were provided a rationed amount
of Purina certied primate diet (No. 5048) daily, with supplemen-
tal fruits and vegetables provided 23 times weekly, and municipal
water, processed by reverse-osmosis ltration and UV-light treat-
ment, ad libitum.
2.2. Experimental design
Three pregnant female monkeys were used in the study (5.15.3
years old), all receiving treatment with a single intravaginal dose
(1 mL) of 0.75% metronidazole gel (Sandoz Inc., Princeton, NJ,
molecular weight = 171) during the early fetal period [Gestation
Day (GD) 60, 70, or 71 for Female Nos. 1502, 1501, and 1503, respec-
tively]. The pharmacokinetics of intravaginal metronidazole gel in
humans are well characterized [8]; compared to oral administra-
tion, absorption of metronidazole from intravaginal gel is more
prolonged, incomplete, and variable with peak systemic exposures
observed 6 to 12 h after vaginal dosing. Since the drug is already dis-
solved in gel, this formulation is easily absorbed as the gel improves
contact with the vaginal surfaces relative to tablet or cream appli-
cations. Given all of these factors, metronidazole gel was selected
for use in this study. Doses were given with a 1 mL syringe, inserted
approximately 0.250.5 inch into the vagina of chair-restrained
pregnant females. The gel was dyed blue with food grade blue
Please cite this article in press as: K.E. Thompson, et al., Evaluation of early fetal exposure to vaginally-administered metronidazole in
pregnant cynomolgus monkeys, Reprod Toxicol (2015), http://dx.doi.org/10.1016/j.reprotox.2015.10.013
dye immediately prior to use to ensure the gel could be visualized
for potential leakage. Females were continuously monitored for an
hour after dosing to check for expulsion of gel and provided edible
enrichment during this time.
2.3. Cesarean-sections and sample collection
Approximately 7 h after dosing (within the range of expected
Cmax ), each monkey was anesthetized and a cesarean section was
conducted. Immediately prior to surgery, a 1-mL blood sample was
collected from the femoral artery of each female. At C-section, a
3-mL sample of amniotic uid was collected and retained. Follow-
ing removal of the fetus from the gestational sac, the umbilical
cord was clamped and cut 23 cm from the umbilicus; umbilical
blood was collected. Fetuses were anesthetized with phenytoin
and pentobarbital (Beuthanasia -D) via intraperitoneal injection.
Fetal blood collection was attempted by cardiac puncture, decapi-
tation, and/or sampling of the vena cava. Fetal body weights were
measured. All maternal and fetal blood samples were collected into
tubes with anticoagulant (K2 EDTA) and then processed to plasma.
Samples were stored at 80 C until analyzed. Female monkeys
were returned to the facility stock colony following recovery from
the surgical procedure.
2.4. Sample concentration bioanalysis
Monkey plasma and amniotic uid samples were prepared by
liquidliquid extraction in ethyl acetate. Analytes were injected
into a Phenomenex MAX RP analytical column and analyzed
by Turboionspray Ionization (TIS) tandem mass spectrometry
(MS/MS) in the positive ion multiple reaction monitoring mode.
The concentrations of metronidazole and the major metabolite,
2-hydroxymetronidazole, in plasma and amniotic uid quality con-
trols and unknown samples were calculated by weighted linear
regression from plasma calibration standards run simultaneously
with the quality controls and unknown samples. Data was analyzed
with Sciex program Analyst, Version 1.4.1. Overall precision and
accuracy for the calibration standards and quality control samples
met the established acceptance criteria.
3. Results
Single, xed volume, vaginal doses of 7.5 mg metronidazole
were well tolerated by pregnant cynomolgus monkeys. No clini-
cal observations were noted after dose administration. Leakage of
the test article material from the vagina was noted for the rst mon-
key dosed. As such, insertion of the dosing syringe was increased
from 0.25 to 0.5 inches into the vagina for the 2 subsequent animals
tested. The modied dosing technique eliminated issues related to
leakage and loss of the test material.
Fetal blood sample collection was attempted by several routes
following euthanasia; the largest blood samples were collected
from the umbilical cord or at the site of umbilicus attachment.
Collections attempted by cardiac puncture or from the vena cava
were unsuccessful. Sufcient blood was obtained from the umbili-
cus, the umbilical cord, and fetal trunk blood harvested from the
fetus on GD 71 (Female No. 1503) such that individual bioanalysis
could be conducted on samples from each site (Table 1). Based on
the results from this single fetus, the site of blood sampling had
no effect on the concentration of metronidazole or the metabolite
hydroxymetronidazole; all values were comparable (Fig. 1).
Maternal exposures to metronidazole were variable when
assessed 7 h after a single xed volume vaginal dose (Table 2);
individual maternal plasma values ranged from 94 to 756 ng/mL.
Relatedly, there were similar variations in the fetal plasma
and amniotic uid concentrations measured for metronidazole
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Table 1 Table 5
Fetal plasma concentrations from conceptus of female No. 1503. Concentrations of 2-hydroxymetronidazole (2-HM) in amniotic uid.

