Tetradecafluorohexane

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

In-life phase: December 1-31, 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Reference

Reference Type:

other company data

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Principles of method if other than guideline:

Specific Aim
To assess the teratogenicity of AF0150 in rabbits (Segment II) Methods

Animal Preparation: New Zealand white female rabbits (98, sexually mature and virgin, 5 months old with body weight of 2525-3787 g). Standard procedures were followed for housing, handling, feeding and care of the animals. After acclimation for 26 days, animals meeting good health and acceptable body weight requirements (6 month old, 3000-4500 g) were randomly assigned to 4 groups, 22/group for insemination and treatment (Table 1).
Insemination: Semen was individually collected from 11 resident male rabbits of the same strain and supplier as the females. Semen with greater than 50% motility was diluted with saline to a final concentration of more than 3 million motility sperms/ml. The diluted semen from one male was used to inseminate two females in each group. A 0.25-0.5 ml of the diluted semen was deposited into the anterior vagina of each female with a glass insemination pipette, immediately

followed by IV injection of HCG (100 USP units) to ensure ovulation. The insemination day was designated as gestation day 0.
Table I. Teratology Study Design for AF0150 IV Injection in Rabbits
Group AF0150 Dose*
(mg/kg/day) Number of
Female Rabbits IV Volume
(ml/kg/day)
1 (Control) (Saline) 22 5.0
2 (AF0150) 50 22 1.25
3 (AF0150) 100 22 2.5
4 (AF0150) 200 22 5.0

* AF0150 was reconstituted in SWFI to final concentration of 40 mg/ml. Animals were dosed from the gestation day 7 to day 20, and sacrificed on gestation day 29.
AF0150 Preparation and Administration: AF0150 (400 mg fill) was reconstituted with 10 ml SWFI to a final concentration of 40 mg/kg, and used within 30 minutes after reconstitution. The solutions were not used if they turned clear and could not be restored (by shaking) to an opaque appearance. On the first day of dosing, AF0150 dose was verified by osmolality measurement. AF0150 solution and saline (0.9% sodium chloride for injection, as a negative control) were administered daily by IV injection via a marginal ear vein (dilated with warm water if necessary), from gestation days 7 through 20. The AF0150 dosages were 50, 100 and 200 mg/kg/day for the assigned groups, and 5 ml/kg/day of saline for the control group (Table 1) based on the most recent body weights. All animals were treated at approximately the same time each day.
Maternal Observations:
Clinical Signs: All animals were observed twice a day for mortality and moribundity, and detailed clinical signs were recorded daily till scheduled sacrifice day (gestation days 0-29).
Food consumption: Daily food consumption was recorded from gestation days 0-29 and reported as g/animal/day and g/kg/day for each corresponding body weight changes.
Body Weights and Gravid Uterine Weights: Maternal body weights were recorded on gestation days 0, 7-21 (daily), 24 and 29. A group mean body weight was calculated for each time point. Mean body weight changes were calculated for each corresponding interval and also for gestation days 7-10, 10-13, 13-21, 21-29 and 0-29. The net body weights were determined by exclusion of the uterus and content weights on gestation day 29 (scheduled laparohysterectomy) from body weight on Day 29. Net body weight changes were calculated by exclusion of the uterus and content weights from the day 0-29 body weight change.
Gestation Day 20 Laparohysterectomy: All animals were sacrificed on gestation day 29 followed by necropsy including the thoracic, abdominal, pelvic cavities and their contents. The number of corpora lutea in each ovary was recorded. The trimmed uterus was weighed and opened. The number and location of all fetuses, early and late resorptions, and the total number of implantation sites were collected.
Fetal Observations
All fetuses were examined for sex, weight, external and visceral malformation. The skeleton was examined by stereomicroscopy. External, visceral and skeletal findings were recorded as developmental variations (alterations in anatomic structure without significant biological effect, representing slight deviations from normal) or malformations (structural anomalies that alter general body conformity, disrupt or interfere with body fonction, or are generaily thought to be incompatible with life).

