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. 2013 Dec;57(12):6110-21.
doi: 10.1128/AAC.01420-13. Epub 2013 Sep 30.

GRL-04810 and GRL-05010, difluoride-containing nonpeptidic HIV-1 protease inhibitors (PIs) that inhibit the replication of multi-PI-resistant HIV-1 in vitro and possess favorable lipophilicity that may allow blood-brain barrier penetration

Pedro Miguel Salcedo Gómez  1 , Masayuki Amano, Sofiya Yashchuk, Akira Mizuno, Debananda Das, Arun K Ghosh, Hiroaki Mitsuya
Affiliations

GRL-04810 and GRL-05010, difluoride-containing nonpeptidic HIV-1 protease inhibitors (PIs) that inhibit the replication of multi-PI-resistant HIV-1 in vitro and possess favorable lipophilicity that may allow blood-brain barrier penetration

Pedro Miguel Salcedo Gómez et al. Antimicrob Agents Chemother. 2013 Dec.

Abstract

We designed, synthesized, and identified two novel nonpeptidic human immunodeficiency virus type 1 (HIV-1) protease inhibitors (PIs), GRL-04810 and GRL-05010, containing the structure-based designed privileged cyclic ether-derived nonpeptide P2 ligand, bis-tetrahydrofuranylurethane (bis-THF), and a difluoride moiety, both of which are active against the laboratory strain HIV-1LAI (50% effective concentrations [EC50s], 0.0008 and 0.003 μM, respectively) with minimal cytotoxicity (50% cytotoxic concentrations [CC50s], 17.5 and 37.0 μM, respectively, in CD4(+) MT-2 cells). The two compounds were active against multi-PI-resistant clinical HIV-1 variants isolated from patients who had no response to various antiviral regimens. GRL-04810 and GRL-05010 also blocked the infectivity and replication of each of the HIV-1NL4-3 variants selected by up to 5 μM lopinavir (EC50s, 0.03 and 0.03 μM, respectively) and atazanavir (EC50s, 0.02 and 0.04 μM, respectively). Moreover, they were active against darunavir (DRV)-resistant variants (EC50 in 0.03 to 0.034 μM range for GRL-04810 and 0.026 to 0.043 μM for GRL-05010), while DRV had EC50s between 0.02 and 0.174 μM. GRL-04810 had a favorable lipophilicity profile as determined with the partition (log P) and distribution (log D) coefficients of -0.14 and -0.29, respectively. The in vitro blood-brain barrier (BBB) permeability assay revealed that GRL-04810 and GRL-05010 may have a greater advantage in terms of crossing the BBB than the currently available PIs, with apparent penetration indexes of 47.8 × 10(-6) and 61.8 × 10(-6) cm/s, respectively. The present data demonstrate that GRL-04810 and GRL-05010 exert efficient activity against a wide spectrum of HIV-1 variants in vitro and suggest that two fluorine atoms added to their bis-THF moieties may well enhance their penetration across the BBB.

