Nucleos(t)ide analogues entecavir (ETV) and tenofovir disoproxil fumarate (TDF) are recommended as first-line monotherapies for chronic hepatitis B (CHB). Multiple HBV genotypes/subgenotypes have been described, but their impact on treatment response remains largely elusive. We investigated the effectiveness of ETV/TDF on HBV/D-subgenotypes, D1/D2/D3/D5, studied the structural/functional differences in subgenotype-specific reverse transcriptase (RT) domains of viral polymerase, and identified novel molecules with robust inhibitory activity on various D-subgenotypes. Transfection of Huh7 cells with full-length D1/D2/D3/D5 and in vitro TDF/ETV susceptibility assays demonstrated that D1/D2 had greater susceptibility to TDF/ETV while D3/D5 exhibited poorer response. Additionally, HBV load was substantially reduced in TDF-treated CHB patients carrying D1/D2 but minimally reduced in D3/D5-infected patients. Comparison of RT sequences of D-subgenotypes led to identification of unique subgenotype-specific residues, and molecular modeling/docking/simulation studies depicted differential bindings of TDF/ETV to the active site of their respective RTs. Replacement of signature residues in D3/D5 HBV clones with corresponding amino acids seen in D1/D2 improved their susceptibility to TDF/ETV. Using high throughput virtual screening, we identified N(9)-[3-fluoro-2-(phosphonomethoxy)propyl] (FPMP) derivatives of purine bases, including N6-substituted (S)-FPMP derivative of 2,6-diaminopurine (DAP) (OB-123-VK), as potential binders of RT of different D-subgenotypes. We synthesized (S)-FPMPG prodrugs (FK-381-FEE/FK-381-SEE/FK-382) and tested their effectiveness along with OB-123-VK. Both OB-123-VK and FK-381-FEE exerted similar antiviral activities against all D-subgenotypes, although FK-381-FEE was more potent. Our study highlighted the natural variation in therapeutic response of D1/D2/D3/D5 and emphasized the need for HBV subgenotype determination before treatment. Novel molecules described here could benefit future design/discovery of pan-D-subgenotypic inhibitors. IMPORTANCE Current treatment of chronic hepatitis B relies heavily on nucleotide/nucleoside analogs in particular, tenofovir disoproxil fumarate (TDF) and entecavir (ETV) to keep HBV replication under control and prevent end-stage liver diseases. However, it was unclear whether the therapeutic effects of TDF/ETV differ among patients infected with different HBV genotypes and subgenotypes. HBV genotype D is the most widespread of all HBV genotypes and multiple D-subgenotypes have been described. We here report that different subgenotypes of HBV genotype-D exhibit variable response toward TDF and ETV and this could be attributed to naturally occurring amino acid changes in the reverse transcriptase domain of the subgenotype-specific polymerase. Further, we identified novel molecules and also synthesized prodrugs that are equally effective on different D-subgenotypes and could facilitate management of HBV/D-infected patients irrespective of D-subgenotype.
- MeSH
- antivirové látky chemie farmakologie terapeutické užití MeSH
- chronická hepatitida B farmakoterapie virologie MeSH
- genotyp MeSH
- guanin analogy a deriváty chemie farmakologie terapeutické užití MeSH
- inhibitory reverzní transkriptasy chemie farmakologie terapeutické užití MeSH
- lidé MeSH
- mutace MeSH
- organofosfonáty chemie farmakologie MeSH
- prekurzory léčiv MeSH
- proteinové domény MeSH
- racionální návrh léčiv * MeSH
- reverzní transkriptasa chemie genetika MeSH
- tenofovir chemie farmakologie terapeutické užití MeSH
- virová léková rezistence účinky léků genetika MeSH
- virová nálož účinky léků MeSH
- virus hepatitidy B účinky léků enzymologie genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
To elucidate the structure-geometry-activity relationship in diarylpyrimidine family (DAPYs) containing carbonyl linker between the central pyrimidine core and phenyl type B-arm, a series of (2,6-difluorophenyl)(2-(phenylamino)pyrimidin-4-yl)methanones was designed, prepared and tested for their anti-HIV-1 activity. The carbonyl linker bearing B phenyl arm was successfully attached at both C-2 and C-4 positions of the central pyrimidine ring using a new synthetic approach. Further modifications of target compounds are present at C-5 position of the pyrimidine ring. In vitro anti-HIV-1 activity study performed on a series of 22 compounds confirmed the crucial importance of both conformational rigidity between phenyl B arm and the pyrimidine core linked through the carbonyl bridge, as well as presence of fluoro substituents in ortho-positions of phenyl B moiety. The most potent derivative of the series, compound 17, having almost perpendicular angle within the two planes made from the B aromatic arm and the pyrimidine ring, exhibited low nanomolar anti-HIV-1 activity (EC50 = 4 nM) with no significant toxicity (CC50 > 57.1 μM).
