Fortunatus Chidolue Ezebuo

Lassa fever (LF), a viral hemorrhagic fever disease with a case fatality rate that can be over 20% among hospitalized LF patients, is endemic to many West African countries. Currently, no vaccines or therapies are specifically licensed to prevent or treat LF, hence the significance of developing therapeutics against the mammarenavirus Lassa virus (LASV), the causative agent of LF. We used in silico docking approaches to investigate the binding affinities of 2015 existing drugs to LASV proteins known to play critical roles in the formation and activity of the virus ribonucleoprotein complex (vRNP) responsible for directing replication and transcription of the viral genome. Validation of docking protocols were achieved with reference inhibitors of the respective targets. Our in silico docking screen identified five drugs (dexamethasone, tadalafil, mefloquine, ergocalciferol, and flunarizine) with strong predicted binding affinity to LASV proteins involved in the formation of the vRNP. We used cell-based functional assays to evaluate the antiviral activity of the five selected drugs. We found that flunarizine, a calcium-entry blocker, inhibited the vRNP activity of LASV and LCMV and virus surface glycoprotein fusion activity required for mammarenavirus cell entry. Consistently with these findings, flunarizine significantly reduced peak titers of LCMV in a multi-step growth kinetics assay in human A549 cells. Flunarizine is being used in several countries worldwide to treat vertigo and migraine, supporting the interest in exploring its repurposing as a candidate drug to treat LASV infections.

Lassa fever (LF), a viral hemorrhagic fever disease with a case fatality rate that can be over 20% among hospitalized LF patients, is endemic to many West African countries. Currently, no vaccines or therapies are specifically licensed to prevent or treat LF. Hence, the significance of developing therapeutics against the mammarenavirus Lassa virus (LASV), the causative agent of LF. We used in silico docking approaches to investigate the binding affinities of 2015 existing drugs to LASV proteins known to play critical roles in the formation and activity of the virus ribonucleoprotein complex (vRNP) responsible for directing replication and transcription of the viral genome. Validation of docking protocols were achieved with reference inhibitors of the respective targets. Our in silico docking screen identified five drugs (dexamethasone, tadalafil, mefloquine, ergocalciferol, and flunarizine) with strong predicted binding affinity to LASV proteins involved in the formation of the vRNP. We used cell-based functional assays to evaluate the antiviral activity of the five selected drugs. We found that flunarizine, a calcium-entry blocker, inhibited the vRNP activity of LASV and LCMV, and virus surface glycoprotein fusion activity required for mammarenavirus cell entry. Consistent with these findings, flunarizine significantly reduced peak titers of LCMV in a multi-step growth kinetics assay in human A549 cells. Flunarizine is being used in several countries worldwide to treat vertigo and migraine, supporting the interest in exploring its repurposing as a candidate drug to treat LASV infections.

Zika virus (ZIKV) infection is one of the mosquito-borne flaviviruses of human importance with more than 2 million suspected cases and more than 1 million people infected in about 30 countries. There are reported inhibitors of the zika virus replication machinery, but no approved effective antiviral therapy including vaccines directed against the virus for treatment or prevention is currently available. The study investigated the chemoinformatic design and profiling of derivatives of dasabuvir, efavirenz, and tipranavir as potential inhibitors of the zika virus RNA-dependent RNA polymerase (RdRP) and/or methyltransferase (MTase). The three-dimensional (3D) coordinates of dasabuvir, efavirenz, and tipranavir were obtained from the PubChem database, and their respective derivatives were designed with DataWarrior-5.2.1 using an evolutionary algorithm. Derivatives that were not mutagenic, tumorigenic, or irritant were selected; docked into RdRP and MTase; and further subjected to absorption, distribution, metabolism, excretion, and toxicity (ADMET) evaluation with Swiss-ADME and pkCSM web tools. Some of the designed compounds are Lipinski's rule-of-five compliant, with good synthetic accessibilities. Compounds 20d, 21d, 22d, and 1e are nontoxic with the only limitation of CYP1A2, CYP2C19, and/or CYP2C9 inhibition. Replacements of -CH₃ and -NH- in the methanesulfonamide moiety of dasabuvir with -OH and -CH₂- or -CH₂CH₂-, respectively, improved the safety/toxicity profile. Hepatotoxicity in 5d, 4d, and 18d is likely due to -NH- in their methanesulfonamide/sulfamic acid moieties. These compounds are potent inhibitors of N-7 and 2'-methylation activities of ZIKV methyltransferase and/or RNA synthesis through interactions with amino acid residues in the priming loop/"N-pocket" in the virus RdRP. Synthesis of these compounds and wet laboratory validation against ZIKV are recommended.

