Biology subjects

Vasin, A.

Publications and source records attributed to Vasin, A..

5 recordsLinked to original sources

bioRxiv2023

Cell-penetrating peptide and cationic liposomes mediated siRNA delivery to arrest growth of chronic myeloid leukemia cells in vitro

Gene silencing through RNA interference (RNAi) is a promising therapeutic approach for a wide range of disorders, including cancer. Non-viral gene therapy, using specific siRNAs against BCR-ABL, can be a supportive or alternative measure to traditional chronic myeloid leukemia (CML) tyrosine kinase inhibitor (TKIs) therapies, given the prevalence of clinical TKI resistance. The main challenge for such approaches remains the development of the effective delivery system for siRNA tailored to the specific disease model. The purpose of this study was to examine and compare the efficiency of endosomolytic cell penetrating peptide (CPP) EB1 and PEG2000-decorated cationic liposomes composed of polycationic lipid 1,26-bis(cholest-5-en-3-yloxycarbonylamino)-7,11,16,20-tetraazahexacosane tetrahydrochloride (2X3) and helper lipid 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) for anti-bcr-abl siRNA delivery into the K562 human CML cell line. We show that both EB1 and 2X3-DOPE-DSPE-PEG2000 (0.62% mol.) liposomes effectively deliver siRNA into K562 cells by endocytic mechanisms, and the use of liposomes leads to more effective inhibition of expression of the targeted gene (BCR-ABL) and cancer cell proliferation. Taken together, these findings suggest that PEG-decorated cationic liposomes mediated siRNA delivery allows an effective antisense suppression of certain oncogenes, and represents a promising new class of therapies for CML.

molecular biology↗

bioRxiv2021

Kinetics Of Interferon-{lambda} And Receptor Expression In Response To In Vitro Respiratory Viral Infection

The major protective immune response against viruses is production of type I and III interferons (IFNs). IFNs induce the expression of hundreds of IFN-stimulated genes (ISGs) that block viral replication and further viral spread. The ability of respiratory viruses to suppress induction of IFN-mediated antiviral defenses in infected epithelial cells may be a factor contributing to the particular pathogenicity of several strains. In this report, we analyzed expression of IFNs and some ISGs in an alveolar epithelial cell subtype (A549) in response to infection with: influenza A viruses (A/California/07/09pdm (H1N1), A/Texas/50/12 (H3N2)); influenza B virus (B/Phuket/3073/13); adenovirus type 5 and 6; or respiratory syncytial virus (strain A2). IFNL and ISGs expression significantly increased in response to infection with all RNA viruses 24 hpi. Nevertheless, only IBV led to early increase in IFNL and ISGs mRNA level. IBV and H1N1 infection led to elevated proinflammatory cytokine production. We speculate that augmented IFN-, IFN-{beta}, IL-6 levels negatively correlate to SOCS1 expression. Importantly, we showed a decrease in IFNLR1 mRNA in case of IBV infection that implies the existence of negative ISGs expression regulation at IFN{lambda}R level. It could be either a specific feature of IBV or a consequence of early IFNL expression.

immunology↗

bioRxiv2020

Clean and folded: optimized production of high quality recombinant human interferon-{lambda}1

Type III interferons exhibit antiviral activity against influenza viruses, coronaviruses, rotaviruses, and others. In addition, this type of interferon theoretically has therapeutic advantages, in comparison with type I interferons, due to its ability to activate a narrower group of genes in a relatively small group of target cells. Hence, it can elicit more targeted antiviral or immunomodulatory responses. Obtaining biologically-active interferon lambda (hIFN-{lambda}1) is fraught with difficulties at the stage of expression in soluble form or, in the case of expression in the form of inclusion bodies, at the stage of refolding. In this work, hIFN-{lambda}1 was expressed in the form of inclusion bodies, and a simple, effective refolding method was developed. Efficient and scalable methods for chromatographic purification of recombinant hIFN-{lambda}1 were also developed. High-yield, high-purity product was obtained through optimization of several processes including: recombinant protein expression; metal affinity chromatography; cation exchange chromatography; and an intermediate protein refolding stage. The obtained protein was shown to have expected, specific biological activity in line with published effects: induction of MxA gene expression in A549 cells.

