Development of High-Throughput Screening Assay for Antihantaviral Therapeutics.
Humans acquire hantavirus infection by the inhalation of aerosolized excreta of infected rodent hosts. There is no treatment for hantavirus diseases at present. Therapeutic intervention during early stages of viral infection can improve the outcome of this zoonotic viral illness. The interaction between an evolutionary conserved sequence at the 5' terminus of hantaviral genomic RNA and hantavirus nucleocapsid protein plays a critical role in the hantavirus replication cycle. This unique interaction is a novel target for therapeutic intervention of hantavirus disease. We developed a very sensitive, tractable, and cost-effective fluorescence-based assay to monitor the interaction between the nucleocapsid protein and the target RNA sequence. The assay was optimized for high-throughput screening of chemical libraries to identify molecules that interrupt this RNA-protein interaction. The assay was validated using a library of 6880 chemical compounds. This validation screen demonstrated the reproducibility and validity of required statistical criteria for high-throughput screening. The assay is ready to use for high-throughput screening of large chemical libraries to identify antihantaviral therapeutic molecules and can be amenable to similar targets in other viruses.
1 High-Throughput Screening Lab, University of Kansas, Lawrence, KS, USA.
Anuradha
A
Roy
2 College of Veterinary Medicine, Western University of Health Sciences, Pomona, CA, USA.
Mohammad A
MA
Mir
D000998:Antiviral Agents / Q000494:pharmacology*; D055808:Drug Discovery / Q000379:methods; D006476:Orthohantavirus / Q000187:drug effects* / Q000302:isolation & purification / Q000502:physiology; D018778:Hantavirus Infections / Q000188:drug therapy / Q000821:virology; D057166:High-Throughput Screening Assays; D006801:Humans; D011485:Protein Binding; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D020506:Untranslated Regions; D014764:Viral Proteins / Q000378:metabolism; D014779:Virus Replication / Q000187:drug effects
D000998:Antiviral Agents; D012367:RNA, Viral; D020506:Untranslated Regions; D014764:Viral Proteins
Bunyaviridae family; hantavirus replication; high-throughput screen; negative-strand RNA viruses; nucleocapsid protein