A novel HBV antisense RNA gene delivery system targeting hepatocellular carcinoma

AIM: 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. RTPCR showed HBV antisense RNA expressed specifically in liver tumor cells of tumor-bearing nude mice. After 4injections of AFP-enhancing 4-element complex containing 0.2 μg 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).CONCLUSION: 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.

A Multicenter Comparative Clinical Study of Sino-Levonorgestrel-Releasing Implants— No. I and No. II withNorplant

Objective To compare the efficacy, side effects and acceptability of two Sino-implants with Norplant Methods A randomized, prospective multicenter comparative clinical study was conducted in 10 clinical centers in China. Results Totally 1 001, 1 000 and 998 cases were recruited for Sino-implant No. I, No. II and Norplant, respectively, in 1993. The follow-up rate was 99.8% totally in 5 years. Three and five pregnancies occurred respectively in Sino-implant No. I and No. II group, while no pregnancy occurred in the Norplant group. The cumulative pregnancy rate was 0.4, 0.7 and 0 per 100 women respectively in implant No.I, No. II and Norplant group for five years, meaning that there was no statistical difference. There was no ectopic pregnancy in the three groups. The cumulative discontinuation rates at the end of five years were not significantly different among the three groups, either. Men- strual problems were the main reason for termination. The menstrual blood loss decreased significantly and no serious health problems arose fromimplants use. Conclusion The two Sino-implants provided similar high efficacy and safety to Norplant, therefore, they can be used by women at reproductive age who are from different areas,different of races, educational background and occupation.

Effect of NS-398 on colon cancer cells

AIM: To study the effect of NS-398, a selective cydooxygenase 2 (COX-2) inhibitor, on invasion of colon cancer cell line HT-29 in vitro and to explore its mechanisms. METHODS: Invasive behaviors of the malignant colon cancer cell line HT-29 were investigated in this study. Expressions of COX-2 and CD44v6 in HT-29 cells were detected by flow cytometry. Cellular survival rate was determined by MTT assay. The invasive capacity was quantified by a modified Boyden chamber model. Alterations of cytoskeleton component F-actin were observed by confocal laser scanning microscope. RESULTS: Flow cytometry analysis showed that COX-2 was highly expressed in HT-29 cells. The invasive capability of HT-29 cells could be greatly inhibited by NS-398 at the experimental concentrations of 0.1,1.0 and 10 μmol/L with an inhibitory rate of 22.74%, 42.35% and 58.61% (P<0.01), respectively. MTT assay showed that NS-398 at the experimental concentrations had no significant influence on cellular viability, indicating that such anti-invasive effects had no relationship with cytotoxicity. F-actin was mainly distributed around nuclei forming annular structure in HT-29 cells. After exposure to NS-398 of 10 μmol/L, the annular structure around nuclei disappeared and the fluorescence intensity of F-actin decreased obviously. Treatment with NS-398 could down-regulate the expression of CD44v6 as well. CONCLUSION: NS-398 has anti-invasive effects on colon cancer HT-29 cells in vitro, which may be mediated by a novel mechanism of disruption of cytoskeleton. Down-regulation of CD44v6 expression may be related to alterations of cytoskeleton.