bioengineering↗

bioRxiv2019

Cold and distant: structural features of the nucleoprotein complex of a cold-adapted influenza A virus strain

Two influenza A nucleoprotein variants (wt: G102R; and mutant: G102R and E292G) were studied with regard to macro-molecular interactions in oligomeric form (24-mers). The E292G mutation has been previously shown to provide cold adaptation. Molecular dynamics simulations of these complexes and trajectory analysis showed that the most significant difference between the obtained models was distance differences between nucleoprotein complex strands. Influenza virus nucleoprotein complexes were isolated from strains bearing the corresponding NP amino acid substitutions. The isolated complexes were characterized by transmission electron microscopy and differential scanning fluorimetry (DSF). Presence of the E292G substitution was shown by DSF to affect nucleoprotein complex melting temperature. In the filament interface peptide model, it was shown that the peptide corresponding in primary structure to the wild-type NP (SGYDFEREGYS, wild type peptide) is prone to temperature-dependent self-association, unlike the peptide carrying the substitution corresponding to E292G (SGYDFGREGYS, mutant peptide). It was also shown that the SGYDFEREGYS peptide (wt) is capable of interacting with a recombinant full-size monomeric nucleoprotein (with primary structure corresponding to wild type); this interactions equilibrium dissociation constant is five orders of magnitude lower than for the SGYDFGREGYS peptide. Using small-angle neutron scattering (SANS), the supramolecular structures of isolated complexes of these proteins was studied at temperatures of 15, 32, and 37{degrees}C. SANS data show that the structures of the studied complexes (mutant or normal proteins with RNA) at elevated temperature differ from the rod-like particle model and react differently to temperature changes. The data suggest that the mechanism behind cold adaptation with E292G is associated with a weakening of the interaction between strands of the ribonucleoprotein complex and, as a result, the appearance of inter-chain interface flexibility necessary for complex function at low temperature.

biophysics↗

bioRxiv2018

Novel Computational Method to Define RNA PSRs Explains Influenza A Virus Nucleotide Conservation

ABSTRACTRNA molecules often fold into evolutionarily selected functional structures. Yet, the literature offers neither a satisfactory definition for \"structured RNA regions\", nor a computational method to accurately identify such regions. Here, we define structured RNA regions based on the premise that both stems and loops in functional RNA structures should be conserved among RNA molecules sharing high sequence homology. In addition, we present a computational approach to identify RNA regions possessing evolutionarily conserved secondary structures, RNA ISRAEU (RNA Identification of Structured Regions As Evolutionary Unchanged). Applying this method to H1N1 influenza mRNAs revealed previously unknown structured RNA regions that are potentially essential for viral replication and/or propagation. Evolutionary conservation of RNA structural elements may explain, in part, why mutations in some nucleotide positions within influenza mRNAs occur significantly more often than in others. We found that mutations occurring in conserved nucleotide positions may be more disruptive for structured RNA regions than single nucleotide polymorphisms in positions that are more prone to changes. Finally, we predicted computationally a previously unknown stem-loop structure and demonstrated that oligonucleotides complementing the stem (but not the loop or unrelated sequences) reduce viral replication in vitro. These results contribute to understanding influenza A virus evolution and can be applied to rational design of attenuated vaccines and/or drug designs based on disrupting conserved RNA structural elements.\n\nAUTHOR SUMMARYRNA structures play key biological roles. However, the literature offers neither a satisfactory definition for \"structured RNA regions\" nor the computational methodology to identify such regions. We define structured RNA regions based on the premise that functionally relevant RNA structures should be evolutionarily conserved, and devise a computational method to identify RNA regions possessing evolutionarily conserved secondary structural elements. Applying this method to influenza virus mRNAs of pandemic and seasonal H1N1 influenza A virus generated Predicted Structured Regions (PSRs), which were previously unknown. This explains the previously mysterious sequence conservation among evolving influenza strains. Also, we have experimentally supported existence of a computationally predicted stem-loop structure predicted computationally. Our approach may be useful in designing live attenuated influenza vaccines and/or anti-viral drugs based on disrupting necessary conserved RNA structures.

bioinformatics↗