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Tetradecafluorohexane

EC Number:

206-585-0

EC Name:

Tetradecafluorohexane

Cas Number:

355-42-0

Molecular formula:

C6F14

IUPAC Name:

tetradecafluorohexane

Test material form:

liquid

Remarks:

Preparation for iv injection

Details on test material:

Imagent® Kit for the preparation of perflexane lipid microspheres for injectable suspension,
is a sterile, non-pyrogenic white powder with a diluted perflexane headspace that, after
reconstitution into a suspension of microspheres, is used for contrast enhancement during the
indicated ultrasound imaging procedures.
The contents of the 200 mg Imagent powder vial are sterile and non-pyrogenic. Each vial
of Imagent® powder contains 9.2 mg 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC);
75 mg hydroxyethyl starch; 2.1 mg poloxamer 188; 75 mg sodium chloride; and 36 mg
sodium phosphate buffer in a vial filled with a mixture of 17% v/v perflexane vapor in
nitrogen.
After reconstitution with 10 mL of the provided Sterile Water for Injection, USP, the
contents of the vial yield an opaque white suspension for injection. The reconstituted
suspension must be withdrawn from the vial with the supplied vented 5 µm filter dispensing
pin.
Each mL of reconstituted aqueous suspension contains a maximum of 13.7 x 108
microspheres, 92 µg perflexane, 0.92 mg DMPC; 7.5 mg hydroxyethyl starch; and 0.21 mg
poloxamer 188. The reconstituted product is iso-osmolar and has a pH between 6.7 to 7.7.
Table 1. Microsphere Size Distribution
DIAMETER
Mean Volume Weighted Median: 6 µm
Number per mL
Mean (% of Total)
ALL SIZES (Total) 5.9-13.7 x 108
(100%)
<3 µm 7 x 108
(78.8%)
3 - 10 µm 2 x 108
(21.0 %)
>10 µm 0.01 x 108
(0.2 %)
Upper limit 20 µm
The active moiety, the microsphere, comprises two critical components: perflexane, the
gaseous component, and DMPC, the lipid membrane component.
Perflexane is chemically characterized as n-perfluorohexane with a molecular weight of 338
atomic mass units and an empirical formula of C6F14. Perflexane has the following structural
formula:
FF FF F
F
F
F F F
F
F
F F
DMPC is a semi-synthetic (not of animal origin) phospholipid and is chemically
characterized as 1, 2,-dimyristoyl-sn-glycero-3-phosphocholine with a molecular weight of
678 atomic mass units and an empirical formula of C36H72NO8P. DMPC has the following
structural formula:
O H C
O
O CH2
H2C
O
O
P
O O
O
N(CH3)3 -
+
Imagent Kit for the Preparation of Perflexane-Lipid Microspheres Injectable Suspension is
supplied for single-use and each kit contains a 10-mL glass vial containing 200 mg of
Imagent powder, a 20-mL plastic vial of Sterile Water for Injection, a 10-mL disposable
plastic sterile syringe, a sterile, vented 5 µm filter dispensing pin, and a package insert.
The powder vial must be reconstituted with 10 mL supplied Sterile Water for Injection and
then withdrawn from the vial with the provided vented 5 µm filter dispensing pin as
described under DOSAGE AND ADMINISTRATION – Drug Handling and Preparation.

Test animals

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

New Zealand white female rabbits (98, sexually mature and virgin, 5 months old with body weight of 2525-3787 g). Standard procedures were followed for housing, handling, feeding and care of the animals. After acclimation for 26 days, animals meeting good health and acceptable body weight requirements (6 month old, 3000-4500 g) were randomly assigned to 4 groups, 22/group for insemination and treatment (Table 1).

Table I. Teratology Study Design for AF0150 IV Injection in Rabbits
Group AF0150 Dose*
(mg/kg/day) Number of
Female Rabbits IV Volume
(ml/kg/day)
1 (Control) (Saline) 22 5.0
2 (AF0150) 50 22 1.25
3 (AF0150) 100 22 2.5
4 (AF0150) 200 22 5.0

* AF0150 was reconstituted in SWFI to final concentration of 40 mg/ml. Animals were dosed from the gestation day 7 to day 20, and sacrificed on gestation day 29.