Site of fetal Metronidazole 2- Metabolite-to- Female Gestation day of 2-HM (ng/mL) 2-HM-to Maternal 2-HM
blood (ng/mL) Hydroxymetronidazole parent number evaluation parent ratio plasma-to-
collection (ng/mL) ratio amniotic uid
ratio
Umbilicus 552 21.6 0.04
Umbilical cord 542 20.4 0.04 1501 70 2.01 0.01 1.8
Trunk blood 523 21.3 0.04 1502 60 8.69 0.01 3.0
Mean 539 14.7 21.1 0.0625 0.04 0 1503 71 13.4 0.02 1.5
Mean SD 8.03 5.7 0.01 0.006 2.1 0.8

to fetal plasma concentration ratios of hydroxymetronidazole were

0.91.1 (Table 4); and 1.53.0 for amniotic uid (Table 5).

4. Discussion

Previous work has shown that a 1 mg/kg intravaginal dose of

Fig. 1. Molecular structures of (A) metronidazole (molecular weight: 171) and (B)
2-hydroxymetronidazole (molecular weight: 187).
metronidazole (given as a solution formulation) in rhesus mon-
keys was approximately 74% bioavailable and resulted in systemic
concentrations of approximately 315 ng/mL [9] at 7 h postdose,
Table 2
Concentrations of metronidazole in plasma and maternal-to-fetal ratios.
the approximate Cmax . Assuming dose proportionality and simi-
lar bioavailability with the gel formulation, a 7.5 mg dose would be
Female Gestation day Maternal Fetal plasma Maternal-to- predicted to result in systemic exposures of 473 ng/mL, which is
number of evaluation plasma (ng/mL) (ng/mL) fetal plasma
ratio
very similar to the mean results measured in the present study
for metronidazole, 448 ng/mL. Concentrations of the parent and
1501 70 94.4 105 0.9
metabolite in maternal plasma, fetal plasma, and amniotic uid for
1502 60 756 735 1.0
1503 71 494 539 0.9 Female No. 1501 were lower than those observed for the latter 2
Mean SD 448 333 460 322 1.0 0.06 monkeys (Female Nos. 1502 and 1503) evaluated. This difference
was likely related to the shallower insertion of the dosing syringe
into the vagina (0.25 in.) and associated documented leakage of
Table 3
the gel from the vaginal opening following dose administration.
Concentrations of metronidazole in amniotic uid and maternal ratios.
Deeper insertion of the syringe (0.5 in.) used with the 2 other ani-
Female Gestation day Maternal Amniotic uid Maternal mals included on study resulted in no notable leakage of the gel,
number of evaluation plasma (ng/mL) plasma-to-
and relatedly, higher systemic concentrations in the mother and
(ng/mL) amniotic uid
ratio
fetus, as well as in the amniotic uid. Regardless of dosing tech-
nique, vaginal administration of drugs has been associated with
1501 70 94.4 145 0.7
increased inter-individual variability in systemic exposure relative
1502 60 756 649 1.2
1503 71 494 817 0.6 to other routes of exposure [10].
Mean SD 448 333 537 345 0.8 0.3 2-Hydroxymetronidazole has been found to be a circulating
metabolite of metronidazole in humans following oral and vagi-
nal administration of a single dose, formed by hydroxylation of the
(Tables 2 and 3). Variability among concentrations for the parent 2-methyl group [8,11]. Following a single 20-min intravenous infu-
in all 3 matrices was primarily attributed to the results from the sion of metronidazole in healthy volunteers, AUC (024 h) values
rst monkey dosed (Female No. 1501, 0.10.2 lower) relative to for 2-hydroxymetabolite were 62% that measured for metronida-
the subsequent 2 monkeys treated. However, for each respective zole [12]. Metabolism has been primarily attributed to hepatic
individual, maternal-to-fetal plasma ratios for metronidazole were biotransformation and in vitro studies implicate CYP2A6 as the pre-
similar, ranging from 0.9 to 1 (Table 2); and maternal-to-amniotic dominant catalyst of this hydroxylation (>96%; [13]). In the present
uid ratios for individuals were 0.61.2 (Table 3). study, circulating concentrations of the hydroxy-metabolite in
Similarly, exposure to the metabolite 2-hydroxymetronidazole pregnant cynomolgus monkeys as well as early stage fetuses were
demonstrated variability across individuals, with maternal plasma much lower than that observed for metronidazole (34% of parent),
concentrations of 3.6626.5 ng/mL (Table 4). As observed for demonstrating similar metabolism to humans, albeit to a lesser
the parent, concentration values for the metabolite were lowest extent. Following a single vaginal dose of metronidazole gel in
for Female No. 1501, relative to the other 2 animals evaluated. women, Cmax for hydroxymetronidazole occurred later than that
Values for the metabolite in fetal plasma and amniotic uid for the parent; 1124 h after dosing [8], as such, it is likely that in
were respectively variable (Tables 4 and 5). However, individual the absence of a full AUC prole in the present work, metabolite-to-
metabolite-to-parent ratios were calculated to be 0.04 in females, parent ratios are underestimated. Cynomolgus monkeys have been
0.030.04 in fetuses, and 0.01 to 0.02 in amniotic uid. Maternal found to express CYP2A23, CYP2A24, and CYP2A26 in liver, with