Schistosomiasis is a neglected disease of considerable health importance in tropical and subtropical regions. Its treatment relies on the use of praziquantel or oxamniquine but there are reported cases of treatment failures due to resistance or tolerance. Again, derivatives of praziquantel and oxamniquine have not shown significant activities than their parent compounds. The study predicted approved drugs with possible antischistosomal activities. Four schistosomal drug targets were obtained from Protein Data Bank and six hundred and twelve (612) approved drugs including their isomers were selected based on their Molinspiration® bioscore similarities with reference compounds (praziquantel, oxamniquine, [(2S,3S,4S,5S,6S)-3,4,5-triacetyloxy-6-sulfanyloxan-2-yl] methyl acetate, [propylamino-3-hydroxy-buta-1,4-dionyl]-isoleucylproline). The selected drugs and drug targets were obtained and prepared for molecular docking simulations. The molecular docking simulations were performed using AutoDockvina^®-1.1.2 after validation of docking protocols while molecular dynamics simulations were performed with GROMACS-4.5.5. The binding energies were calculated using MMPBSA (Molecular Mechanics Poisson-Boltzmann Surface Area). Tolmetin was predicted as potential antischistosomal drug with binding energies of −231.064 ± 18.550 and −338.636 ± 36.900 KJ/mol for sulfotransferase and thioredoxin glutathione reductase (TGR) respectively. Also diflunisal was predicted as potential antischistosomal drug with binding energies of −168.641 ± 20.370 and −290.117 ± 43.800 KJ/mol for sulfotransferase and TGR respectively. Non-covalent interactions and conformational changes were responsible for molecular recognitions and specificities and average bond measurement showed that carboxylic functional groups in diflunisal and tolmetin may interact covalently with -SH group of Cys159 in TGR. Confirmation of covalent interactions and in vitro validations are recommended. Communicated by Ramaswamy H. Sarma

Background: Sulfotransferase family comprises key enzymes involved in drug metabolism. Oxamniquine is a pro-drug converted into its active form by schistosomal sulfotransferase. The conformational dynamics of side-chain amino acid residues at the binding site of schistosomal sulfotransferase towards activation of oxamniquine has not received attention.Objective: The study investigated the conformational dynamics of binding site residues in free and oxamniquine bound schistosomal sulfotransferase systems and their contribution to the mechanism of oxamniquine activation by schistosomal sulfotransferase using molecular dynamics simulations and binding energy calculations.Methods: Schistosomal sulfotransferase was obtained from Protein Data Bank and both the free and oxamniquine bound forms were subjected to molecular dynamics simulations using GROMACS-4.5.5 after modeling it’s missing amino acid residues with SWISS-MODEL. Amino acid residues at its binding site for oxamniquine was determined and used for Principal Component Analysis and calculations of side-chain dihedrals. In addition, binding energy of the oxamniquine bound system was calculated using g_MMPBSA.Results: The results showed that binding site amino acid residues in free and oxamniquine bound sulfotransferase sampled different conformational space involving several rotameric states. Importantly, Phe45, Ile145 and Leu241 generated newly induced conformations, whereas Phe41 exhibited shift in equilibrium of its conformational distribution. In addition, the result showed binding energy of -130.091 ± 8.800 KJ/mol and Phe45 contributed -9.8576 KJ/mol.Conclusion: The results showed that schistosomal sulfotransferase binds oxamniquine by relying on hybrid mechanism of induced fit and conformational selection models. The findings offer new insight into sulfotransferase engineering and design of new drugs that target sulfotransferase.