Administration / exposure

Route of administration:

intravenous

Vehicle:

water

Details on exposure:

AF0150 (400 mg fill) was reconstituted with 10 ml SWFI to a final concentration of 40 mg/kg, and used within 30 minutes after reconstitution. The solutions were not used if they turned clear and could not be restored (by shaking) to an opaque appearance. On the first day of dosing, AF0150 dose was verified by osmolality measurement. AF0150 solution and saline (0.9% sodium chloride for injection, as a negative control) were administered daily by IV injection via a marginal ear vein (dilated with warm water if necessary), from gestation days 7 through 20. The AF0150 dosages were 50, 100 and 200 mg/kg/day for the assigned groups, and 5 ml/kg/day of saline for the control group (Table 1) based on the most recent body weights. All animals were treated at approximately the same time each day.

Analytical verification of doses or concentrations:

yes

Remarks:

osmolality measurement.

Details on analytical verification of doses or concentrations:

On the first day of dosing, AF0150 dose was verified by osmolality measurement.

Details on mating procedure:

Insemination: Semen was individually collected from 11 resident male rabbits of the same strain and supplier as the females. Semen with greater than 50% motility was diluted with saline to a final concentration of more than 3 million motility sperms/ml. The diluted semen from one male was used to inseminate two females in each group. A 0.25-0.5 ml of the diluted semen was deposited into the anterior vagina of each female with a glass insemination pipette, immediately followed by IV injection of HCG (100 USP units) to ensure ovulation. The insemination day was designated as gestation day 0.

Duration of treatment / exposure:

from gestation days 7 through 20from gestation days 7 through 20

Frequency of treatment:

Daily IV bolus

Duration of test:

29 days

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day

Remarks:

Control

Doses / concentrations 2

Dose / conc.:

50 mg/kg bw/day

Doses / concentrations 3

Dose / conc.:

100 mg/kg bw/day

Doses / concentrations 4

Dose / conc.:

200 mg/kg bw/day

No. of animals per sex per dose:

Table I. Teratology Study Design for AF0150 IV Injection in Rabbits
Group AF0150 Dose*
(mg/kg/day) Number of
Female Rabbits IV Volume
(ml/kg/day)
1 (Control) (Saline) 22 5.0
2 (AF0150) 50 22 1.25
3 (AF0150) 100 22 2.5
4 (AF0150) 200 22 5.0

* AF0150 was reconstituted in SWFI to final concentration of 40 mg/ml. Animals were dosed from the gestation day 7 to day 20, and sacrificed on gestation day 29.

Control animals:

yes, concurrent vehicle

Examinations

Maternal examinations:

Clinical Signs: All animals were observed twice a day for mortality and moribundity, and detailed clinical signs were recorded daily till scheduled sacrifice day (gestation days 0-29).
Food consumption: Daily food consumption was recorded from gestation days 0-29 and reported as g/animal/day and g/kg/day for each corresponding body weight changes.
Body Weights and Gravid Uterine Weights: Maternal body weights were recorded on gestation days 0, 7-21 (daily), 24 and 29. A group mean body weight was calculated for each time point. Mean body weight changes were calculated for each corresponding interval and also for gestation days 7-10, 10-13, 13-21, 21-29 and 0-29. The net body weights were determined by exclusion of the uterus and content weights on gestation day 29 (scheduled laparohysterectomy) from body weight on Day 29. Net body weight changes were calculated by exclusion of the uterus and content weights from the day 0-29 body weight change.

Gestation Day 20 Laparohysterectomy: All animals were sacrificed on gestation day 29 followed by necropsy including the thoracic, abdominal, pelvic cavities and their contents. The number of corpora lutea in each ovary was recorded. The trimmed uterus was weighed and opened. The number and location of all fetuses, early and late resorptions, and the total number of implantation sites were collected.

