title: Differential Mechanisms of Tenofovir and Tenofovir Disoproxil Fumarate Cellular Transport and Implications for Topical Preexposure Prophylaxis.
abstract: Intravaginal rings releasing tenofovir (TFV) or its prodrug, tenofovir disoproxil fumarate (TDF), are being evaluated for HIV and herpes simplex virus (HSV) prevention. The current studies were designed to determine the mechanisms of drug accumulation in human vaginal and immune cells. The exposure of vaginal epithelial or T cells to equimolar concentrations of radiolabeled TDF resulted in over 10-fold higher intracellular drug levels than exposure to TFV. Permeability studies demonstrated that TDF, but not TFV, entered cells by passive diffusion. TDF uptake was energy independent but its accumulation followed nonlinear kinetics, and excess unlabeled TDF inhibited radiolabeled TDF uptake in competition studies. The carboxylesterase inhibitor bis-nitrophenyl phosphate reduced TDF uptake, suggesting saturability of intracellular carboxylesterases. In contrast, although TFV uptake was energy dependent, no competition between unlabeled and radiolabeled TFV was observed, and the previously identified transporters, organic anion transporters (OATs) 1 and 3, were not expressed in human vaginal or T cells. The intracellular accumulation of TFV was reduced by the addition of endocytosis inhibitors, and this resulted in the loss of TFV antiviral activity. Kinetics of drug transport and metabolism were monitored by quantifying the parent drugs and their metabolites by high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). Results were consistent with the identified mechanisms of transport, and the exposure of vaginal epithelial cells to equimolar concentrations of TDF compared to TFV resulted in ∼40-fold higher levels of the active metabolite, tenofovir diphosphate. Together, these findings indicate that substantially lower concentrations of TDF than TFV are needed to protect cells from HIV and HSV-2.
authors.affiliation: Department of Microbiology & Immunology, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Ekaterina
authors.identifier: http://orcid.org/0000-0002-4884-1373
authors.initials: E
authors.lastname: Taneva
authors.affiliation: Particle Sciences, Inc., Bethlehem, Pennsylvania, USA.
authors.forename: Kerry
authors.initials: K
authors.lastname: Crooker
authors.affiliation: Department of Biomedical Engineering, Northwestern University, Evanston, Illinois, USA Department of Chemistry, Northwestern University, Evanston, Illinois, USA Chemistry of Life Processes Institute, Northwestern University, Evanston, Illinois, USA.
authors.forename: Sung Hyun
authors.initials: SH
authors.lastname: Park
authors.affiliation: Department of Biomedical Engineering, Northwestern University, Evanston, Illinois, USA.
authors.forename: Jonathan T
authors.initials: JT
authors.lastname: Su
authors.affiliation: Department of Biomedical Engineering, Northwestern University, Evanston, Illinois, USA.
authors.forename: Adina
authors.initials: A
authors.lastname: Ott
authors.affiliation: Department of Pediatrics, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Natalia
authors.initials: N
authors.lastname: Cheshenko
authors.affiliation: Department of Biomedical Engineering, Northwestern University, Evanston, Illinois, USA Department of Chemistry, Northwestern University, Evanston, Illinois, USA Chemistry of Life Processes Institute, Northwestern University, Evanston, Illinois, USA.
authors.forename: Igal
authors.initials: I
authors.lastname: Szleifer
authors.affiliation: Department of Biomedical Engineering, Northwestern University, Evanston, Illinois, USA.
authors.forename: Patrick F
authors.initials: PF
authors.lastname: Kiser
authors.affiliation: Particle Sciences, Inc., Bethlehem, Pennsylvania, USA.
authors.forename: Bruce
authors.initials: B
authors.lastname: Frank
authors.affiliation: Department of Pediatrics, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Pedro M M
authors.initials: PM
authors.lastname: Mesquita
authors.affiliation: Department of Microbiology & Immunology, Albert Einstein College of Medicine, Bronx, New York, USA Department of Pediatrics, Albert Einstein College of Medicine, Bronx, New York, USA betsy.herold@einstein.yu.edu.
authors.forename: Betsy C
authors.initials: BC
authors.lastname: Herold
mesh_terms: D000282:Administration, Intravaginal; D019380:Anti-HIV Agents / Q000627:therapeutic use; D001692:Biological Transport / Q000187:drug effects*; D002265:Carboxylic Ester Hydrolases / Q000378:metabolism; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D004705:Endocytosis / Q000187:drug effects; D004847:Epithelial Cells / Q000187:drug effects*; D005260:Female; D015658:HIV Infections / Q000188:drug therapy / Q000517:prevention & control*; D015497:HIV-1 / Q000187:drug effects*; D006558:Herpes Genitalis / Q000188:drug therapy / Q000517:prevention & control*; D018258:Herpesvirus 2, Human / Q000187:drug effects*; D006801:Humans; D009596:Nitrophenols / Q000494:pharmacology; D009943:Organophosphorus Compounds / Q000494:pharmacology; D013601:T-Lymphocytes / Q000187:drug effects; D053719:Tandem Mass Spectrometry; D000068698:Tenofovir / Q000008:administration & dosage / Q000494:pharmacology*
chemical_list: D019380:Anti-HIV Agents; D009596:Nitrophenols; D009943:Organophosphorus Compounds; C008644:nitrophenylphosphate; D000068698:Tenofovir; D002265:Carboxylic Ester Hydrolases