Publications by authors named "Hanieh Ghassabian"

4 Publications

  • Page 1 of 1

: An Screening Targeting HCMV ppUL44 Processivity Factor Homodimerization Identifies Small Molecules Inhibiting Viral Replication.

Viruses 2021 05 20;13(5). Epub 2021 May 20.

Department of Molecular Medicine, University of Padova, 35121 Padova, Italy.

Human cytomegalovirus (HCMV) is a leading cause of severe diseases in immunocompromised individuals, including AIDS patients and transplant recipients, and in congenitally infected newborns. The utility of available drugs is limited by poor bioavailability, toxicity, and emergence of resistant strains. Therefore, it is crucial to identify new targets for therapeutic intervention. Among the latter, viral protein-protein interactions are becoming increasingly attractive. Since dimerization of HCMV DNA polymerase processivity factor ppUL44 plays an essential role in the viral life cycle, being required for oriLyt-dependent DNA replication, it can be considered a potential therapeutic target. We therefore performed an screening and selected 18 small molecules (SMs) potentially interfering with ppUL44 homodimerization. Antiviral assays using recombinant HCMV TB4-UL83-YFP in the presence of the selected SMs led to the identification of four active compounds. The most active one, B3, also efficiently inhibited HCMV AD169 strain in plaque reduction assays and impaired replication of an AD169-GFP reporter virus and its ganciclovir-resistant counterpart to a similar extent. As assessed by Western blotting experiments, B3 specifically reduced viral gene expression starting from 48 h post infection, consistent with the inhibition of viral DNA synthesis measured by qPCR starting from 72 h post infection. Therefore, our data suggest that inhibition of ppUL44 dimerization could represent a new class of HCMV inhibitors, complementary to those targeting the DNA polymerase catalytic subunit or the viral terminase complex.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/v13050941DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8160850PMC
May 2021

Generation of Combinatorial Lentiviral Vectors Expressing Multiple Anti-Hepatitis C Virus shRNAs and Their Validation on a Novel HCV Replicon Double Reporter Cell Line.

Viruses 2020 09 18;12(9). Epub 2020 Sep 18.

Department of Molecular Medicine, University of Padua, 35121 Padua, Italy.

Despite the introduction of directly acting antivirals (DAAs), for the treatment of hepatitis C virus (HCV) infection, their cost, patient compliance, and viral resistance are still important issues to be considered. Here, we describe the generation of a novel JFH1-based HCV subgenomic replicon double reporter cell line suitable for testing different antiviral drugs and therapeutic interventions. This cells line allowed a rapid and accurate quantification of cell growth/viability and HCV RNA replication, thus discriminating specific from unspecific antiviral effects caused by DAAs or cytotoxic compounds, respectively. By correlating cell number and virus replication, we could confirm the inhibitory effect on the latter of cell over confluency and characterize an array of lentiviral vectors expressing single, double, or triple cassettes containing different combinations of short hairpin (sh)RNAs, targeting both highly conserved viral genome sequences and cellular factors crucial for HCV replication. While all vectors were effective in reducing HCV replication, the ones targeting viral sequences displayed a stronger antiviral effect, without significant cytopathic effects. Such combinatorial platforms as well as the developed double reporter cell line might find application both in setting-up anti-HCV gene therapy approaches and in studies aimed at further dissecting the viral biology/pathogenesis of infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/v12091044DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7551853PMC
September 2020

New Frontiers of Corneal Gene Therapy.

Hum Gene Ther 2019 08 30;30(8):923-945. Epub 2019 May 30.

1Department of Molecular Medicine, University of Padova, Padova, Italy.

Corneal diseases are among the most prevalent causes of blindness worldwide. The transparency and clarity of the cornea are guaranteed by a delicate physiological, anatomic, and functional balance. For this reason, all the disorders, including those of genetic origin, that compromise this state of harmony can lead to opacity and eventually vision loss. Many corneal disorders have a genetic etiology, and some are associated with rather rare and complex syndromes. Conventional treatments, such as corneal transplantation, are often ineffective, and to date, many of these disorders are still incurable. Gene therapy carries the promise of being a potential cure for many of these diseases, with solutions and strategies that did not seem possible until a few years ago. With its potential to treat genetic disease by means of deletion, replacement, or editing of a defective gene, the challenge can also be extended to corneal disorders in order to achieve long-term, if not definitive, relief. The aim of this paper is to review the state of the art of the different gene therapy approaches as potential treatments for corneal diseases and the future perspectives for the development of personalized gene-based medicine.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1089/hum.2019.026DOI Listing
August 2019

Contribution of the residue at position 4 within classical nuclear localization signals to modulating interaction with importins and nuclear targeting.

Biochim Biophys Acta Mol Cell Res 2018 08 8;1865(8):1114-1129. Epub 2018 May 8.

Department of Molecular Medicine, University of Padua, Via Gabelli 63, 35121 Padua, Italy. Electronic address:

Nuclear import involves the recognition by importin (IMP) superfamily members of nuclear localization signals (NLSs) within protein cargoes destined for the nucleus, the best understood being recognition of classical NLSs (cNLSs) by the IMPα/β1 heterodimer. Although the cNLS consensus [K-(K/R)-X-(K/R) for positions P2-P5] is generally accepted, recent studies indicated that the contribution made by different residues at the P4 position can vary. Here, we apply a combination of microscopy, molecular dynamics, crystallography, in vitro binding, and bioinformatics approaches to show that the nature of residues at P4 indeed modulates cNLS function in the context of a prototypical Simian Virus 40 large tumor antigen-derived cNLS (KKRK, P2-5). Indeed, all hydrophobic substitutions in place of R impaired binding to IMPα and nuclear targeting, with the largest effect exerted by a G residue at P4. Substitution of R with neutral hydrophobic residues caused the loss of electrostatic and van der Waals interactions between the P4 residue side chains and IMPα. Detailed bioinformatics analysis confirmed the importance of the P4 residue for cNLS function across the human proteome, with specific residues such as G being associated with low activity. Furthermore, we validate our findings for two additional cNLSs from human cytomegalovirus (HCMV) DNA polymerase catalytic subunit UL54 and processivity factor UL44, where a G residue at P4 results in a 2-3-fold decrease in NLS activity. Our results thus showed that the P4 residue makes a hitherto poorly appreciated contribution to nuclear import efficiency, which is essential to determining the precise nuclear levels of cargoes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bbamcr.2018.05.006DOI Listing
August 2018
-->