Table 4
Concentrations of 2-hydroxymetronidazole (2-HM) and exposure ratios.

Female Gestation day of Maternal plasma Maternal 2-HM-to Fetal plasma Fetal 2-HM-to Maternal-to-fetal
number evaluation (ng/mL) parent ratio (ng/mL) parent ratio 2-HM plasma ratio

1501 70 3.66 0.04 4.17 0.04 0.9

1502 60 26.5 0.04 23.4 0.03 1.1
1503 71 19.6 0.04 21.1 0.04 0.9
Mean SD 16.6 11.7 0.04 0 16.2 10.5 0.04 0.006 1.0 0.1

Please cite this article in press as: K.E. Thompson, et al., Evaluation of early fetal exposure to vaginally-administered metronidazole in
pregnant cynomolgus monkeys, Reprod Toxicol (2015), http://dx.doi.org/10.1016/j.reprotox.2015.10.013
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similar amino acid sequence (9195% identical) and functionality
to CYP2A6 in humans [14] and metabolize human CYP2A6 sub-
strates similarly [15]. In humans, RNA for CYP2A6 was not detected
in fetal livers collected at 1124 weeks of gestation [16] and CYP2A6
protein was found in only 1 of 6 fetal livers evaluated from Ges-
tation Week 1418 samples [17]. Given the immature metabolic
capacity of the early age fetuses included in the present work, it is
likely that the measured hydroxymetronidazole measured in fetal
plasma was generated by maternal metabolism and then passed
from maternal circulation to fetal, rather than generated de novo
from metronidazole within the fetus.
Interestingly, amniotic uid demonstrated similar concentra-
tions of metronidazole relative to fetal and maternal plasma, but
appeared to have lower levels of the hydroxylated metabolite. The
metabolite-to-parent ratio for hydroxymetronidzaole in amniotic
uid was 24 lower than that observed for plasma. While this is
possibly within the variation of the bioanalytical assay, especially
for a low abundance metabolite, it is unclear why distribution of
the metabolite in the amniotic uid appeared reduced relative to
plasma when similar variability was not observed for the parent.
Prior to keratinization of the fetal skin in mid gestation, amniotic
uid represents the maternal and fetal plasma. Water and solutes
diffuse bi-directionally from maternal circulation to fetus, and then
across fetal membranes to the amnionic compartment driven by
hydrostatic and osmotic gradients [18]. Even though the early kid-
ney begins to form urine by the end of organogenesis, fetal urine
represents only a minor portion of amniotic uid until the sec-
ond half of pregnancy, after the skin has keratinized. Prior to this,
amniotic uid represents an extension of the fetal extracellular
compartment, and is similar to fetal plasma, with less protein [19].
All samples in the present study were collected shortly after the
embryonic period in cynomolgus monkeys, in the early fetal phase
and prior to keratinization of the developing skin. As such, amniotic
uid samples would be predicted to be at steady-state with the fetal
plasma. Both metronidazole and its hydroxymetabolite have lim-
ited protein binding (1421% bound in plasma [20]), with similar
chemical properties (polar and soluble), predicting ready diffusion
into the fetal and amniotic spaces. Literature for amniotic uid does
not describe a metabolic capacity within this compartment in early
