A novel HBV antisense RNA gene delivery system targeting hepatocellular carcinoma.
OBJECTIVE
To construct a novel HBV antisense RNA delivery system targeting hapatocellular carcinoma and study its inhibitory effect in vitro and in vivo.


METHODS
GE7,a 16-peptide specific to EGFR, and HA20,a homologue of N-terminus of haemagglutinin of influenza viral envelope protein, were synthesized and conjugated with polylysin. The above conjugates were organized into the pEBAF-as-preS2, a hepatocarcinoma specific HBV antisense expression vector, to construct a novel HBV antisense RNA delivery system, named AFP-enhancing 4-element complex. Hepatocelluar carcinoma HepG2.2.15 cells was used to assay the in vitro inhibition of the complex on HBV. Expression of HBV antigen was assayed by ELISA. BALB/c nude mice bearing HepG2.2.15 cells were injected with AFP-enhancing 4-element complex. The expression of HBV antisense RNA was examined by RT-PCR and the size of tumor in nude mice were measured.


RESULTS
The AFP-enhancing 4-element complex was constructed and DNA was completely trapped at the slot with no DNA migration when the ratio of polypeptide to plasmid was 1:1. The expression of HBsAg and HBeAg of HepG2.2.15 cells was greatly decreased after being transfected by AFP-enhancing 4-element complex. The inhibitory rates were 33.4 % and 58.5 % respectively. RT-PCR showed HBV antisense RNA expressed specifically in liver tumor cells of tumor-bearing nude mice. After 4 injections of AFP-enhancing 4-element complex containing 0.2 micro g DNA, the diameter of the tumor was 0.995 cm+/-0.35, which was significantly smaller than that of the control groups(2.215 cm+/-0.25, P<0.05).


CONCLUSIONS
AFP-enhancing 4-element complex could deliver HBV antisense RNA targeting on hepatocarcinoma and inhibit both HBV and liver tumor cells in vitro and in vivo.
Ma
Chun-Hong
CH
Institute of Immunology,Medical College of Shandong University, Jinan 250012, Shandong Province, China.
Sun
Wen-Sheng
WS
Tian
Pei-Kun
PK
Gao
Li-Fen
LF
Liu
Su-Xia
SX
Wang
Xiao-Yan
XY
Zhang
Li-Ning
LN
Cao
Ying-Lin
YL
Han
Li-Hui
LH
Liang
Xiao-Hong
XH
D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000235:genetics*; D018014:Gene Transfer Techniques; D006515:Hepatitis B virus / Q000235:genetics*; D008113:Liver Neoplasms / Q000235:genetics*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D016372:RNA, Antisense; D012367:RNA, Viral / Q000235:genetics*; D014407:Tumor Cells, Cultured
D016372:RNA, Antisense; D012367:RNA, Viral