Fullerene derivatives with hydrophilic substituents have been shown to exhibit a range of biological activities, including antiviral ones. For a long time, the anti-HIV activity of fullerene derivatives was believed to be due to their binding into the hydrophobic pocket of HIV-1 protease, thereby blocking its activity. Recent work, however, brought new evidence of a novel, protease-independent mechanism of fullerene derivatives' action. We studied in more detail the mechanism of the anti-HIV-1 activity of N,N-dimethyl[70]fulleropyrrolidinium iodide fullerene derivatives. By using a combination of in vitro and cell-based approaches, we showed that these C70 derivatives inhibited neither HIV-1 protease nor HIV-1 maturation. Instead, our data indicate effects of fullerene C70 derivatives on viral genomic RNA packaging and HIV-1 cDNA synthesis during reverse transcription-without impairing reverse transcriptase activity though. Molecularly, this could be explained by a strong binding affinity of these fullerene derivatives to HIV-1 nucleocapsid domain, preventing its proper interaction with viral genomic RNA, thereby blocking reverse transcription and HIV-1 infectivity. Moreover, the fullerene derivatives' oxidative activity and fluorescence quenching, which could be one of the reasons for the inconsistency among reported anti-HIV-1 mechanisms, are discussed herein.
- MeSH
- fullereny metabolismus farmakologie MeSH
- genom virový účinky léků MeSH
- genové produkty gag - virus lidské imunodeficience metabolismus MeSH
- HEK293 buňky MeSH
- HIV-1 účinky léků genetika metabolismus fyziologie MeSH
- látky proti HIV metabolismus farmakologie MeSH
- lidé MeSH
- nukleokapsida - proteiny metabolismus MeSH
- reverzní transkripce MeSH
- RNA virová metabolismus MeSH
- svlékání virového obalu účinky léků MeSH
- vazba proteinů MeSH
- virion metabolismus MeSH
- zabalení virového genomu účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Special attention is required when pharmacological treatment is indicated for a pregnant woman. P-glycoprotein (MDR1) is a well-known transporter localized in the maternal blood-facing apical membrane of placental syncytiotrophoblast and is considered to play an important role in protecting the developing fetus. Maraviroc, a MDR1 substrate that is registered for treatment of HIV infection, shows a low toxicity profile, suggesting favorable tolerability also if administered to pregnant women. Nevertheless, there is only poor understanding to date regarding the extent to which it permeates across the placental barrier and what are the transport mechanisms involved. Endeavoring to clarify the passage of maraviroc across placenta, we used in this study the method of closed-circuit perfusion of maraviroc across human placental cotyledon. The data obtained confirmed slight involvement of MDR1, but they also suggest possible interaction with other transport system(s) working in the opposite direction from that of MDR1. Complementary in vitro studies, including cellular experiments on choriocarcinoma BeWo cells as well as transporter-overexpressing MDCKII and A431 cell lines and accumulation in placental fresh villous fragments, revealed maraviroc transport by MRP1, OATP1A2, and OATP1B3 transporters. Based on mRNA expression data in the placental tissue, isolated trophoblasts, and fetal endothelial cells, especially MRP1 and OATP1A2 seem to play a crucial role in cooperatively driving maraviroc into placental tissue. By the example of maraviroc, we show here the important interplay of transporters in placental drug handling and its possibility to overcome the MDR1-mediated efflux.
