Experimental study of thymidine kinase gene therapy of neuroblastoma in vitro and in vivo

摘要

Neuroblastoma arises as a direct result of genetic disorder; therefore, it should be well treated and conquered by gene therapy in future. In this study, neuroblastoma cell line SH-SY5Y experiments, in vitro and in nude mice in vivo, were subjected to research thymidine kinase suicide gene to treat neuroblastoma. The plasmid LXpsp-hytk and a plasmid LXSH were transduced separately by lipofectin into human neuroblastoma cell line SH-SY5Y. SH-SY5Y-hy and SH-SY5Y-hytk were selected by hygromycin B. Different ganciclovir (GCV) concentration was given to SH-SY5Y-hytk to determine optimal GCV concentration. The cytotoxic effect of GCV on SH-SY5Y-hytk, SH-SY5Y-hy, and SH-SY5Y cells was determined. Scapular subcutaneous tumors were established in nude mice by inoculating 2.5×106 SH-SY5Y-hytk on their left sides and 2.5×106 SH-SY5Y-hy cells on their right sides for every mouse of treatment group and control group, respectively. After 1 week, mass grew in both sides of all the mice, and from the eighth day on, every mouse in treatment group received daily intraperitoneal injection of GCV 50 mg/kg body weight for 14 days; every mouse in control group received daily intraperitoneal injection of 1 ml saline for 14 days. On day 22 tumors were excised and weighed on the left and right sides, respectively, and apoptosis was detected by TUNEL method. Apoptotic index was calculated on the left and on the right sides, respectively, for every mouse in treatment group and control group. The lowest concentration of hygromycin B was 60 μg/ml. The cytotoxic effect of GCV on SH-SY5Y-hytk cells was obvious (IC50=0.03 μM), whereas GCV showed almost no cytotoxic effect on SH-SY5Y and SH-SY5Y-hy cells (IC50