Ovaries and uterine content:

Gestation Day 20 Laparohysterectomy: All animals were sacrificed on gestation day 29 followed by necropsy including the thoracic, abdominal, pelvic cavities and their contents. The number of corpora lutea in each ovary was recorded. The trimmed uterus was weighed and opened. The number and location of all fetuses, early and late resorptions, and the total number of implantation sites were collected.

Fetal examinations:

Fetal Observations
All fetuses were examined for sex, weight, external and visceral malformation. The skeleton was examined by stereomicroscopy. External, visceral and skeletal findings were recorded as developmental variations (alterations in anatomic structure without significant biological effect, representing slight deviations from normal) or malformations (structural anomalies that alter general body conformity, disrupt or interfere with body fonction, or are generaily thought to be incompatible with life).

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

no effects observed

Description (incidence and severity):

One animal in the 50 mg/kg/day group had a spontaneous abortion on gestation day 27 without remarkable observations at necropsy. All other animals survived to the scheduled necropsy on gestation day 29. AF0150 did not result in significant toxic effects at any time point or dose level.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

One animal in the 50 mg/kg/day group had a spontaneous abortion on gestation day 27 without remarkable observations at necropsy. All other animals survived to the scheduled necropsy on gestation day 29.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

AF0150 had no significant effects on mean body weights, body weight gains, gravid uterine weights, net body weights and net body weight gains at any dose level.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

Food consumption slightly increased in the 100 mg/kg/day A10150 group during gestation days 19-20 and 21-24 (statistically significant as compared to the control group). No food consumption change was noted in the other dose groups and time points.

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

Maternal Macroscopic Examination: At the scheduled necropsy (gestation day 29), 3, 3, 4 and 1 animals in the control, 50, 100 and 200 mg/kg/day groups, respectively, had an accessory spleen. One animal in each of the control and 100 mg/kg/day groups had cystic oviducts. One animals in the 50 mg/kg/day group had white purulent material in the right uterine horn. One female in the 200 mg/kg/day group had white amniotic fluid. No other AF0150-related internal findings were noted at any dose level.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Fetal Morphology: Malformation findings in the litters and fetuses available for morphological evaluation were summarized in Table 2. An increased frequency of malformations was noted in some fetuses from the high AF0150 dose groups (100 and 200 mg/kg/day). The NOAEL was 50 mg/kg/day (HED: 16.2 mg/kg/day and HDM: 130-fold of PCD).
Table 2. Fetal Morphological Examination
AF0150 (mg/kg/day), n=22 Rabbits
0 50 100 200
Number of Litters 17 19 18 19
Number of Fetuses 105 116 108 136
Malformation
(Fetuses/Litters)
External 0/0 0/0 3/2 1/1
Soft Tissue 0/0 0/0 0/0 1/1
Skeletal 2/2 1/1 2/2 8/5
Total Malformation
Fetuses/Litters 2/2 1/1 4/3 8/5
% Total Fetuses* 1.9 0.9 3.7 5.9
% Total Litters 11.8 5.3 16.7 26.3

*p = 0.12 with multi-group chi-square analysis

Histopathological findings: non-neoplastic:

no effects observed

Maternal developmental toxicity

Number of abortions:

effects observed, non-treatment-related

Description (incidence and severity):

One animal in the 50 mg/kg/day group had a spontaneous abortion on gestation day 27.

Pre- and post-implantation loss:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

no effects observed

Changes in number of pregnant:

no effects observed

Effect levels (maternal animals)

1

Dose descriptor:

NOAEL

Effect level:

>= 200 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

clinical signs

Remarks on result:

not determinable due to absence of adverse toxic effects

2

Dose descriptor:

NOAEL

Effect level:

>= 200 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

mortality

Remarks on result:

not determinable due to absence of adverse toxic effects

3

Dose descriptor:

NOAEL

Effect level:

>= 200 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

body weight and weight gain

Remarks on result:

not determinable due to absence of adverse toxic effects

4

Dose descriptor:

