Evaluation of Potential Anti-Hepatitis A Virus 3C Protease Inhibitors Using Molecular Docking

Hepatitis A virus (HAV) infection is a major cause of acute hepatitis worldwide and occasionally causes acute liver failure and can lead to death in the absence of liver transplantation. Although HAV vaccination is available, the prevalence of HAV vaccination is not adequate in some countries. Additionally, the improvements in public health reduced our immunity to HAV infection. These situations motivated us to develop potentially new anti-HAV therapeutic options. We carried out the in silico screening of anti-HAV compounds targeting the 3C protease enzyme using the Schrodinger Modeling software from the antiviral library of 25,000 compounds to evaluate anti-HAV 3C protease inhibitors. Additionally, in vitro studies were introduced to examine the inhibitory effects of HAV subgenomic replicon replication and HAV HA11-1299 genotype IIIA replication in hepatoma cell lines using luciferase assays and real-time RT-PCR. In silico studies enabled us to identify five lead candidates with optimal binding interactions in the active site of the target HAV 3C protease using the Schrodinger Glide program. In vitro studies substantiated our hypothesis from in silico findings. One of our lead compounds, Z10325150, showed 47% inhibitory effects on HAV genotype IB subgenomic replicon replication and 36% inhibitory effects on HAV genotype IIIA HA11-1299 replication in human hepatoma cell lines, with no cytotoxic effects at concentrations of 100 μg/mL. The effects of the combination therapy of Z10325150 and RNA-dependent RNA polymerase inhibitor, favipiravir on HAV genotype IB HM175 subgenomic replicon replication and HAV genotype IIIA HA11-1299 replication showed 64% and 48% inhibitory effects of HAV subgenomic replicon and HAV replication, respectively. We identified the HAV 3C protease inhibitor Z10325150 through in silico screening and confirmed the HAV replication inhibitory activity in human hepatocytes. Z10325150 may offer the potential for a useful HAV inhibitor in severe hepatitis A.     

Temperature and orientation dependence of the deformation and fracture in boron-doped Ni76Al19Ti5 single crystals

Tensile tests were performed on a boron-doped Ni₇₆Al₁₉Ti₅ single crystal as a function of temperature and orientation to investigate the effect of boron additions on the deformation and fracture behavior of single crystalline Ni₃Al-based intermetallic compounds. By boron addition, the peak temperature, T_p, where the stress reaches a maximum, was shifted to a higher temperature, except for the [001] orientation, which was interpreted by the increased stress level required for operation of the primary cube slip systems. In contrast, the temperature dependence of slip line morphology and elongation below and aboveT _p was similar for boron-doped and undoped crystals at each orientation. By boron addition, the fracture belowT _p occurred predominantly on {111} slip planes ({111} cracking) and noncrystallographic planes, depending on orientation with a suppression of {001} cleavage fracture. AboveT _p , an increment in elongation was found as a result of the formation of kink bands ([113] orientation) and restraint of {111} cracking ([111] orientation) in a similar way to the case of boron-free crystal. Formerly Graduate Student, Tohoku University.     

A calmodulin-target peptide hybrid molecule with unique calcium-binding properties

This paper describes the production and properties of a hybridprotein comprising the full length of the Xenopus laevis cabnodulin(CaM) sequence, followed, through a gh/cylgh/dne linker, bythe 26-residue CaM-binding region of myosin light-chain kinase(M13). This hybrid molecule appears to have high thermal stability(T_m > 75°C in the presence of Ca²⁺) as well as unusualCa²⁺-binding properties: (i) a wide-range biphasic Ca²⁺-bindingresponse (extending over pCa 4.8-7.4) and (ii) a high apparentbinding constant (pCa_50% = 6.3, a 10-fold increase from thatof wild-type CaM). NMR and CD data indicate that the CaM-M13hybrid molecule exists in equilibrium in an approximate 1:1ratio between two major conformations, one of which is similarto the compact globular structure of the CaM-M13 complex [M.Daira,G.M.Clore, A.M.Gronenborn, G.Zhu, C.B.Klee and A.Bax (1992)Science, 256, 632-638] and the other to the dumbbell-like structureof the wild type CaM [Y.S.Babu, C.E.Bugg and W.J.Cook (1988)J. Mol. Biol., 204, 191-204]. The biphasic Ca²⁺-binding curvecan be interpreted using a linear combination of two Hill bindingcurves with significantly different dissociation constants (2x 10^-6 M and 8 x 10^-6 M), which can be attributed to the twoconformations in equilibrium. The present study has opened anavenue to engineer proteins with higher Ca²⁺-binding affinitiesusing the known CaM structures as a template Received April 23, 1993; revised August 5, 1993; accepted August 25, 1993.     

Fusion variations of pancreatic ducts in patients with anomalous arrangement of pancreaticobiliary ductal system

Portal vein flow was recorded by color Doppler sonography in 31 patients with chronic heart failure and 18 control subjects. Compared with patients showing a forward flow (Group A), those with reversed portal vein flow (Group B) had higher prevalence of tricuspid regurgitation (75% vs. 43%), hepatic congestion (100% vs. 30%) and ascites (50% vs. 18%), and showed higher right atrial pressure (25.3 +/- 3.01 mmHg vs. 11.8 +/- 5.75 mmHg, p < 0.01). In controls, portal vein pulsatility ratio was 0.66 +/- 0.08, in Group A it was 0.46 +/- 0.28 (p < 0.01), in Group B -0.60 +/- 0.19 (p < 0.01). Portal vein pulsatility ratio negatively correlated with right atrial pressure (r = -0.87; p < 0.01). In Group A, hepatic congestion, ascites and tricuspid regurgitation were associated with a higher portal vein pulsatility. This study indicates that portal vein pulsatility ratio reflects the level of impairment of the right heart.     

