title: Synthesis, characterization and preclinical formulation of a dual-action phenyl phosphate derivative of bromo-methoxy zidovudine (compound WHI-07) with potent anti-HIV and spermicidal activities.
abstract: In a systematic effort to develop a microbicide contraceptive capable of preventing transmission of human immunodeficiency virus (HIV), as well as providing fertility control, we have previously identified novel phenyl phosphate derivatives of zidovudine (ZDV) with 5-halo 6-alkoxy substitutions in the thymine ring and halo substitution in the phenyl moiety respectively. Here, we describe the synthesis, characterization, and successful preclinical formulation of our lead compound, 5-bromo-6-methoxy-3'-azidothymidine-5'-(p-bromophenyl) methoxyalaninyl phosphate (WHI-07), which exhibits potent anti-HIV and sperm immobilizing activities. The anti-HIV activity of WHI-07 was tested by measuring viral p24 antigen production and reverse transcriptase activity as markers of viral replication in HIV-1 infected human peripheral blood mononuclear cells (PBMC). WHI-07 inhibited replication of HIV in a concentration-dependent fashion with nanomolar IC50 values. The effects of WHI-07 on human sperm motion kinematics were analysed by computer-assisted sperm analysis (CASA), and its effects on sperm membrane integrity were examined by confocal laser scanning microscopy (CLSM), and high-resolution low-voltage scanning electron microscopy (HR-LVSEM). WHI-07 caused cessation of sperm motility in a concentration- and time-dependent fashion. The in-vitro cytotoxicities of WHI-07 and nonoxynol-9 (N-9) were compared using normal human ectocervical and endocervical epithelial cells by the MTT cell viability assay. Unlike N-9, WHI-07 had no effect upon sperm plasma and acrosomal membrane integrity. N-9 was cytotoxic to normal human ectocervical and endocervical cells at spermicidal doses, whereas WHI-07 was selectively spermicidal. The in-vivo vaginal absorption and vaginal toxicity of 2% gel-microemulsion of WHI-07 was studied in the rabbit model. The sperm immobilizing activity of WHI-07 was 18-fold more potent than that of N-9. Over a 10 day period, there was no irritation or local toxicity to the vaginal epithelia or systemic absorption of WHI-07. Therefore, as a potent anti-HIV agent with spermicidal activity, and lack of mucosal toxicity, WHI-07 may have the clinical potential to become the active ingredient of a vaginal contraceptive for women who are at high risk for acquiring HIV by heterosexual vaginal transmission.
authors.affiliation: Department of Reproductive Biology, Hughes Institute, St Paul, MN 55113, USA.
authors.forename: O J
authors.initials: OJ
authors.lastname: D'Cruz
authors.forename: M J
authors.initials: MJ
authors.lastname: Shih
authors.forename: S H
authors.initials: SH
authors.lastname: Yiv
authors.forename: C L
authors.initials: CL
authors.lastname: Chen
authors.forename: F M
authors.initials: FM
authors.lastname: Uckun
mesh_terms: D020101:Acrosome Reaction / Q000187:drug effects; D000818:Animals; D019380:Anti-HIV Agents / Q000138:chemical synthesis / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects; D002584:Cervix Uteri / Q000166:cytology / Q000187:drug effects; D054306:Dideoxynucleotides; D004353:Drug Evaluation, Preclinical; D004655:Emulsions / Q000737:chemistry / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects; D005260:Female; D015497:HIV-1 / Q000187:drug effects; D006801:Humans; D008297:Male; D011817:Rabbits; D013081:Sperm Motility / Q000187:drug effects; D013089:Spermatocidal Agents / Q000138:chemical synthesis / Q000494:pharmacology*; D013094:Spermatozoa / Q000187:drug effects; D013938:Thymidine Monophosphate / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000494:pharmacology; D014621:Vagina / Q000187:drug effects / Q000473:pathology; D014779:Virus Replication / Q000187:drug effects; D015215:Zidovudine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology
chemical_list: C114570:5-bromo-6-methoxy-5, 6-dihydro-3'-azidothymidine-5'-(p-bromophenyl) methoxyalaninyl phosphate; D019380:Anti-HIV Agents; D054306:Dideoxynucleotides; D004655:Emulsions; D013089:Spermatocidal Agents; D013938:Thymidine Monophosphate; D015215:Zidovudine