pregnancy. Therefore, in the absence of a plausible explanation for
a difference in the metabolite values, the apparent lower amniotic
concentrations to 2-hydroxymetronidazole were considered most
likely related to variation in the bioanalytical assay and are not
interpreted to be biologically relevant.
Importantly, it was noted that concentrations of metronida-
zole and its metabolite hydroxymetronidazole, were equivalent
between maternal and fetal circulation for all 3 animals evalu-
ated; individual maternal-to-fetal exposure ratios were close to 1,
with no indication of higher exposures in fetuses relative to that
observed in maternal circulation. These data demonstrate that the
embryo-fetal compartment had no evidence of drug accumulation
following vaginal exposure to the dam relative to maternal sys-
temic exposures, and any potential direct movement of drug from
the vagina to the conceptus was negligible.
While the present work is limited to evaluation of a single drug,
this study was conducted in the most relevant nonclinical in vivo
model. The macaque has been well established to have reproduc-
tive physiology, endocrinology, and anatomy highly similar to that
in human [21,22]. Further, cynomolgus monkeys have hemochorial,
discoidal placentas, with a similar interhaemal barrier and pattern
of circulation in the intervillous space, to that in humans, conrm-
ing macaques as the model of choice for studies evaluating drug
distribution to the embryo-fetal compartment [23]. Interestingly,
cynomolgus monkeys have also been described as an appropriate
model for transcervical movement of sperm in humans [24]. As
Please cite this article in press as: K.E. Thompson, et al., Evaluation of early fetal exposure to vaginally-administered metronidazole in
pregnant cynomolgus monkeys, Reprod Toxicol (2015), http://dx.doi.org/10.1016/j.reprotox.2015.10.013
such, the results from this study are likely highly translational to
the movement of metronidazole in pregnant women.
The ndings from this study in pregnant monkeys extend those
by Hui et al. [10], describing the exposures to thalidomide in preg-
nant rabbits and their embryos following oral and vaginal dosing,
as well as those by Thompson et al. [25] evaluating the movement
of vaginally-delivered vital dyes in pregnant rats and mice. In the
majority of cases, vaginal delivery of a small molecule drug during
pregnancy will result in exposure to the embryo-fetal compart-
ment. However, in the 4 predominant preclinical model species
used in developmental and reproductive toxicity testing, no evi-
dence for preferential distribution to the conceptus was found
following vaginal dosing to the dam. While each study evaluated
a limited number of test compounds, given the diversity of study
designs and animals models employed, the body of evidence gen-
erated by these studies portrayed a consistent outcome. Taken
together, these data suggest that there are no previously uniden-
tied risks of fetal exposure from drugs in semen, and further,
traditional modeling for embryo-fetal exposure to small molecule
drugs in semen in support of risk assessment for pharmaceutical
agents is sufciently conservative [5,26].
To apply this body of work more broadly to risk assessment for
