Molecular docking,pharmacophore based virtual screening and molecular dynamics studies towards the identification of potential leads for the management of H. pylori

The enzyme pantothenate synthetase panC is one of the potential new antimicrobial drug targets, but it is poorly characterized in H. pylori. H. pylori infection can cause gastric cancer and the management of H. pylori infection is crucial in various gastric ulcers and gastric cancer. The current study describes the use of innovative drug discovery and design approaches like comparative metabolic pathway analysis (Metacyc), exploration of database of essential genes (DEG), homology modelling, pharmacophore based virtual screening, ADMET studies and molecular dynamics simulations in identifying potential lead compounds for the H. pylori specific panC. The top ranked virtual hits STOCK1N-60270, STOCK1N-63040, STOCK1N-44424 and STOCK1N-63231 can act as templates for synthesis of new H. pylori inhibitors and they hold a promise in the management of gastric cancers caused by H. pylori.

Computational approaches such as pharmacophore modeling, virtual screening and MD simulations were explored to find the potential hits as H. pylori specific panC inhibitors for the management of gastric ulcers and gastric cancers.     

The suppression of lung tumorigenesis by aerosol-delivered folate–chitosan-graft-polyethylenimine/Akt1 shRNA complexes through the Akt signaling pathway

RNA interference (RNAi) represents a promising new approach to the inhibition of gene expression in vitro and in vivo, and has therapeutic potential for human diseases. Efficient delivery of small interfering RNA (siRNA) or small hairpin RNA (shRNA) is a critical concern in RNAi studies. Here we report the development of a new polymeric gene carrier for cancer cell-targeting, designed to enhance the intracellular delivery of shRNA and reduce cytotoxicity. Folate–chitosan-graft-polyethylenimine (FC-g-PEI) copolymer was prepared by an imine reaction between periodate-oxidized folate–chitosan (FC) and low molecular weight polyethylenimine (PEI). FC-g-PEI copolymer was investigated as a potential cancer cell-targeting gene carrier. The composition of FC-g-PEI was characterized using ¹H nuclear magnetic resonance (¹H NMR), and particle size and zeta potential of FC-g-PEI/shRNA complexes were measured using dynamic light scattering (DLS). FC-g-PEI showed good shRNA condensation ability and high protection of shRNA from nuclease attack. It also exhibited lower cytotoxicity compared to PEI 25K control, and showed good cancer cell-targeting ability. Furthermore, aerosol delivery of FC-g-PEI/Akt1 shRNA complexes suppressed lung tumorigenesis in a urethane-induced lung cancer model mouse through the Akt signaling pathway. Together, these results suggest that FC-g-PEI may be useful for shRNA-based gene therapy.     

Chitosan-graft-polyethylenimine as a gene carrier.

Chitosans have been proposed as biocompatible alternative cationic polymers that are suitable for non-viral delivery. However, the transfection efficiency of chitosan-DNA nanoparticles is still very low. To improve transfection efficiency, we prepared chitosan-graft-polyethylenimine (CHI-g-PEI) copolymer by an imine reaction between periodate-oxidized chitosan and polyethylenimine (PEI). The molecular weight and composition of the CHI-g-PEI copolymer were characterized, using multi-angle laser scattering (GPC-MALS) and (1)H nuclear magnetic resonance ((1)H NMR), respectively. The copolymer was complexed with plasmid DNA (pDNA) in various copolymer/DNA (N/P) charge ratios, and the complex was characterized. CHI-g-PEI showed good DNA binding ability and high protection of DNA from nuclease attack. Also, with an increase in charge ratio, the sizes of the CHI-g-PEI/DNA complex showed a tendency to decrease, whereas the zeta potential of the complex showed an increase. The CHI-g-PEI copolymer had low cytotoxicity, compared to PEI 25K from cytotoxicity assays. At high N/P ratios, the CHI-g-PEI/DNA complex showed higher transfection efficiency than PEI 25K in HeLa, 293T and HepG2 cell lines. Our results indicate that the CHI-g-PEI copolymer has potential as a gene carrier in vitro.     

Galactosylated chitosan-graft-polyethylenimine as a gene carrier for hepatocyte targeting

Chitosans have been proposed as alternative, biocompatible cationic polymers for nonviral gene delivery. However, the low transfection efficiency and low specificity of chitosan need to be addressed before clinical application. We prepared galactosylated chitosan-graft-polyethylenimine (GC-g-PEI) copolymer by an imine reaction between periodate-oxidized GC and low-molecular-weight PEI. The molecular weight and composition were characterized using gel permeation chromatography column with multi-angle laser scattering and (1)H nuclear magnetic resonance, respectively. The copolymer was complexed with plasmid DNA in various copolymer/DNA (N/P) charge ratios, and the complexes were characterized. GC-g-PEI showed good DNA-binding ability and superior protection of DNA from nuclease attack and had low cytotoxicity compared to PEI 25K. GC-g-PEI/DNA complexes showed higher transfection efficiency than PEI 25K in both HepG2 and HeLa cell lines. Transfection efficiency into HepG2, which has asialoglycoprotein receptors, was higher than that into HeLa, which does not. GC-g-PEI/DNA complexes also transfected liver cells in vivo after intraperitoneal (i.p.) administration more effectively than PEI 25K. These results suggest that GC-g-PEI can be used in gene therapy to improve transfection efficiency and hepatocyte specificity in vitro and in vivo.     