- MeSH
- akridiny farmakologie MeSH
- buňky MDCK MeSH
- látky proti HIV krev metabolismus farmakologie MeSH
- lékové interakce MeSH
- lidé MeSH
- maravirok krev metabolismus MeSH
- nádorové buněčné linie MeSH
- P-glykoproteiny antagonisté a inhibitory genetika metabolismus MeSH
- perfuze MeSH
- placenta účinky léků metabolismus MeSH
- placentární oběh MeSH
- přenašeče organických aniontů antagonisté a inhibitory genetika metabolismus MeSH
- protein OATP1B3 antagonisté a inhibitory genetika metabolismus MeSH
- proteiny spojené s mnohočetnou rezistencí k lékům genetika metabolismus MeSH
- psi MeSH
- regulace genové exprese MeSH
- ritonavir farmakologie MeSH
- těhotenství MeSH
- tetrahydroisochinoliny farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- psi MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- AIDS * farmakoterapie MeSH
- antiretrovirové látky farmakologie metabolismus MeSH
- dějiny 20. století MeSH
- dějiny 21. století MeSH
- HIV infekce * farmakoterapie MeSH
- kombinovaná farmakoterapie MeSH
- látky proti HIV farmakologie metabolismus MeSH
- lidé MeSH
- objevování léků trendy MeSH
- vakcíny proti AIDS MeSH
- Check Tag
- dějiny 20. století MeSH
- dějiny 21. století MeSH
- lidé MeSH
Abacavir is a preferred antiretroviral drug for preventing mother-to-child human immunodeficiency virus transmission; however, mechanisms of its placental transfer have not been satisfactorily described to date. Because abacavir is a nucleoside-derived drug, we hypothesized that the nucleoside transporters, equilibrative nucleoside transporters (ENTs, SLC29A) and/or Na+-dependent concentrative nucleoside transporters (CNTs, SLC28A), may play a role in its passage across the placenta. To test this hypothesis, we performed uptake experiments using the choriocarcinoma-derived BeWo cell line, human fresh villous fragments, and microvillous plasma membrane (MVM) vesicles. Using endogenous substrates of nucleoside transporters, [3H]-adenosine (ENTs, CNT2, and CNT3) and [3H]-thymidine (ENTs, CNT1, and CNT3), we showed significant activity of ENT1 and CNT2 in BeWo cells, whereas experiments in the villous fragments and MVM vesicles, representing a model of the apical membrane of a syncytiotrophoblast, revealed only ENT1 activity. When testing [3H]-abacavir uptakes, we showed that of the nucleoside transporters, ENT1 plays the dominant role in abacavir uptake into placental tissues, whereas contribution of Na+-dependent transport, most likely mediated by CNTs, was observed only in BeWo cells. Subsequent experiments with dually perfused rat term placentas showed that Ent1 contributes significantly to overall [3H]-abacavir placental transport. Finally, we quantified the expression of SLC29A in first- and third-trimester placentas, revealing that SLC29A1 is the dominant isoform. Neither SLC29A1 nor SLC29A2 expression changed over the course of placental development, but there was considerable interindividual variability in their expression. Therefore, drug-drug interactions and the effect of interindividual variability in placental ENT1 expression on abacavir disposition into fetal circulation should be further investigated to guarantee safe and effective abacavir-based combination therapies in pregnancy.
- MeSH
- adenosin metabolismus MeSH
- biologický transport fyziologie MeSH
- dideoxynukleosidy metabolismus MeSH
- ekvilibrační přenašeč nukleosidů 1 metabolismus MeSH
- ekvilibrační přenašeč nukleosidů 2 metabolismus MeSH
- krysa rodu rattus MeSH
- látky proti HIV metabolismus MeSH
- lidé MeSH
- membránové transportní proteiny metabolismus MeSH
- nádorové buněčné linie MeSH
- nukleosidy metabolismus MeSH
- placenta metabolismus MeSH
- potkani Wistar MeSH
- proteiny přenášející nukleosidy metabolismus MeSH
- těhotenství MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Tenofovir disoproxil fumarate (TDF, form I) is an orally delivered pharmaceutical salt used for the treatment of HIV and chronic hepatitis, which acts as an inhibitor of nucleotide reverse transcriptase. There are many solid forms of TDF described in the literature; 2 of them were identified in the drug products: form I and form A. It seems that during formulation, the active pharmaceutical ingredient undergoes partial to total conversion of TDF form I to TDF form A. The goals of this study were to investigate when and why did the conversion occur and whether the conversion could be avoided and how. The influence of pH and possible interaction with excipients were studied. The conditions enabling using wet granulation in technology while preventing the undesired conversion were found. The stabilization was achieved either by replacement of used disintegrants or by acid addition to the current composition of formulation.