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Figures

Fig 1
Fig 1
Structures of GRL-04810, GRL-05010, and darunavir. MW, molecular weight.
Fig 2
Fig 2
Selection of in vitro variants resistant to GRL-04810 and GRL-05010. MT-4 cells were exposed to the wild-type strain HIV-1NL4-3 and cultured in the presence of increasing concentrations of GRL-04810, GRL-05010, DRV, and LPV. Culture supernatants were collected weekly, and if the amounts of p24 Gag were found to surpass the cutoff value of 200 ng/ml, those supernatants were used for the subsequent passage. Each passage was conducted in a cell-free manner.
Fig 3
Fig 3
Amino acid sequences of the protease-encoding region of HIV-1 variants selected in the presence of GRL-04810 and GRL-05010. Shown are the amino acid sequences deduced from the nucleotide sequences of the protease-encoding region of proviral DNA isolated from HIV-1NL4-3 variants selected in the presence of GRL-04810, GRL-05010, and DRV at passages 20, 15, and 20, respectively. Identified amino acid substitutions in the protease-encoding region and their frequencies (far-right column) are shown. The amino acid sequence of the wild-type HIV-1NL4-3 protease (pNL4-3 PR) is shown at the top as a reference.
Fig 4
Fig 4
Replication kinetics of GRL-04810- and GRL-05010-resistant HIV-1 variants generated in vitro. GRL-04810- and GRL-05010-resistant viruses were obtained from the selection assay illustrated in Fig. 2. Two time point passages, HIV-1GRL-04810RP18 and HIV-1GRL-05010RP10, were chosen. HIV-1NL4-3 or two resistant viruses were propagated in CD4+ MT-4 cells in the presence of 0.01 μM (▲) and 0.001 μM (■) GRL-04810 and GRL-05010 or in the absence (◆) of the compounds in culture flasks. The amounts of p24 in each culture flask were quantified every 2 days for 1 week. Sup, supernatant.
Fig 5
Fig 5
Antiviral activity of GRL-04810brain, GRL-05010brain, DRVbrain, AZTbrain, IDVbrain and SQVbrain against HIV-1LAI. Compounds recovered from the brain side of the medium in the BBB in vitro assay were challenged with HIV-1LAI, as indicated, in a p24 assay using PHA-PBMC. Dilution ranges were between 10-fold and 10,000-fold, and the percentage of viral inhibition was used as an endpoint. It is assumed that the greater the percentage of inhibition of HIV-1LAI, the greater the concentration of the drug in the brain side medium. Assays were conducted in duplicate, and error bars show standard deviations.
Fig 6
Fig 6
Antiviral activity of GRL-04810brain, GRL-05010brain, DRVbrain, AZTbrain, IDVbrain and SQVbrain against HIV-1ERS104pre. Compounds recovered from the brain side of the medium in the BBB in vitro assay were challenged with HIV-1ERS104pre, as indicated, in a p24 assay using PHA-PBMC. Dilution ranges were between 10-fold and 10,000-fold, and the percentage of viral inhibition was used as an endpoint. It is assumed that the greater the percentage of inhibition of HIV-1ERS104pre, the greater the concentration of the drug in the brain side medium. Assays were conducted in duplicate, and error bars show standard deviations.
Fig 7
Fig 7
Structural interactions of GRL-04810 and GRL-05010 with HIV-1 protease. A model of GRL-04810 bound to protease is shown (A). GRL-04810 is shown in thick sticks, and residues in the protease active site are shown in wire. The following atom colors are used: carbons in gray, oxygens in red, fluorines in green, nitrogens in blue, and sulfurs in yellow. The polar interactions (yellow dotted lines) of DRV, GRL-04810, and GRL-05010, along with other residues in the active site, are shown in panels B, C, and D, respectively. Nonpolar hydrogens are not shown. Figures were generated using Maestro, version 9.3. (Schrödinger, LLC, New York, NY).
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References

    1. Coiras M, Lopez-Huertas M, Perez-Olmeda M, Alcami J. 2009. Understanding HIV-1 latency provides clues for the eradication of long-term reservoirs. Nat. Rev. Microbiol. 7:798–812 - PubMed
    1. De Clercq. 2002. New anti-HIV agents and targets. Med. Res. Rev. 22:531–565 - PubMed
    1. Gorantla S, Poluektova L, Gendelman HE. 2012. Rodent models for HIV-associated neurocognitive disorders. Trends Neurosci. 35:197–208 - PMC - PubMed
    1. Lewin SR, Murray J, Solomon A, Wightman F, Cameron PU, Purcell DJ, Zaunders JJ, Grey P, Bloch M, Smith D, Cooper DA, Kelleher AD. 2008. Virological determinants of success after structured treatment interruptions of antiretrovirals in acute HIV-1 infection. J. Acquir. Immune Defic. Syndr. 47:140–147 - PubMed
    1. Siliciano JD, Siliciano RF. 2004. A long-term latent reservoir for HIV-1: discovery and clinical implications. J. Antimicrob. Chemother. 54:6–9 - PubMed

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