18 Feb

Aryl Phosphate Derivatives of Bromo-Methoxy-Azidothymidine: RESULTS(10)


In Vivo Contraceptive Efficacy of WHI-07

Because fertilization in vivo is dependent on successful sperm transport through the female genital tract, we next determined whether exposure of sperm to the lead compound, WHI-07, in vivo affected the subsequent fertility outcome. Hormonally primed adult female Swiss (CD-1) mice were artificially inseminated with motile epididy-mal sperm via the cervix with and without prior intravaginal application of 1% WHI-07 in a cream base. ventolin 100 mcg

Females were examined 8 days later for the presence or absence of embryos in uteri. The results of three fertility trials are summarized in Table 3. In mice receiving WHI-07 intravagin-ally prior to artificial insemination, the percentage of fertility rate was drastically reduced in comparison to that in controls (7.6% vs. 38.4%; p < 0.001), which demonstrates a substantially impaired ability of WHI-07-exposed sperm to reach the site of fertilization.

Lack of Local Toxicity of WHI-07

Next, the local tissue alterations and inflammatory response to repetitive intravaginal applications of N-9 versus WHI-07 in mice were compared. Two groups of 15 adult female Swiss (CD-1) mice were treated for 5 (group A) or 20 (group B) consecutive days with either 5% N-9, 5% WHI-07, or control vehicle in a cream base, and the cer-vicovaginal tissue sections were examined for histopatho-logical changes and influx of inflammatory cells (Fig. 7).
Fig7Aryl Phosphate Derivatives
FIG. 7. A-D) Histological and confocal fluorescence images of WHI-07- and N-9-treated mouse cervicovaginal sections. Left) Representative hematoxylin- and eosin-stained, paraffin-embedded sections of the cervicovaginal region of mouse treated intravaginally with 5% WHI-07 (A) or 5% N-9 (B) in a cream base for 20 and 5 consecutive days, respectively. The mucosa consisted of stratified squamous epithelium (SE) over a vascular submucosa. Note the lack of inflammatory infiltrate of neutrophils and the intactness of squamous epithelia in WHI-07-treated sections in contrast to N-9-treated sections (arrow); original magnification: X200-300. Right) Two-color laser scanning confocal fluorescence images of the cervicovaginal region of mice treated intravaginally with 5% WHI-07 (C) or 5% N-9 (D) for 20 and 5 consecutive days, respectively. Double labeling of the cervicovaginal epithelia with a rat monoclonal antibody to neutrophils and FITC-conjugated anti-rat IgG as the secondary antibody and counterstaining of cell nuclei with PI (red), showed lack of neutrophils (green fluorescence) in WHI-07-treated tissue section, while N-9-treated cervicovaginal epithelia shows clusters (arrow) of brightly green fluorescent neutrophils. L, Lumen; SE, squamous epithelium; original magnification X300-400.

TABLE 3. Fertility of female mice after artificial insemination of sperm via cervix with and without intravaginal application of WHI-07.

Experiment No. of mice inseminated/ group No. of control mice fertileab (%) No. of WHI-07-treated mice fertile’(%)
1 27 10 2
2 27 11 1
3 24 9 3
Total 78 30 6 d

a Animals were killed on Day 8 of the expected pregnancy. b Fertility was determined by examining the uteri for the presence or absence of embryos.
c Mice were given intravaginally (50 ^l) of a single dose of 1% WHI-07 via a cream base prior to artificial insemination. d Significantly different from control group (p < 0.001).

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