- MeSH
- difrakce rentgenového záření metody MeSH
- koncentrace vodíkových iontů MeSH
- látky proti HIV chemie metabolismus MeSH
- pomocné látky chemie metabolismus MeSH
- příprava léků metody MeSH
- stabilita léku MeSH
- tenofovir chemie metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Assembly of human immunodeficiency virus (HIV-1) represents an attractive target for antiretroviral therapy which is not exploited by currently available drugs. We established high-throughput screening for assembly inhibitors based on competition of small molecules for the binding of a known dodecapeptide assembly inhibitor to the C-terminal domain of HIV-1 CA (capsid). Screening of >70000 compounds from different libraries identified 2-arylquinazolines as low micromolecular inhibitors of HIV-1 capsid assembly. We prepared focused libraries of modified 2-arylquinazolines and tested their capacity to bind HIV-1 CA to compete with the known peptide inhibitor and to prevent the replication of HIV-1 in tissue culture. Some of the compounds showed potent binding to the C-terminal domain of CA and were found to block viral replication at low micromolar concentrations.
- MeSH
- chinazoliny chemická syntéza chemie farmakologie MeSH
- HIV-1 účinky léků metabolismus MeSH
- kapsida účinky léků metabolismus MeSH
- knihovny malých molekul MeSH
- látky proti HIV metabolismus farmakologie MeSH
- lidé MeSH
- molekulární modely MeSH
- rekombinantní proteiny biosyntéza MeSH
- replikace viru účinky léků MeSH
- reprodukovatelnost výsledků MeSH
- rychlé screeningové testy MeSH
- termodynamika MeSH
- viabilita buněk účinky léků MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
OBJECTIVE AND DESIGN: Tenofovir (TFV) is used in pregnant women as a part of combination antiretroviral treatment to prevent mother-to-child transmission of HIV infection. We aimed to detect whether TFV and/or its prodrug, tenofovir disoproxil fumarate (TDF), are substrates of ATP-binding cassette (ABC) transporters that are functionally expressed in the placenta, namely P-glycoprotein (ABCB1/MDR1), Breast Cancer Resistance Protein (ABCG2/BCRP) and Multidrug Resistance-Associated Protein 2 (ABCC2/MRP2). We employed in-vitro cell-based assays and in-situ animal model to assess possible role of the efflux transporters in transplacental pharmacokinetics of TFV and TDF. METHODS: In-vitro transport assays were performed in MDCKII cells transduced with human ABCB1, ABCG2 or ABCC2. To quantify the effect of these transporters on TFV/TDF transplacental passage, we employed the in-situ model of dually perfused rat term placenta in open and closed setup. RESULTS: In-vitro assays revealed that TDF is a dual substrate of ABCB1 and ABCG2 but not of ABCC2. In contrast, TFV transport was not influenced by any of these transporters. Applying concentration-dependent studies and selective inhibitors, we further confirmed these findings in situ on the organ level; both ABCB1 and ABCG2 limited mother-to-fetus transfer of TDF whereas TFV transplacental passage was not affected by these ABC transporters. CONCLUSION: We propose limited mother-to-fetus transport of both TFV and TDF. While placental transport of TFV is restricted passively, by physical-chemical properties of the molecule, mother-to-fetus passage of TDF is actively hindered by placental ABCB1 and ABCG2 transporters, pumping this compound from trophoblast back to maternal circulation.
- MeSH
- ABC transportéry metabolismus MeSH
- adenin analogy a deriváty metabolismus farmakokinetika MeSH
- buněčné linie MeSH
- krysa rodu rattus MeSH
- látky proti HIV metabolismus farmakokinetika MeSH
- lidé MeSH
- nádorové proteiny metabolismus MeSH
- organofosfonáty metabolismus farmakokinetika MeSH
- P-glykoprotein metabolismus MeSH
- potkani Wistar MeSH
- proteiny spojené s mnohočetnou rezistencí k lékům metabolismus MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