Fusion of the MLL gene with two different genes, AF-6 and AF-5alpha, by a complex translocation involving chromosomes 5, 6, 8 and 11 in infant leukemia

We analysed a complex translocation involving chromosomes 5, 6, 8 and 11 in a case of infant leukemia. Molecular analysis of the MLL gene revealed that MLL was fused with two different genes, AF-6 on chromosome 6q27 and AF-5alpha. AF-5alpha, the 11th partner gene fused with MLL, is a novel gene mapped to chromosome 5q12, which encodes a 31 kDa protein of 269 amino acids and contains a possible nuclear targeting sequence, a potential leucine zipper dimerization motif and an alpha-helical coiled-coil domain. In situ hybridization and molecular cloning analyses demonstrated that two different types of chromosomal recombination had occurred in the cells. One was a three-way translocation among chromosomes 6, 8 and 11, and the other was an insertion of a chromosome 5-derived segment into the breakpoint of chromosomes 8 and 11. Accordingly, the karyotype was defined as del(5)(q11.2q12), der(6)t(6;8) (q27;q11.2), der(8)(8pter-->8q11.2::5q11.2-->5q12::11q23-->++ +11qter), der(11)t(6;11) (q27;q23). Thus, the MLL gene created two different fusion mRNAs, since the chromosome 11 split into two different chromosomes 5 and 6. This is the first report demonstrating fusion of the MLL gene with two different genes by a complex translocation.     

Masticatory function in patients with mandibular prognathism before and after orthognathic surgery

Masticatory function was analyzed in 54 patients with mandibular prognathism before or after orthognathic treatment and in 40 adults with normal occlusion. Masticatory efficiency was evaluated spectrophotometrically by measuring the amount of adenosine triphosphate eluted from masticated adenosine triphosphate granules. Occlusal contact was evaluated by calculating the number and area of occlusal contacts from the illuminated images of a silicone record of the occlusion that was stored in the image analysis system with the aid of a charge-coupled device camera. The mean masticatory efficiency of the preoperative group was approximately half that of the control group. The value for the postoperative group was slightly higher than that for the preoperative group, but there was no statistical difference between the two groups. Likewise, the number and area of occlusal contacts in the preoperative group were also approximately half or less as compared with those of the control group, and they did not show significant changes postoperatively. Statistically, there were correlations between the masticatory efficiency and the number and area of occlusal contacts. The results indicate that although the occlusal relationship of the upper and lower teeth were greatly improved by orthognathic treatment, the postoperative occlusion is not tight enough and may need further adjustment at the end of the treatment.     

Toughness and strength characteristics of Nb-W-Si ternary alloys prepared by Arc melting

This article describes the room-temperature and high-temperature mechanical properties and failure modes of series Nb-W-Si alloys—Nb-10W, Nb-10Si, Nb-10Si-5W, Nb-10W-5Si, and Nb-10W-10Si—prepared by arc melting. For the Nb-10W alloy, the microstructure was a monolithic Nb solid solution (Nb _ss ) with a grain size up to a few hundred microns, while the other four alloys consisted of primary Nb _ss and a eutectic of Nb _ss /Nb₅Si₃ (5-3 silicide) as a result of replacing Nb with Si. Among all alloys, the Nb-10W showed the highest fracture toughness of about 15.3 MPa√m^1/2 and the lowest 0.2 pct yield compressive strength of 90 MPa at 1670 K. Conversely, the Nb-10Si-10W had the highest 0.2 pct yield strength of about 330 MPa at 1670 K and the lowest fracture toughness of 8.2 MPa√m^1/2. It is suggested that toughness is supplied by the metallic Nb _ss phase, while high-temperature strength is mainly provided by the brittle silicide phase. For the Nb-10W alloy with the monolithic Nb _ss , intergranular cleavagelike crack propagation is the fracture mode at room temperature, and dislocation movement within the grains and grain-boundary sliding are the dominant modes of high-temperature failure. With two-phase Nb _ss /Nb₅Si₃ microstructures, the compressive damage of all four alloys at high temperature was dominated by debonding of the interfaces between the Nb _ss and the silicide; however, the fracture mode at room temperature is transgranular, controlled by the primary Nb _ss cleavage.     

Preparation of (vinylbenzylimino)oligo(dl-phenylalanine NCA) macromer with narrow molecular weight distribution

Summary Polymerization conditions of dl-phenylalanine N-carboxyanhydride (NCA) by m, p-vinylbenzylamine were examined for the preparation of (vinylbenzylimino)oligo(dl-phenylanaline NCA) macromer (VB-OPhe) with narrow molecular weight distribution. According to the results obtained, i.e., under the condition of short polymerization time (ca. 2 h) and in relatively high concentrations of the NCA in THF, the polymerizations of the NCA were carried out to give VB-OPhe macromers, which were found to have very narrow molecular weight distributions. Furthermore, their functionalities were proved to be approximately unity by the polymerization method.