Cervicovaginal safety of the formulated, biguanide-based human immunodeficiency virus type 1 (HIV-1) inhibitor NB325 in a murine model

Abstract

Vaginal microbicides that reduce or eliminate the risk of HIV-1 sexual transmission must do so safely without adversely affecting the integrity of the cervicovaginal epithelium. The present studies were performed to assess the safety of the biguanide-based antiviral compound NB325 in a formulation suitable for topical application. Experiments were performed using a mouse model of cervicovaginal microbicide application, which was previously shown to be predictive of topical agent toxicity revealed in microbicide clinical trials. Mice were exposed vaginally to unformulated NB325 or NB325 formulated in the hydroxyethyl cellulose "universal placebo." Following exposures to formulated 1% NB325 for 10 min to 24 h, the vaginal and cervical epithelia were generally intact, although some areas of minimal vaginal epithelial damage were noted. Although formulated NB325 appeared generally safe for application in these studies, the low but observable level of toxicity suggests the need for improvements in the compound and/or formulation.

Figure 1

Exposure to saline results in no damage to the cervicovaginal epithelia. Mice were exposed to saline and sacrificed at 10 min, 2 h, 4 h, or 8 h after application. H&E-stained vaginal (a) and cervical (b) epithelial tissues were analyzed for damage. Representative micrograph fields from the 2 h exposure are shown. Images are representative of three mice per exposure duration.

Figure 2

Exposure to N-9 results in severe damage to cervical epithelial tissues. Mice were exposed to N-9 and sacrificed at 10 min, 2 h, 4 h, 8 h, or 24 h after application. H&E-stained vaginal ((a), (c), (e)) and cervical ((b), (d), (f)) epithelial tissues were analyzed for damage. Representative micrograph fields from the 2 h, 4 h, and 8 h exposures are shown. Images are representative of three mice per exposure duration. Tissues exposed to N-9 for 10 min and 24 h were similar to saline-exposed tissues (data not shown).

Figure 3

Short exposures to 1% unformulated NB325 result in isolated damage to the vaginal epithelium. Mice were exposed to 1% unformulated NB325 for 10 min, 2 h, or 4 h prior to sacrifice and cervicovaginal tissue isolation. H&E-stained vaginal ((a), (c), (e)) and cervical ((b), (d), (f)) tissue sections were analyzed for epithelial damage. Representative micrograph fields for each exposure are shown. Images are representative of three mice per exposure duration.

Figure 4

Exposure to 1% unformulated NB325 for 8 h or 24 h is characterized by little or no change in cervicovaginal epithelial integrity. Mice were exposed to 1% unformulated NB325 for 8 h or 24 h prior to sacrifice and cervicovaginal tissue isolation. H&E-stained tissue sections were analyzed for damage to the vaginal ((a), (c)) and cervical ((b), (d)) epithelia. Representative micrograph fields for each exposure are shown. Images are representative of three mice per exposure duration.

Figure 5

Application of gel only (placebo) causes no damage to the cervicovaginal epithelium. Mice were exposed to the HEC-based formulation placebo and sacrificed after application for 10 min, 2 h, 4 h, or 8 h. Vaginal (a) and cervical epithelial tissues (b) were subsequently isolated and analyzed for damage. Representative micrograph fields from the 2 h exposure are shown. Images are representative of three mice per exposure duration.

Figure 6

Short exposures to NB325 formulated at 1% result in mild vaginal epithelial toxicity. Mice were exposed to 1% formulated NB325 for 10 min, 2 h, or 4 h prior to sacrifice and isolation of reproductive tract tissues. H&E-stained tissue sections were analyzed for toxicity to the vaginal ((a), (c), (e)) and cervical ((b), (d), (f)) epithelia. Representative micrograph fields for each exposure are shown. Images are representative of three mice per exposure duration.

Figure 7

Cervicovaginal epithelial integrity after exposure to 1% formulated NB325 for 8 h or 24 h is similar to that of controls. Mice were exposed to 1% formulated NB325 for 8 h or 24 h prior to sacrifice and isolation of reproductive tract tissues. H&E-stained tissue sections were analyzed for toxicity to the vaginal ((a), (c)) and cervical ((b), (d)) epithelia. Representative micrograph fields for each exposure are shown. Images are representative of three mice per exposure duration.