Optimal design and validation of antiviral siRNA for targeting hepatitis B virus.
OBJECTIVE
Optimal design of antiviral short-interfering RNA (siRNA) targeting highly divergent hepatitis B virus (HBV) was validated by quantitative structure activity relationship (QSAR) analysis.


METHODS
The potency of 23 synthetic siRNAs targeting 23 sites throughout HBV pregenomic RNA were evaluated at 10 nmol/L by determining the inhibition on the expression of S/P/pregenomic mRNA and hepatitis B surface antigen (HBsAg) quantitatively in HepG2.2.15 cells. Genotype homology within HBV genomes was identified through plentiful computational analysis and the multiple linear regression analysis was made to validate the relationship between the functional siRNAs and primary characteristics. Based on the preliminary results, relationships between different determined endpoints [S/P mRNA, HBsAg, C/P mRNA, hepatitis B e antigen (HBeAg) and viral DNA load] and siRNA efficacy evaluation were investigated.


RESULTS
Genotype homology, open reading frame (ORF) S/P, X and C had tight correlation with the ability of siRNAs on inhibiting the expression of S/P/Pregenomic mRNA and HBsAg (P<0.01), of which, ORF C was negatively correlated with the siRNA potency (P<0.05). Further study showed that siRNA potency evaluation was influenced by different determined endpoints. P-target siRNAs showed significant inhibition on the S mRNA and HBsAg expression. S-target siRNAs inhibited the expression of S mRNA and HBsAg strongly. X-target siRNAs played active roles in inhibiting all 5 determined endpoints. C-target siRNAs blocked the expression of C mRNA, HBeAg and viral DNA load significantly.


CONCLUSIONS
The antiviral potency of siRNA was relevant to its primary characteristics and determined endpoints were important for siRNA efficacy evaluation for complex genome with overlapping ORF, which was helpful for siRNA optimal design.
Fu
Jie
J
Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing 100850, China.
Tang
Zhong-ming
ZM
Gao
Xin
X
Zhao
Fan
F
Zhong
Hui
H
Wen
Mao-rong
MR
Sun
Xiao
X
Song
Hai-feng
HF
Qian
Xiao-hong
XH
D000818:Animals; D000998:Antiviral Agents; D002460:Cell Line; D016679:Genome, Viral; D005838:Genotype; D006514:Hepatitis B Surface Antigens / Q000235:genetics / Q000378:metabolism; D006513:Hepatitis B e Antigens / Q000235:genetics / Q000378:metabolism; D006515:Hepatitis B virus / Q000235:genetics*; D006801:Humans; D016366:Open Reading Frames; D021281:Quantitative Structure-Activity Relationship; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results
D000998:Antiviral Agents; D006514:Hepatitis B Surface Antigens; D006513:Hepatitis B e Antigens; D034741:RNA, Small Interfering