Arthropod-borne flaviviruses such as tick-borne encephalitis virus (TBEV), West Nile virus (WNV), Zika virus (ZIKV), Dengue virus (DENV), and yellow fever virus (YFV) cause several serious life-threatening syndromes (encephalitis, miscarriages, paralysis, etc.). No effective antiviral therapy against these viruses has been approved yet. We selected, via in silico modeling, 12 U.S. Food and Drug Administration (FDA)-approved antiviral drugs (paritaprevir, dolutegravir, raltegravir, efavirenz, elvitegravir, tipranavir, saquinavir, dasabuvir, delavirdine, maraviroc, trifluridine, and tauroursodeoxycholic acid) for their interaction with ZIKV proteins (NS3 helicase and protease, non-structural protein 5 (NS5) RNA-dependent RNA polymerase, and methyltransferase). Only three of them were active against ZIKV, namely, dasabuvir (ABT-333), efavirenz, and tipranavir. These compounds inhibit virus replication of ZIKV (MR-766 and Paraiba_01) in Vero cells; therefore, we tested these compounds against other medically important flaviviruses WNV (13-104 and Eg101) and TBEV (Hypr). Dasabuvir was originally developed as an antiviral drug against hepatitis C virus (HCV); tipranavir and efavirenz are used for treating human immunodeficiency virus (HIV) infection. The antiviral effects of efavirenz, tipranavir, and dasabuvir were tested for ZIKV in HUH-7, astrocytes (HBCA), and UKF-NB-4 cells, where we also identified a significant inhibition effect of these compounds. For Vero cells, efavirenz inhibited all investigated viruses with EC₅₀ ranging from 9.70 to 29.26 µM; the tipranavir inhibition effect was from 16.19 (WNV 13-104) to 27.47 µM (TBEV), while the strongest and most robust antiviral effect was demonstrated in the case of dasabuvir (EC₅₀ values ranging from 9.09 (TBEV) to 10.85 µM (WNV 13-104)). These results warrant further research of these drugs, either individually or in combination, as possible pan-flavivirus inhibitors.

Vector-borne flaviviruses (VBFs) affect human health worldwide, but no approved drugs are available specifically to treat VBF-associated infections. Here, we performed in silico screening of a library of U.S. Food and Drug Administration-approved antiviral drugs for their interaction with Zika virus proteins. Twelve hit drugs were identified by the docking experiments and tested in cell-based antiviral assay systems. Efavirenz, tipranavir, and dasabuvir at micromolar concentrations were identified to inhibit all VBFs tested; i.e., two representatives of mosquito-borne flaviviruses (Zika and West Nile viruses) and one representative of flaviviruses transmitted by ticks (tick-borne encephalitis virus). The results warrant further research into these drugs, either individually or in combination, as possible pan-flavivirus inhibitors.

Cancer is a difficult disease to treat, and few effective drugs are available. Hence, it is of great importance to develop effective anti-cancer therapeutic agents with well-defined pharmacokinetic properties. Although, ginger (Zingiber officinale) has a number of proven pharmacological activities, its effect on colon cancer has not received much attention. This study therefore investigated the potential colon cancer drug of compounds found in ginger. Dihydropyrimidine dehydrogenase was modeled using comparative homology modeling and virtual screening was performed locally on a Linux platform using AutoDock Vina®. The results showed that human dihydropyrimidine dehydrogenase is a homolog of pig dihydropyrimidine dehydrogenase. The leads of potential colon cancer drugs were beta-sitosterol, 6-Shogoal, Alloaromadedrene, and Zingiberol. They had similar binding site with levamisole for tumor necrosis factor ligand superfamily member 6 with His 148 and Tyr 192 common at their binding site whereas they had different binding sites from 5-fluorouracil for dihydropyrimidine dehydrogenase. The leads had better bioactivities compared with reference drugs (5-flourouracil and Levamisole) approved clinically for the treatment of colon cancer. In vitro, ex vivo and/or in vivo validations of the leads against colon cancer are recommended.

Lipoxygenases comprise a family of non-heme iron-containing enzymes that catalyze the stereospecific dioxygenation of polyunsaturated fatty acids with 1, 4-cis-cis-pentadiene structure. Hemoglobin, a heme iron-containing protein has been reported to have lipoxygenase activity but the assay conditions that could enhance the activity remain obscure. Therefore, establishment of optimum assay conditions for lipoxygenase activity of hemoglobin could allow modeling of hemoglobin as lipoxygenase. Hemoglobin was extracted from blood of an identified individual of genotype AA. The hemoglobin was dialyzed at 4 degrees C for 24 h against 50 mM Tris-HCl buffers (pH 8.5 and 7.2) and effects of sodium dodecyl sulphate (SDS) and linoleic studied at pH 5.0 and 7.2 with UV-VIS Titration Spectrophotometry. The results show that 3.3, 8.6 and 88.1% concentrations of met-hemoglobin were found in presence of 0.0 mM SDS at pH 5.0 and 7.2, 1.043 mM SDS at pH 7.2 and 0.404 mM SDS at pH 5.0 respectively. Also, the difference spectra of hemoglobin in presence of linoleic acid showed positive peak at 285 nm which suggest the presence of oxodienes--a reaction product of hydroperoxidase activity of lipoxygenase. Formation of met-hemoglobin/met-myoglobin is highly correlated with lipid oxidation. Since highest concentration of met-hemoglobin (88.1%) was observed in presence of 0.404 mM SDS at pH 5.0, lipoxygenase activity of hemoglobin was enhanced in presence of SDS under these conditions.