Experimental Study on PEG10 Antisense Gene Therapy for Hepatocellular Carcinoma Mediated by Nanoscale Ultrasound Contrast Agent

Abstract

Objective: To explore the application of nanoscale ultrasound contrast agent mediating PEG10 antisense gene transfection in the treatment of hepatocellular carcinoma. Methods: The optimal concentration and ultrasound parameters of nanoscale ultrasound contrast agent were screened via cell viability assay. PEG10 antisense oligodeoxynucleotide (ASODN) and fluorescein-labeled specific ligand galactose terminal glycol-poly-L-lysine (G-PLL) were combined with nanoscale ultrasound contrast agent. Then PEG10 ASODN was transfected into human HepG2 cells under ultrasonic irradiation. RT-PCR was used to detect the expression of PEG10. MTT assay was conducted to assess the proliferation inhibition of hepatocellular carcinoma cells. And the tumor inhibition rate in vivo was analyzed by establishing the nude mice transplantation tumor model. Results: Of all the conjugates, the optimal molar ratio of PEG10 ASODN to G-PLL was 1:2. The best parameters after screening showed that nanoscale ultrasound contrast agent of 15% microbubble concentration and ultrasound exposure at 1.5 MHz, 1.0 W/cm 2 , and 10% duty cycle for 60 s had no significant influence on the viability of HepG2 cells (P > 0 05). Compared with other groups, PEG10 ASODN+G-PLL+microbubble group had the weakest PEG10 expression, the most significant cell growth inhibition, and the highest tumor inhibition rate up to 38.5%. Conclusion: Under ultrasonic irradiation, G-PLL conjugating nanoscale microbubble ultrasound contrast agent and antisense gene can effectively enhance the inhibition of antisense gene on PEG10 expression, which has targetedly inhibitory effect on hepatoma cells. This is expected to provide an efficient means for the treatment of hepatocellular carcinoma.