Browsing by Subject "inhibitors"

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Kinetics of the Viral Cycle Influence Pharmacodynamics of Antiretroviral Therapy
(2011-09) Sedaghat, Ahmad R.; Wilke, Claus O.; Wilke, Claus O.
Background: More and more antiretroviral therapies are being developed for treatment of HIV infection. The in-vivo efficacy of these drugs is commonly predicted based on in-vitro measures of antiviral effect. One primary invitro measure is the IC50, the amount of drug required for 50% inhibition of viral replication. We have previously shown that HIV life-cycle kinetics impact clinically observed HIV viral dynamics. Here we present a mathematical model of how they affect the pharmacodynamics of antiretroviral drugs. Results: We find that experimentally measured antiretroviral IC50s are determined by three factors: (i) intrinsic drug properties (e. g. drug-target binding), (ii) kinetics of the HIV life cycle, and (iii) kinetics of drug-inhibited infected cells. Our model predicts that the IC50 is a declining function of the duration of the drug-susceptible stage in the host cell. We combine our model with known viral life-cycle kinetics to derive a measure of intrinsic properties, reflecting drug action, for known antiretroviral drugs from previously measured IC50s. We show that this measure of intrinsic drug property correlates very well with in vitro-measured antiviral activity, whereas experimentally measured IC50 does not. Conclusions: Our results have implications for understanding pharmacodynamics of and improving activity of antiretroviral drugs. Our findings predict that drug activity can be improved through co-administration of synergistic drugs that delay the viral life cycle but are not inhibitory by themselves. Moreover, our results may easily extend to treatment of other pathogens.
Synthesis of 2-Substituted 9-Oxa-Guanines {5-Aminooxazolo 5,4-D Pyrimidin-7(6H)-Ones} and 9-Oxa-2-Thio-Xanthines{5-Mercaptooxazolo 5,4-D Pyrimidin-7(6H)-Ones}
(2008-07) Mandal, Subrata; Li, Wen Tai; Bai, Yan; Robertus, Jon D.; Kerwin, Sean M.; Mandal, Subrata; Li, Wen Tai; Bai, Yan; Robertus, Jon D.; Kerwin, Sean M.
Oxazolo[5,4-d] pyrimidines can be considered as 9-oxa-purine analogs of naturally occurring nucleic acid bases. Interest in this ring system has increased due to recent reports of biologically active derivatives. In particular, 5-aminooxazolo[5,4-d]pyrimidine-7(6H)-ones (9-oxa-guanines) have been shown to inhibit ricin. The preparation of a series of 2-substituted 5-aminooxazolo[5,4-d] pyrimidin-7(6H)-ones and related 5-thio-oxazolo[5,4-d] pyrimidines is described, including analogs suitable for further elaboration employing "click" chemistry utilizing copper-catalyzed Huisgen 1,3-dipolar cycloadditions. Two of the compounds prepared were found to inhibit ricin with IC(50) ca. 1-3 mM.
Virtual drug screening of potential and novel inhibitors of Escherichia coli dihydrofolate reductase
(2012) Nguyen, Adam Van; Robertus, Jon D.
Dihydrofolate Reductase (DHFR) is a major drug target because it is an enzyme that acts in a reaction mechanism to help produce tetrahydrofolic acid, which is then used by all cells to synthesize DNA precursors. The activity of this enzyme has also been implicated in the functions of cancerous cells, as they tend to over-replicate and thus require the enzyme more. Virtual Drug Screening is a relatively cheap and efficient method of screening through large libraries of compounds to find ones that have the highest binding affinities to the target protein/enzyme of interest. Specifically in this project, the virtual screening of ligands that can bind and possibly inhibit ecDHFR (Esherichia coli dihydrofolate reductase) was performed. E. Coli provides a target for developing antibiotic drugs since it can be responsible for certain illnesses caused in humans. At the same time, it is particularly difficult to develop antibacterial to E. Coli due to the sheer number of strains present. The actual ecDHFR enzyme was cloned into a pNIC-Bsa4 expression vector, expressed in BL21(DE3) cells, purified using Ni-NTA affinity chromatography, and characterized on a PAGE-gel. Concurrent with the wet lab, virtual screening of over 400,000 ligands also took place. A non-inhibitor enzyme assay was performed to validate the activity of the grown-up ecDHFR enzyme. Further testing in the wet-lab of some of the top-scoring ligands from the virtual screening (NRB00358 from Maybridge/Ryan Scientific; 5282931, 7722615, and 6407567 from cb-306_3d) via spectrophotometric enzyme inhibition assays showed no inhibition of ecDHFR activity. This shows that virtual screening is an imperfect measure of the ligand’s binding affinity, and may not necessarily predict its inhibitory qualities on the target protein. Even as such, it is hoped that further analysis of other top ligands from the virtual screening results will yield potential hits against this enzyme.