Synthesis and anticancer activity of new [(Indolyl)pyrazolyl]-1,3,4-oxadiazole thioglycosides and acyclic nucleoside analogs

New [(Indolyl)pyrazolyl]-1,3,4-oxadiazole compounds and their derived thioglycosides as well as the corresponding sugar hydrazones were synthesized. The acyclo C-nucleoside analogs of the oxadiazoline base system were also prepared by reaction of acid hydrazides with aldehydo sugars followed by one pot process encompassing acetylation and cyclization of the synthesized hydrazones. The anticancer activity of the newly synthesized compounds was studied against colorectal carcinoma (HCT116), breast adenocarcinoma (MCF7) and prostate cancer (PC3) human tumor cell lines and a number of compounds showed moderate to high activities.     

Molecular characterization of exons 6, 8 and 9 of ABCB4 gene in children with Progressive Familial Intrahepatic Cholestasis type 3

Aim of work: To estimate the frequency of mutations involving exons 6, 8 and 9 of Adenosine triphosphate-binding cassette, subfamily B, member 4 (ABCB4) gene among children with progressive intrahepatic cholestasis with high γ-GT activity (PFIC3).

Subjects and methods: Cross sectional study was conducted on 30 children with PFIC3. Genotyping was performed by sequencing analysis of exons 6, 8 and exon 9 of ABCB4 gene.

Results: Heterozygous synonymous polymorphic variant was detected in exon 6 (rs 1202283) and in exon 8 (rs 2109505). No mutations in studied exons were detected.

Conclusion: Exons 6, 8 and 9 mutations of ABCB4 gene are not common among Egyptian children with PFIC3.     

Formulation and <Emphasis Type="Italic">In Vitro</Emphasis> and <Emphasis Type="Italic">In Vivo</Emphasis> Evaluation of Lipid-Based Terbutaline Sulphate Bi-layer Tablets for Once-Daily Administration

The objective of this study was to prepare and evaluate terbutaline sulphate (TBS) bi-layer tablets for once-daily administration. The bi-layer tablets consisted of an immediate-release layer and a sustained-release layer containing 5 and 10 mg TBS, respectively. The sustained-release layer was developed by using Compritol®888 ATO, Precirol® ATO 5, stearic acid, and tristearin, separately, as slowly eroding lipid matrices. A full 4?×?2² factorial design was employed for optimization of the sustained-release layer and to explore the effect of lipid type (X ₁), drug–lipid ratio (X ₂), and filler type (X ₃) on the percentage drug released at 8, 12, and 24 h (Y ₁, Y ₂, and Y ₃) as dependent variables. Sixteen TBS sustained-release matrices (F1–F16) were prepared by melt solid dispersion method. None of the prepared matrices achieved the targeted release profile. However, F2 that showed a relatively promising drug release was subjected to trial and error optimization for the filler composition to develop two optimized matrices (F17 and F18). F18 which consisted of drug–Compritol®888 ATO at ratio (1:6 w/w) and Avicel PH 101/dibasic calcium phosphate mixture of 2:1 (w/w) was selected as sustained-release layer. TBS bi-layer tablets were evaluated for their physical properties, in vitro drug release, effect of storage on drug content, and in vivo performance in rabbits. The bi-layer tablets showed acceptable physical properties and release characteristics. In vivo absorption in rabbits revealed initial high TBS plasma levels followed by sustained levels over 24 h compared to immediate-release tablets.     

A synthesis of 3-O-(α-d-mannopyranosyl)-d-mannose and its protein conjugate

Methods for the synthesis of 3-O-(α-d-mannopyranosyl)-d-mannose and 2-(4-aminophenyl)ethyl 3-O-(α-d-mannopyranosyl)-α-d-mannopyranoside have been investigated by a number of sequences. Glycosidations with 2,3-di-O-acetyl-4,6-di-O-benzyl-d-mannopyranosyl and 2-O-benzoyl-3,4,6-tri-O-benzyl-d-mannopyranosyl p-toluenesulfonates were found to give better yields than the Helferich modification, the use of a peracylated d-mannopyranosyl halide, or the use of triflyl leaving group. Only the α anomer was obtained. Factors influencing glycosidation reactions are discussed. A mercury(II) complex was used for selective 2-O-acylation of 4,6-di-O-benzyl-α-d-mannopyranosides. A disaccharide—protein conjugate was prepared by the isothiocyanate method.     