NOAEL

Effect level:

>= 200 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

food consumption and compound intake

Remarks on result:

not determinable due to absence of adverse toxic effects

5

Dose descriptor:

NOAEL

Effect level:

>= 200 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

organ weights and organ / body weight ratios

Remarks on result:

not determinable due to absence of adverse toxic effects

6

Key result

Dose descriptor:

NOAEL

Effect level:

>= 200 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

changes in pregnancy duration

effects on pregnancy duration

necropsy findings

number of abortions

pre and post implantation loss

total litter losses by resorption

Remarks on result:

not determinable due to absence of adverse toxic effects

Maternal abnormalities

Abnormalities:

no effects observed

Localisation:

not specified

Description (incidence and severity):

At the scheduled necropsy (gestation day 29), 3, 3, 4 and 1 animals in the control, 50, 100 and 200 mg/kg/day groups, respectively, had an accessory spleen. One animal in each of the control and 100 mg/kg/day groups had cystic oviducts. One animals in the 50 mg/kg/day group had white purulent material in the right uterine horn. One female in the 200 mg/kg/day group had white amniotic fluid. No other AF0150-related internal findings were noted at any dose level.

Results (fetuses)

Fetal body weight changes:

not specified

Reduction in number of live offspring:

no effects observed

Description (incidence and severity):

There were no significant differences in postimplantation loss, viable lifter size, fetal body weights, fetal sex ratios, the numbers of corpora lutea and implantation sites between the AF0150-treated animals at all dose levels and the control group.

Changes in sex ratio:

no effects observed

Description (incidence and severity):

There were no significant differences in postimplantation loss, viable lifter size, fetal body weights, fetal sex ratios, the numbers of corpora lutea and implantation sites between the AF0150-treated animals at all dose levels and the control group.

Changes in litter size and weights:

no effects observed

Description (incidence and severity):

There were no significant differences in postimplantation loss, viable lifter size, fetal body weights, fetal sex ratios, the numbers of corpora lutea and implantation sites between the AF0150-treated animals at all dose levels and the control group.

Changes in postnatal survival:

not examined

External malformations:

effects observed, treatment-related

Description (incidence and severity):

up to 3 fetuses in the 100 and/or 200 mg/kg/day group had external malformations such as microphthalmia, spina bifida, mandibular agnathia, astomia and an open left eyelid.

Skeletal malformations:

effects observed, treatment-related

Description (incidence and severity):

The malformations were noted at slightly high incidence in the 200 mg/kg/day group, shown as vertebral anomalies with or without associated rib anomalies. These malformation consisted primarily of extra arches or fused arches (smaller or larger than normal, absent or malpositioned); extra centra or fused centra (absent or malpositioned); and extra ribs or fused ribs (thickened, absent or forked). One fetus in each of the 100 and 200 mg/kg/day groups had only 5 sternebrae. One Fetus in each of the control and 200 mg/kg/day groups had skull anomalies that included an absent zygomatic arch, and/or fused frontal and/or posterior nasal portions. Two fetuses in the 200 mg/kg/day group had rib anomalies (in each case, fused and/or forked ribs). Fused sternebrae were noted in one fetus of the 200 mg/kg/day group.

Visceral malformations:

effects observed, treatment-related

Description (incidence and severity):

The visceral malformations were shown in one fetus of the 200 mg/kg/day group, including hydrocephaly with increased cavitation of both lateral ventricles and the third ventricle.

Effect levels (fetuses)

1

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 100 mg/kg bw (total dose)

Based on:

test mat.

Sex:

not specified

Basis for effect level:

external malformations

2

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 100 mg/kg bw/day

Based on:

test mat.

Sex:

not specified

Basis for effect level:

visceral malformations

3

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 50 mg/kg bw/day

Based on:

test mat.

Sex:

not specified

Basis for effect level:

skeletal malformations

4

Key result

Dose descriptor:

NOAEL

Effect level:

>= 200 mg/kg bw/day

Based on:

test mat.

