Saturday, February 18, 2012
Exhibit Hall A-B1 (VCC West Building)
Background: Myocarditis is a major heart disease in infants and young adults. It is commonly caused by coxsackievirus B3 (CVB3) infection. However, no specific treatment or vaccine is available currently. RNA interference (RNAi)-based antiviral therapy is a promising strategy with great potential to inhibit viral replication. However, viral mutational escape from drug suppression and the targeted delivery of the reagents to specific cell populations are hampering the further development of this treatment. Methods: In this study, we designed artificial microRNAs (AmiRs) targeting the 3' untranslated region (3'UTR) of CVB3 genome with mismatches to the central region of their targeting sites. To achieve specific delivery, we then linked AmiRs to the folate-conjugated bacterial phage packaging RNA (pRNA) and delivered the complexes without the need of transfection into HeLa cells, a widely used cell line as an in vitro model for CVB3 infection, via folate-mediated specific internalization. Results: Antiviral evaluation showed that vector based expression of AmiR-1 and AmiR-2 suppressed CVB3 replication approximately 100-fold in infected cells. After conjugation with pRNA, the designed AmiRs were still correctly processed and preserved the anti-viral function. We further demonstrated that pRNA-AmiR delivered through folate mediated internalization was successful with similar inhibitory effect on CVB3 replication as the ones delivered by transfection. By mutational analyses, we found that our designed pRNA-AmiRs conjugates were tolerable to target mutations and have great potential to suppress viral mutational escape. Little effect on triggering interferon induction was observed in our designed pRNA-AmiR. Conclusion/Significance: This study provides important clues for designing AmiRs targeting the 3'UTR of viral genome. It also proves the feasibility of specific deliver of miRNA mimics using conjugated pRNA vehicles through receptor and its legend interactions. These small AmiRs combined with pRNA-folate conjugates could form a promising system for antiviral drug development.