Identification of potential inhibitors for HCV NS5b of genotype 4a by combining dynamic simulation,protein–ligand interaction fingerprint, 3D pharmacophore,docking and 3D QSAR

Abstract

HCV NS5B polymerase has been one of the most attractive targets for developing new drugs for HCV infection and many drugs were successfully developed, but all of them were designed for targeting Hepatitis C Virus genotype 1 (HCV GT1). Hepatitis C virus genotype 4a (HCV GT4a) dominant in Egypt has paid less attention. Here, we describe our protocol of virtual screening in identification of novel potential potent inhibitors for HCV NS5B polymerase of GT4a using homology modeling, protein–ligand interaction fingerprint (PLIF), docking, pharmacophore, and 3D CoMFA quantitative structure activity relationship (QSAR). Firstly, a high-quality 3D model of HCV NS5B polymerase of GT4a was constructed using crystal structure of HCV NS5B polymerase of GT1 (PDB ID: 3hkw) as a template. Then, both the model and the template were simulated to compare conformational stability. PLIF was generated using five crystal structures of HCV NS5B (PDB ID: 4mia, 4mib, 4mk9, 4mka, and 4mkb), which revealed the most important residues and their interactions with the co-crystalized ligands. After that, a 3D pharmacophore model was developed from the generated PLIF data and then used as a screening filter for 17000328 drug-like zinc database compounds. 900 compounds passed the pharmacophore filter and entered the docking-based virtual screening stage. Finally, a 3D CoMFA QSAR was developed using 42 compounds as a training and 19 compounds as a test set. The 3D CoMFA QSAR was used to design and screen some potential inhibitors, these compounds were further evaluated by the docking stage. The highest ranked five hits from docking result (compounds (p1–p4) and compound q1) were selected for further analysis.

Communicated by Ramaswamy H. Sarma     

Dosimetric evaluation study of IMRT and VMAT techniques for prostate cancer based on different multileaf collimator designs

The hypofractionated radiotherapy modality was established to reduce treatment durations and enhance therapeutic efficiency, as compared to conventional fractionation treatment. However, this modality is challenging because of rigid dosimetric constraints. This study aimed to assess the impact of multi-leaf collimator (MLC) widths (10 mm and 5 mm) on plan quality during the treatment of prostate cancer. Additionally, this study aimed to investigate the impact of the MLC mode of energy on the Agility flattening filter (FF), MLC Agility-free flattening filter (FFF), and MLCi2 for patients receiving hypofractionated radiotherapy. Two radiotherapy techniques; Intensity Modulated Radiotherapy (IMRT) and Volumetric Modulated Arc Radiotherapy (VMAT), were used in this research. In the present study, computed tomography simulations of ten patients (six plans per patient) with localized prostate adenocarcinoma were analyzed. Various dosimetric parameters were assessed, including monitor units, treatment delivery times, conformity, and homogeneity indices. To evaluate the plan quality, dose-volume histograms (DVHs) were estimated for each technique. The results demonstrated that the determined dosimetric parameters of planning target volume (PTV)p (such as D mean, conformity, and homogeneity index) showed greater improvement with MLC Agility FF and MLC Agility FFF than with MLCi2. Additionally, the treatment delivery time was reduced in the MLC Agility FF (by 31%) and MLC Agility FFF (by 10.8%) groups compared to the MLCi2 group. It is concluded that for both the VMAT and IMRT techniques, the smaller width (5 mm) MLCs revealed better planning target volume coverage, improved the dosimetric parameters for PTV, reduced the treatment time, and met the constraints for OARs. It is therefore recommended to use 5 mm MLCs for hypofractionated prostate cancer treatment due to better target coverage and better protection of OARs.

     

Efficacy of Nile tilapia (Oreochromis niloticus) juveniles and spinosyns bioinsecticides against aquatic stages of Culex pipiens: An experimental study