Transfection of primary human nasal epithelial cells using a biodegradable poly (ester amine) based on polycaprolactone and polyethylenimine as a gene carrier

The purpose of this study was to prepare and characterize poly (ester amine) (PEA)/pGL3 complexes and investigate their transfection efficiency in human nasal epithelial (HNE) cells. Particle size, zeta potential, and gel retardation characteristics of PEA /pGL3 complexes were also measured. After treatment of DNase-I, protection and release assay of PEA/pGL3 complexes were performed. To assess the transfection efficiency and cytotoxicity, measurement of relative luciferase activity and MTS assay were performed. PEA/pGL3 complexes showed effective and stable DNA condensation with the particle sizes below 200 nm, implicating their potential for intracellular delivery. PEA/pGL3 complexes successfully transfected into the HNE cells with higher viability of the cells. These results suggested that, the PEA can be used as an efficient cationic polymeric vehicle which provides a versatile platform for further investigation of structure property relationship along with the controlled degradation, significant low cytotoxicity, and high transfection efficiency of the primary HNE cells.     

Degradable poly(amido amine)s as gene delivery carriers

INTRODUCTION: In recent years, there has been a great deal of interest in the development of vectors which are being developed based on the capacity of polymers to mediate appropriate interactions with the cellular environment, or to interface with specific cellular processes. Several such vectors have been synthesized, resulting in biomacromolecules with low cytotoxicity and higher gene delivery ability. AREAS COVERED: This review briefly describes the recent success of poly(amido amine)s (PAAs) as non-viral vectors, and highlights their promising future in the development of nucleic acid-based therapy. It also provides an overview on the synthesis, characterization and application of PAAs as gene carriers, which will be useful for various biological motifs. This review helps the readers to better understand the emergence of non-viral vectors for gene therapy, especially PAAs, their properties, their advantages and disadvantages and the gene therapy based on them. EXPERT OPINION: The future of gene-based therapy needs to identify approaches to develop new carriers, depending on the properties of the biological membranes they face, and their physicochemical properties, in order to successfully deliver the genes to the target sites. With the emergence of a variety of non-viral vectors, such as biodegradable polymers, it may not take long before non-viral vectors are observed that are not just safe and tissue-specific, but even more efficient than viral vectors.     

Aerosol delivery of beclin1 enhanced the anti-tumor effect of radiation in the lungs of K-rasLA1 mice

Radiotherapy alone has several limitations for treating lung cancer. Inhalation, a non-invasive approach for direct delivery of therapeutic agents to the lung, may help to enhance the therapeutic efficacy of radiation. Up-regulating beclin1, known as a tumor suppressor gene that plays a major role in autophagy, may sensitize tumors and lead to tumor regression in lungs of K-ras^LA1 lung cancer model mice. To minimize the side-effects of radiotherapy, fractionated exposures (five times, 24-h interval) with low dose (2 Gy) of radiation to the restricted area (thorax, 2 cm) were conducted. After sensitizing the lungs with radiation, beclin1, complexed with a nano-sized biodegradable poly(ester amine), was prepared and delivered into the murine lung via aerosol three times/week for four weeks. In a histopathological analysis, animals treated with beclin1 and radiation showed highly significant tumor regression and low progression to adenocarcinoma. An increase in the number of autophagic vacuoles and secondary lysosomes was detected. Dissociation of beclin1-bcl2 stimulated autophagy activation and showed a synergistic anti-tumor effect by inhibiting the Akt-mTOR pathway, cell proliferation and angiogenesis. The combination of radiation with non-invasive aerosol delivery of beclin1 may provide a prospect for developing novel therapy regimens applicable in clinics.     

Poly (amino ester) Composed of Poly (ethylene glycol) and Aminosilane Prepared by Combinatorial Chemistry as a Gene Carrier

Purpose Application of combinatorial chemistry and high throughput screening for the synthesis and evaluation of mini-library of novel biodegradable poly (β-amino ester)s (PAE)s composed of γ-aminopropyl-triethoxysilane (APES) and poly (ethylene glycol) diacrylate (PEGDA) for gene delivery efficiency and safety in 293T and HeLa cells in the presence of and absence of serum. Materials and methods PAEs were synthesized at different mole ratios of APES and PEGDA by Michael addition reaction and synthesis was confirmed by ¹H nuclear magnetic resonance (¹H-NMR). Ninety six ratios of polyplexes were evaluated for luciferase and MTS assay in 293T and HeLa cells in the presence of and absence of serum. Relationship between transfection efficiency and DNA binding ability of PAEs was studied by gel electrophoresis. Particle sizes and molecular weight of selected PAEs were measured by dynamic light scattering and gel permeation chromatography multi-angle light scattering, respectively. Results ¹H-NMR confirmed the synthesis of PAEs. In both cell lines, transfection efficiency and cell viability were increased for PAEs obtained from R106 (0.7:1, APES:PEGDA) to R121 (6:1, APES:PEGDA) with a marginal increase in APES concentration. Transfection pattern was uniform in the absence of and presence of serum. In both cell lines, PAE obtained from R121 demonstrated high transfection efficiency and low cytotoxicity as compared to polyethylenimine (25 KDa) and Lipofectamine. PAE obtained from R121 showed good DNA binding and condensation with average particle sizes of 133 nm. Conclusion Addition of PEGDA over APES resulted in a novel PAE which has high safety and transfection efficiency. Transfection and cytotoxicity are very sensitive to monomer ratios and mainly governed by concentration of amine monomer.