embryo-fetal exposure to small molecule drugs in semen, a specic
evaluation of drug concentrations in semen would not be needed
in most cases. The experimental evidence generated to date has
found no evidence to suggest that the conceptus is a depot for
drug accumulation. As such, the need for risk management for the
potential sequelae of drugs in semen would be limited those agents
with pharmacological mechanisms related to teratogenesis, a geno-
toxic liability, those with no developmental NOAEL determined in
embryo-fetal development studies, and/or a developmental NOAEL
at least 3-orders of magnitude less than target clinical concentra-
tions [5,26].
Conict of interest
The authors declare that there are no conicts of interest.
Transparency document
The Transparency document associated with this article can be
found in the online version.
Acknowledgements
This research was funded by Bristol-Myers Squibb Company
and Amgen. The authors would like to acknowledge the techni-
cal staff at Charles River Laboratories for study conduct, as well as
Stephanie Rayhon for her assistance with preparation and review
of the manuscript.
References
[1] ICH Harmonised Tripartite Guideline M3(R2). Guidance on Nonclinical Safety
Studies for the Conduct of Human Clinical Trials and Marketing Authorization
for Pharmaceuticals, June 2009.
[2] ICH Harmonised Tripartite Guideline S5(R2). Guideline for Industry: detection
of Toxicity to Reproduction for Medicinal Products & Toxicity to Male
Fertility, November 2005.
[3] S. Pichini, P. Zuccaro, R. Pacici, Drugs in semen, Clin. Pharmacokinet. 26
(1994) 356373.
[4] A.D. Kashuba, J.R. Dyer, L.M. Kramer, R.H. Raasch, J.J. Eron, M.S. Cohen,
Antiretroviral-drug concentrations in semen: implications for sexual
transmission of human immunodeciency virus type 1, Antimicrob. Agents
Chemother. 43 (1999) 18171826.
[5] L. Klemmt, A.R. Scialli, The transport of chemicals in semen, Birth Defects Res.
(Part B) 74 (2005) 119131.
G Model
RTX-7197; No. of Pages 5 ARTICLE IN PRESS
K.E. Thompson et al. / Reproductive Toxicology xxx (2015) xxxxxx
[6] S. Milingos, G. Creatsas, G. Kallipolitis, D. Lolis, M. Pavlatos, D. Kaskarelis, An
appraisal of cephalexin and clindamycin concentration in seminal plasma,
Acta Eur. Fertil. 12 (1981) 312319.
[7] C.L. Chen, B.K. Beyer, W.J. Breslin, A.M. Delise, J.Y. Hui, G.J. Moffat, K.E.
Thompson, Introduction to the HESI DART drugs in semen consortium,
Reprod. Toxicol. 48 (2014) 113114.
[8] F.E. Cunningham, D.M. Kraus, L. Brubaker, J.H. Fischer, Pharmacokinetics of
intravaginal metronidazole gel, J. Clin. Pharmacol. 34 (1994) 10601065.
[9] Y.W. Chien, J. Oppermann, B. Nicolova, H.J. Lambert, Medicated tampons:
intravaginal sustained administration of metronidazole and in vitro-in vivo
relationships, J. Pharm. Sci. 71 (1982) 767771.
[10] J.Y. Hui, M. Hoffman, G. Kumar, Embryo-fetal exposure and developmental
outcome of thalidomide following oral and intravaginal administration to
pregnant rabbits, Reprod. Toxicol. 48 (2014) 115123.
[11] R. Gugler, J.C. Jensen, Interaction between cimetidine and metronidazole, N.
Engl. J. Med. 309 (1983) 15181519.
[12] M.N. Muscara, J. Pedrazzoli Jr., E.L. Miranda, J.G. Ferraz, E. Hofstatter, G. Leite,
A.F. Magalhaes, S. Leonardi, G. Ne Nucci, Plasma