Sex:

not specified

Basis for effect level:

reduction in number of live offspring

changes in sex ratio

changes in litter size and weights

Remarks on result:

not determinable due to absence of adverse toxic effects

Fetal abnormalities

Key result

Abnormalities:

effects observed, treatment-related

Localisation:

external: eye

external: face

skeletal: skull

skeletal: sternum

skeletal: supernumerary rib

visceral/soft tissue: integumentary

Description (incidence and severity):

External and Visceral Malformations: up to 3 fetuses in the 100 and/or 200 mg/kg/day group had external malformations such as microphthalmia, spina bifida, mandibular agnathia, astomia and an open left eyelid. The visceral malformations were shown in one fetus of the 200 mg/kg/day group, including hydrocephaly with increased cavitation of both lateral ventricles and the third ventricle.
Fetal Skeletal Malformations: The malformations were noted at slightly high incidence in the 200 mg/kg/day group, shown as vertebral anomalies with or without associated rib anomalies. These malformation consisted primarily of extra arches or fused arches (smaller or larger than normal, absent or malpositioned); extra centra or fused centra (absent or malpositioned); and extra ribs or fused ribs (thickened, absent or forked). One fetus in each of the 100 and 200 mg/kg/day groups had only 5 sternebrae. One Fetus in each of the control and 200 mg/kg/day groups had skull anomalies that included an absent zygomatic arch, and/or fused frontal and/or posterior nasal portions. Two fetuses in the 200 mg/kg/day group had rib anomalies (in each case, fused and/or forked ribs). Fused sternebrae were noted in one fetus of the 200 mg/kg/day group.

Overall developmental toxicity

Key result

Developmental effects observed:

yes

Lowest effective dose / conc.:

100 mg/kg bw/day

Treatment related:

yes

Relation to maternal toxicity:

developmental effects in the absence of maternal toxicity effects

Dose response relationship:

yes

Relevant for humans:

not specified

Any other information on results incl. tables

Table 2.Fetal Morphological Examination
	AF0150 (mg/kg/day),n=22 Rabbits
	0	50	100	200
Number of Litters	17	19	18	19
Number of Fetuses	105	116	108	136
Malformation
(Fetuses/Litters)
External	0/0	0/0	3/2	1/1
Soft Tissue	0/0	0/0	0/0	1/1
Skeletal	2/2	1/1	2/2	8/5
Total Malformation
Fetuses/Litters	2/2	1/1	4/3	8/5
% Total Fetuses*	1.9	0.9	3.7	5.9
%Total Litters	11.8	5.3	16.7	26.3

 

_*p = 0.12 with multi-group chi-square analysis

Applicant's summary and conclusion

Conclusions:

NOAEL for malformfoetusations was 50 mg/kg/day.

Executive summary:

Pregnant rabbits received IV injection of AF0150 at the doses of 50, 100 and 200 mg/kg/day from gestation days 7 through 20 and the animals were sacrificed on gestation day 29. A slight increase in fetal malformation incidence was observed at the high dose levels, particularly in the 200 mg/kg/day group, as compared to the control group. These malformations included external, visceral (soft tissue) and skeletal anomalies. Percentage of fetuses with malformations was 1.9% in control rabbits, 3.7% and 5.9% in rabbits treated with 100 and 200 mg/kg/day AF0150, respectively. One animal in the 50 mg/kg/day group had a spontaneous abortion on gestation day 27. The NOAEL for malformfoetusations was 50 mg/kg/day (HED: 16.2 mg/kg/day and HDM: 130- fold of PCD).

AF10150 treatment did not affect post-implantation loss, live litter size, mean fetal body weights, fetal sex ratios, fetal developmental variation, the mean number of corpora lutea and implantation sites, as compared to the control group.

No maternal toxicity (clinical signs, body weigh and food consumption changes, and macroscopic examination) was observed at any AF0150 dose level. The highest AF0150 dose (200 mg/kg/day) did not resuit in minimal maternal toxicity. TheNOAELfor maternal toxicity was 200 mg/kg/day.