title: Carrier-mediated delivery improves the efficacy of 9-(2-phosphonylmethoxyethyl)adenine against hepatitis B virus.
abstract: We recently synthesized a lipophilic prodrug of 9-(2-phosphonyl-methoxyethyl)adenine (PMEA), designated PMEA-LO, and incorporated it into reconstituted lactosylated high-density lipoprotein (LacNeoHDL). In a rat model, LacNeoHDL-associated PMEA-LO was internalized by the asialoglycoprotein receptor on parenchymal liver cells and converted into its active diphosphorylated metabolite. To further evaluate the therapeutic potential of the carrier-associated prodrug, we examined in this study the processing of (125)I-labeled PMEA-LO--loaded LacNeoHDL by HepG2 cells. Upon incubation with HepG2 cells, PMEA-LO--loaded LacNeoHDL became rapidly cell-associated. The association was saturable and of high-affinity (k(d) = 3.8 +/- 0.4 nM). Asialofetuin, an established ligand for the asialoglycoprotein receptor, inhibited the association by >75%, which confirms the role of the asialoglycoprotein receptor. Association of the prodrug-loaded particles to HepG2 cells was coupled to degradation. Radiolabeled degradation products appeared in the culture medium with a lag phase of 2 h. Asialofetuin and chloroquine inhibited secretion of degradation products by 75 to 80%, indicating that PMEA-LO--loaded LacNeoHDL is internalized via the asialoglycoprotein receptor and lysosomally processed. The therapeutic potential of LacNeoHDL-associated PMEA-LO was studied by measuring its effects on hepatitis B virus (HBV) replication in Hep AD38 cells (HBV-transfected HepG2 cells). LacNeoHDL-associated PMEA-LO effectively inhibited HBV DNA synthesis. The EC(50) value of carrier-associated PMEA-LO was 35 times lower than that of free PMEA (3.4 +/- 0.4 and 120 +/- 18 ng of PMEA/ml, respectively). We conclude that the present results, combined with our earlier in vivo disposition data, underscore the therapeutic potential and utility of PMEA-LO--loaded LacNeoHDL for treatment of chronic hepatitis B.
authors.affiliation: Division of Biopharmaceutics, Leiden/Amsterdam Center for Drug Research, University of Leiden, Leiden, The Netherlands. bijsterb@lacdr.leidenuniv.nl
authors.forename: M K
authors.initials: MK
authors.lastname: Bijsterbosch
authors.forename: C
authors.initials: C
authors.lastname: Ying
authors.forename: R L
authors.initials: RL
authors.lastname: de Vrueh
authors.forename: E
authors.initials: E
authors.lastname: de Clercq
authors.forename: E A
authors.initials: EA
authors.lastname: Biessen
authors.forename: J
authors.initials: J
authors.lastname: Neyts
authors.forename: T J
authors.initials: TJ
authors.lastname: van Berkel
mesh_terms: D000225:Adenine / Q000008:administration & dosage / Q000031:analogs & derivatives / Q000494:pharmacology*; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000008:administration & dosage / Q000494:pharmacology*; D004337:Drug Carriers; D016503:Drug Delivery Systems; D006515:Hepatitis B virus / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D007785:Lactose / Q000378:metabolism; D008075:Lipoproteins, HDL; D008095:Lithocholic Acid / Q000737:chemistry; D008826:Microbial Sensitivity Tests; D063065:Organophosphonates; D014407:Tumor Cells, Cultured; D014779:Virus Replication / Q000187:drug effects
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D004337:Drug Carriers; D008075:Lipoproteins, HDL; D063065:Organophosphonates; D008095:Lithocholic Acid; C053001:adefovir; D007785:Lactose; D000225:Adenine