The mosquito Culex pipiens is the most widely distributed dipteran species in all regions of Egypt and the principal vector of Wuchereria bancrofti and certain arboviruses in human beings. For controlling C. pipiens vector, biological tools (e.g., larvivorous fish and bioinsecticides) are more potent and safer options to the environment, human beings, and beneficial organisms than chemical pesticides. The efficiency of O. niloticus juveniles as predatory fish species and two bioinsecticides, spinosad 24% and spinetoram 12%, was investigated against the C. pipiens developmental stages in the laboratory. The first trial evaluated the predatory efficacy of small-sized O. niloticus (2.1–2.6 cm; 250–315 mg) and large-sized O. niloticus (2.5–3.2 cm; 250–315 mg) against the 3rd larvae and pupae of C. pipiens. This is the first report in Egypt confirming the predation potential of O. niloticus as efficient predatory fish against the immature C. pipiens. Large-sized O. niloticus predated a greater number of 3rd of C. pipiens larvae and pupae than the small-sized ones. Furthermore, the daily consumption of C. pipiens larvae by small- and large-sized O. niloticus was significantly higher than the pupae. The second trial assessed the toxicity efficacy of spinosad 24% and spinetoram 12% against C. pipiens larvae and pupae. The results confirmed that the tested bioinsecticides showed higher potency toward C. pipiens larvae than pupae after exposure for 24 h and 48 h. Spinosad was more toxic toward 3rd C. pipiens larvae (LC₅₀ = 0.013 and 0.003 mg/L) and pupae (LC₅₀ = 320.69 and 44.28 mg/L) than spinetoram after 24 and 48 h. Herein, O. niloticus juveniles (as promising native predatory fish) and spinosyns bioinsecticides were more effective against C. pipiens in the larval stage than in the pupal stage. In conclusion, Nile tilapia juveniles and biorational compounds, spinosad 24% and spinetoram 12%, might be considered as promising and favorable environmental biological agents for controlling C. pipiens in Egypt. However, further trials are needed to investigate the potential of these agents in the control of this mosquito vector under field conditions.     

Developmental toxicity of orally administered sildenafil citrate (Viagra) in SWR/J mice

Normal adult inbred SWR/J mice were used to investigate the teratogenic and other possible toxic effects of various dose levels of sildenafil citrate (Viagra) on fetuses. Multiple dose levels of 6.5, 13.0, 19.5, 26.0, 32.5 or 40.0 mg of sildenafil citrate/kg body weight (which correspond to the multiples of 1, 2, 3, 4, 5 or 6 of human 50 mg Viagra, respectively) were orally administered into pregnant mice on days 7–9, 10–12 or 13–15 of gestation. On day 17 of pregnancy, all fetuses were removed and examined for toxic phenomena (embryo-fetal toxicity) and for external, internal and skeletal malformations. A total of 285 pregnant mice were used in the present study.None of the dams treated with sildenafil citrate at any of the oral dose levels used in the present study died during the experimental period and all dams treated with the drug failed to reveal overt signs of maternal toxicity. Moreover, the results of the present study clearly demonstrate that none of the multiple oral dose levels of the drug at any time interval used has induced any external, internal or skeletal malformations in the fetuses obtained from treated females.However, the dose level of 40 mg/kg body weight of sildenafil citrate has a growth suppressing effect on alive fetuses when it was administered at all the time intervals used in the present study. Furthermore, the dose levels 26.0, 32.5 and 40 mg/kg of the drug have embryo-fetal toxicity when the drug is applied on days 13–15 of gestation. The possible mechanisms involved in the embryo-fetal toxicity and fetal growth suppressing effects of sildenafil citrate were discussed.The results of this study have important implications for the widespread use of this drug.     

Recombineering Homologous Recombination Constructs in Drosophila

The continued development of techniques for fast, large-scale manipulation of endogenous gene loci will broaden the use of Drosophila melanogaster as a genetic model organism for human-disease related research. Recent years have seen technical advancements like homologous recombination and recombineering. However, generating unequivocal null mutations or tagging endogenous proteins remains a substantial effort for most genes. Here, we describe and demonstrate techniques for using recombineering-based cloning methods to generate vectors that can be used to target and manipulate endogenous loci in vivo. Specifically, we have established a combination of three technologies: (1) BAC transgenesis/recombineering, (2) ends-out homologous recombination and (3) Gateway technology to provide a robust, efficient and flexible method for manipulating endogenous genomic loci. In this protocol, we provide step-by-step details about how to (1) design individual vectors, (2) how to clone large fragments of genomic DNA into the homologous recombination vector using gap repair, and (3) how to replace or tag genes of interest within these vectors using a second round of recombineering. Finally, we will also provide a protocol for how to mobilize these cassettes in vivo to generate a knockout, or a tagged gene via knock-in. These methods can easily be adopted for multiple targets in parallel and provide a means for manipulating the Drosophila genome in a timely and efficient manner.