ABSTRACT
Before pharmaceutical products are evaluated in humans, it is essential that they undergo a rigorous safety assessment using in vitro models and studies in preclinical species. Once products progress into the clinic, additional preclinical studies are needed to support further clinical testing. Although regulatory guidelines provide a good framework for the types of studies that should be performed, there are some areas where it is unclear how these should be applied to microbicides, what study designs should be used, whether certain tests are relevant or if additional assays are appropriate. In this chapter we provide an overview of the key issues for the preclinical development of microbicides, and describe the purpose of each of the tests along with the key considerations to be taken into account when designing the individual safety studies as well as the overall preclinical program.
Subject(s)
Anti-HIV Agents/adverse effects , Drug Delivery Systems/instrumentation , Drug Evaluation, Preclinical , HIV Infections/drug therapy , HIV-1/drug effects , Anti-HIV Agents/pharmacokinetics , Chemistry, Pharmaceutical , Clinical Trials as Topic , Drug Delivery Systems/methods , Drug Evaluation, Preclinical/instrumentation , Drug Evaluation, Preclinical/methods , HIV Infections/virology , HIV-1/physiology , HumansABSTRACT
BACKGROUND: The ex vivo challenge assay is a bio-indicator of drug efficacy and was utilized in this randomized, placebo controlled trial as one of the exploratory endpoints. Fresh and cryopreserved tissues were evaluated for human immunodeficiency virus (HIV) infection and pharmacokinetic (PK)/pharmacodynamic (PD) relationships. METHODS: HIV-negative women used vaginal rings containing 25âmg dapivirine (DPV)/100âmg maraviroc (MVC) (n = 12), DPV only (n = 12), MVC only (n = 12), or placebo (n = 12) for 28 days. Blood plasma, cervicovaginal fluid (CVF), and cervical biopsies were collected for drug quantification and the ex vivo challenge assay; half (fresh) were exposed immediately to HIV while the other half were cryopreserved, thawed, then exposed to HIV. HIV replication was monitored by p24 enzyme-linked immunosorbent assay from culture supernatant. Data were log-transformed and analyzed by linear least squared regression, nonlinear Emax dose-response model and Satterthwaite t test. RESULTS: HIV replication was greater in fresh compared to cryopreserved tissue (P = 0.04). DPV was detected in all compartments, while MVC was consistently detected only in CVF. Significant negative correlations between p24 and DPV levels were observed in fresh cervical tissue (P = 0.01) and CVF (P = 0.03), but not plasma. CVF MVC levels showed a significant negative correlation with p24 levels (P = 0.03); drug levels in plasma and tissue were not correlated with HIV suppression. p24 levels from cryopreserved tissue did not correlate to either drug from any compartment. CONCLUSION: Fresh tissue replicated HIV to greater levels and defined PK/PD relationships while cryopreserved tissue did not. The ex vivo challenge assay using fresh tissue could prioritize drugs being considered for HIV prevention.
Subject(s)
Contraceptive Devices, Female , Cyclohexanes/pharmacology , HIV Fusion Inhibitors/pharmacology , HIV Infections/prevention & control , HIV-1/drug effects , Pyrimidines/pharmacology , Sexually Transmitted Diseases, Viral/prevention & control , Triazoles/pharmacology , Administration, Intravaginal , Adult , Biopsy , Cervix Uteri/virology , Cryopreservation , Cyclohexanes/pharmacokinetics , Double-Blind Method , Enzyme-Linked Immunosorbent Assay , Female , HIV Fusion Inhibitors/pharmacokinetics , Humans , In Vitro Techniques , Maraviroc , Pyrimidines/pharmacokinetics , Triazoles/pharmacokinetics , United StatesABSTRACT
Dapivirine is a nonnucleoside reverse transcriptase inhibitor being developed as a topical microbicide for the prevention of human immunodeficiency virus infection. The distribution of radioactivity and drug in plasma and in vaginal, cervical, and draining lymph node tissues was investigated after daily application of a vaginal gel formulation of [14C]dapivirine to rhesus macaques. This was preceded by a preliminary study with rabbits. Following the intravaginal administration of [14C]dapivirine ( approximately 0.1 mg/ml [15 microCi/ml]) to rabbits (0.5 ml/day) and macaques (1 ml/day) for 7 days, the dapivirine levels associated with vaginal and cervical tissue samples 1 h after the final dose were high (quantities of microg/g of tissue) and remained detectable at 24 h (mean, >or=2.5 ng/g in rabbits) and 48 h (mean, >80 ng/g in macaques). Radioactivity levels were low in the plasma and very low or unquantifiable in the draining lymph nodes of the macaques. Microautoradiography identified drug-related material (DRM) on the surfaces of the vaginal and cervical tissues of the rabbits and macaques. Although DRM was primarily associated with the outermost layer of shedding cells in rabbits, two animals showed some evidence of small quantities in the mucosal epithelium of the cervix. In macaques, DRM was seen within the keratinized layer of the vaginal epithelium and and was found to extend into the superficial cellular layers, and in at least one animal it appeared to be present in the deepest (germinal) layer of the epithelium and in submucosal tissues. The persistence of biologically significant concentrations of dapivirine in vaginal and cervical tissues for >24 h supports the development of dapivirine as a microbicide for once daily application.