hydroxy-metronidazole/metronidazole ratio in patients with liver disease
and in healthy volunteers, Br. J. Clin. Pharmacol. 40 (1995) 477480.
[13] R.E. Pearce, M. Cohen-Wolkowiez, M.R. Sampson, G.L. Kearns, The role of
human cytochrome P450 enzymes in the formation of
2-hydroxymetronidazole: CYP2A6 is the high afnity (Low Km) catalyst, Drug
Metab. Dispos. 41 (2013) 16861894.
[14] S. Uehara, N. Murayama, H. Yamazaki, Y. Uno, A novel CYP2A6 identied in
cynomolgus monkey liver metabolizes coumarin, Xenobiotica 40 (2010)
621629.
[15] Y. Li, N.Y. Li, E.M. Sellers, Comparison of CYP2A6 catalytic activity on
coumarin 7-hydroxylation in human and monkey liver microsomes, Eur. J.
Drug Metab. Pharmacokinet. 22 (1997) 295304.
[16] J. Hakkola, M. Pasanen, R. Purkunen, S. Saarikoski, O. Pelkonen, J. Maenpaa, A.
Rane, H. Raunio, Expression of xenobiotic-metabolizing cytochrome P450
forms in human adult and fetal liver, Biochem. Pharmacol. 48 (1994) 5964.
Please cite this article in press as: K.E. Thompson, et al., Evaluation of early fetal exposure to vaginally-administered metronidazole in
pregnant cynomolgus monkeys, Reprod Toxicol (2015), http://dx.doi.org/10.1016/j.reprotox.2015.10.013
5
[17] J. Gu, T. Su, Y. Chen, Q.Y. Zhang, X. Ding, Expression of biotransformation
enzymes in human fetal olfactory mucosa: potential roles in developmental
toxicity, Toxicol. Appl. Pharmacol. 165 (2000) 158162.
[18] M.A. Underwood, W.M. Gilbert, M.P. Sherman, Amniotic uid: not just fetal
urine anymore, J. Perinatol. 25 (2005) 341348.
[19] R.L. Fischer, R. Depp, Chapter 76: amniotic uid: physiology and assessment,
In: J.J. Sciarra (Ed.), Gynecol. Obstetrics, Lippincott Williams & Wilkins, vol. 3,
2004.
[20] D. Frasca, C. Dahyot-Fizelier, C. Adier, O. Mimoz, B. Debaene, W. Couet, S.
Marchand, Metronidazole and hydroxymetronidazole central nervous system
distribution: 1. microdialysis assessment of brain extracellular uid
concentrations in patients with acute brain injury, Antimicrob. Agents
Chemother. 58 (2014) 10191023.
[21] E.V. Esch, J.M. Cline, E. Buse, C.E. Wood, E.P.C.T. deRijk, G.F. Weinbauer,
Summary comparison of female reproductive system in human and the
cynomolgus monkey (Maccaca fascicularis), Toxicol. Pathol. 36 (2008)
171S172S.
[22] I.D. Bolton. Basic Physiology of Maccaca fascicularis, In: J. Bluemel, S. Korte, E.
Schenck, G.H. Weinbauer (Eds.). The Nonhuman Primate in Nonclinical Drug
Development and Safety Assessment, 2015, 6786 (http://www.ncbi.nlm.nih.
gov/pubmed/17196252).
[23] A.M. Carter, Animal models of human placentationa review, Placenta 21
(2007) S41S47.
[24] E. Behboodi, D.F. Katz, J.W. Overstreet, A.G. Hendrickx, Movement of
cynomolgus and rhesus monkey spermatozoa collected from the lower
female reproductive tract, Gamete Res. 24 (1989) 333342.
[25] K.E. Thompson, S.L. Rayhon, G. Bailey, P. Delille, M.E. McNerney, Assessment
of cervical passage of vital dyes in pregnant, nonpregnant, and mated rats and
mice, Reprod. Toxicol. (2016) (in this issue).
[26] A.R. Scialli, G. Bailey, B.K. Beyer, I.B. Bogh, W.J. Breslin, C.L. Chen, A.M. DeLise,
J.Y. Hui, G.J. Moffat, J. Stewart, K.E. Thompson, Potential seminal transport of
pharmaceuticals to the conceptus, Reprod. Toxicol. (2016) (in this issue).