Prophylactic and therapeutic effects of small interfering RNA targeting SARS-coronavirus.
OBJECTIVE
To identify and characterize the siRNA duplexes that are effective for inhibition of SARS-CoV infection and replication in the non-human primate cells. This in vitro study will serve as the foundation for development of novel anti-SARS therapeutics.


METHODS
48 siRNA sequences were designed for targeting regions throughout entire SARS-CoV genome RNA including open-reading frames for several key proteins. Chemically synthesized siRNA duplexes were transfected into foetal rhesus kidney (FRhK-4) cells prior to or after SARS-CoV infection. The inhibitory effects of the siRNAs were evaluated for reductions of intracellular viral genome copy number and viral titres in the cell culture medium measured by Q-RT-PCR and CPE-based titration, respectively. Four siRNA duplexes were found to achieve potent inhibition of SARS-CoV infection and replication. A prolonged prophylactic effect of siRNA duplexes with up to 90% inhibition that lasted for at least 72 h was observed. Combination of active siRNA duplexes targeting different regions of the viral genome resulted in therapeutic activity of up to 80% inhibition.


CONCLUSIONS
Chemically synthesized siRNA duplexes targeting SARS-CoV genomic RNA are potent agents for inhibition of the viral infection and replication. The location effects of siRNAs were revealed at both genome sequence and open-reading frame levels. The rapid development of siRNA-based SARS-CoV inhibitors marked a novel approach for combating newly emergent infectious diseases.
Zheng
Bo-jian
BJ
Department of Microbiology, Institute of Molecular Biology, Department of Pathology, University of Hong Kong, Hong Kong, China.
Guan
Yi
Y
Tang
Qingquan
Q
Du
Cheng
C
Xie
Frank Y
FY
He
Ming-Liang
ML
Chan
Kwok-Wah
KW
Wong
Kin-Ling
KL
Lader
Eric
E
Woodle
Martin C
MC
Lu
Patrick Y
PY
Li
Baojian
B
Zhong
Nanshan
N
D000818:Animals; D000998:Antiviral Agents / Q000138:chemical synthesis; D002460:Cell Line; D003470:Culture Media; D003588:Cytopathogenic Effect, Viral / Q000502:physiology*; D015316:Genetic Therapy; D016679:Genome, Viral; D008253:Macaca mulatta; D034741:RNA, Small Interfering / Q000138:chemical synthesis / Q000235:genetics*; D012367:RNA, Viral / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D013997:Time Factors; D014162:Transfection; D014779:Virus Replication / Q000235:genetics
D000998:Antiviral Agents; D003470:Culture Media; D034741:RNA, Small Interfering; D012367:RNA, Viral