- MeSH
- HIV reverzní transkriptasa antagonisté a inhibitory chemie metabolismus MeSH
- HIV-1 účinky léků enzymologie MeSH
- inhibitory reverzní transkriptasy chemie metabolismus farmakologie MeSH
- konformace proteinů MeSH
- pyrimidiny chemie metabolismus farmakologie MeSH
- racionální návrh léčiv * MeSH
- simulace molekulového dockingu MeSH
- Publikační typ
- časopisecké články MeSH
Heterocyclic compounds execute a very important role in drug design and discovery. This article provides the basic milestones of the research for pyrroloaryl and pyrroloheteroaryl based components targeting HIV viral replication cycle. Anti-HIV activity is elaborated for several classes of pyrrolo-compounds as pyrrolopyridines, pyrrolopyrimidines, pyrrolopyridazines, pyrrolobenzodiazepinones, pyrrolobenzothiazepines, pyrrolobenzoxazepinones, pyrrolophenanthridines, pyrroloquinoxalines, pyrrolotriazines, pyrroloquinolines, pyrrolopyrazinones, pyrrolothiatriazines, arylthiopyrroles and pyrrolopyrazolones targeting two essential HIV enzymes, reverse transcriptase and integrase as well as attachment/fusion of HIV virons to the host CD-4 cell. Such attempts were resulted in a discovery of highly potent anti-HIV agents suitable for clinical trials, for example, BMS-378806, BMS-585248, BMS-626529, BMS-663068, BMS-488043 and BMS-663749, etc. as anti-HIV attachment agents, triciribine, QX432, BI-1 and BI-2 as HIV RT inhibitors which are in preclinical or clinical development. Mechanism of action of compounds presented in this article towards the suppression of HIV attachment/fusion as well as against the activities of HIV enzymes reverse transcriptase and integrase has been discussed. Relationships of new compounds' molecular framework and HIV viral target has been overviewed in order to facilitate further construction of promising anti-HIV agents in future drug discovery process.
- MeSH
- HIV infekce farmakoterapie virologie MeSH
- HIV-integrasa metabolismus MeSH
- HIV účinky léků enzymologie fyziologie MeSH
- inhibitory HIV fúze chemie farmakologie MeSH
- inhibitory HIV-integrasy chemie farmakologie MeSH
- inhibitory reverzní transkriptasy chemie farmakologie MeSH
- látky proti HIV chemie farmakologie MeSH
- lidé MeSH
- objevování léků MeSH
- pyrroly chemie farmakologie MeSH
- replikace viru účinky léků MeSH
- reverzní transkriptasa metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
While the selection of amino acid insertions in human immunodeficiency virus (HIV) reverse transcriptase (RT) is a known mechanism of resistance against RT inhibitors, very few reports on the selection of insertions in the protease (PR) coding region have been published. It is still unclear whether these insertions impact protease inhibitor (PI) resistance and/or viral replication capacity. We show that the prevalence of insertions, especially between amino acids 30 to 41 of HIV type 1 (HIV-1) PR, has increased in recent years. We identified amino acid insertions at positions 33 and 35 of the PR of HIV-1-infected patients who had undergone prolonged treatment with PIs, and we characterized the contribution of these insertions to viral resistance. We prepared the corresponding mutated, recombinant PR variants with or without insertions at positions 33 and 35 and characterized them in terms of enzyme kinetics and crystal structures. We also engineered the corresponding recombinant viruses and analyzed the PR susceptibility and replication capacity by recombinant virus assay. Both in vitro methods confirmed that the amino acid insertions at positions 33 and 35 contribute to the viral resistance to most of the tested PIs. The structural analysis revealed local structural rearrangements in the flap region and in the substrate binding pockets. The enlargement of the PR substrate binding site together with impaired flap dynamics could account for the weaker inhibitor binding by the insertion mutants. Amino acid insertions in the vicinity of the binding cleft therefore represent a novel mechanism of HIV resistance development.
- MeSH
- buněčné linie MeSH
- chemické modely MeSH
- difrakce rentgenového záření MeSH
- financování organizované MeSH
- HIV-1 enzymologie fyziologie genetika MeSH
- HIV-proteasa genetika chemie izolace a purifikace metabolismus MeSH
- inhibitory reverzní transkriptasy chemie MeSH
- inzerční mutageneze MeSH
- katalýza MeSH
- kinetika MeSH
- konsenzuální sekvence MeSH
- látky proti HIV terapeutické užití MeSH
- ledviny cytologie MeSH
- lidé MeSH
- molekulární sekvence - údaje MeSH
- rekombinantní proteiny chemie izolace a purifikace metabolismus MeSH
- replikace viru MeSH
- RNA virová analýza MeSH
- sekvence aminokyselin MeSH
- vazba proteinů MeSH
- vazebná místa MeSH
- virová léková rezistence MeSH
- Check Tag
- lidé MeSH
