人端粒酶逆转录启动子驱动的单纯疱疹病毒胸苷激酶基因治疗裸鼠胰腺移植瘤的安全性研究  

The safety of HSV-TK gene under the control of human telomerase transcriptase gene Promoter on pancreatic carcinoma xenograft in nude mice

作　　者：谢于[1] 李德春[1] 张子详[1] 朱兴国[1] 王凤力[1] 

机构地区：[1]苏州大学附属第一医院普外科,215006

出　　处：《中华实验外科杂志》2007年第4期391-394,共4页Chinese Journal of Experimental Surgery

基　　金：卫生部科学研究基金资助项目(WKJ2004-2-001)

摘　　要：目的探讨人端粒酶逆转录(hTERT)启动子驱动的单纯疱疹病毒胸苷激酶(HSV- TK)基因治疗裸鼠胰腺癌的安全性。方法应用细菌内同源重组法构建hTERT启动子和常规巨细胞病毒(CMV)启动子驱动的HSV-TK基因真核表达载体PDC312-TP-TK,PSU-CMV-TK。制备腺病毒液,检测病毒滴度为1.9×10^(10)pfu/ml和1.4×10^(10)pfu/ml。建立裸鼠胰腺癌皮下移植瘤模型,分别以两种方法治疗后观察皮下移植瘤消退、HSV-TK基因表达情况、肝脏苏木素-伊红(HE)染色切片、肝功能。结果PDC312-TP-TK组与PSU-CMV-TK组对肿瘤的抑瘤率分别为58.3%和65.6%,治疗组间差异无统计学意义(P>0.05),杀伤方面具有一样的高效性;但其对肝脏的毒副作用远小于PSU-CMV-TK组,PDC312-TP-TK组血清中ALT,AST,ALP分别为(54.00±1.97)、(147.00±12.67)、(33.00±2.59)U/L而PSU-CMV-TK组中为(334.00±9.81)、(511.00±11.24)、(98.00±3.12)。差异有统计学意义(P<0.01)。裸鼠肝脏苏木素-伊红(HE)染色中,仅PSU-CMV-TK组有肝脏坏死改变。逆转录-聚合酶链反应(RT-PCR)中也仅PSU-CMV-TK组检测到1143 bp的TK基因片段。结论hTERT启动子驱动的HSV-TK基因是一种新的靶向治疗方法,在具有治疗高效性的同时,更具有安全性。Objective To evaluate the safety of HSV-TK gene under the control of human telomerase transcriptase gene promoter on pancreatic carcinoma xenograft in nude mice. Methods An expression vector （PDC312-TP-TK） containing TK gene under the hTERT promoter was constructed by the homologous recombination method in bacteria. The newly recombinated PDC312-TP-TK was propagated in 293 cells and purified by cesium chloride gradient centrifugation. Positive clone were selected by using endonuclease to digest the recombinants and the concentration of viral liquid containing TK gene was 1.9 × 10^10pfu/ml. And the same we got PSU-CMV-TK and the concentration of viral liquid containing TK gene was 1.4×10^10pfu/ ml. An orthotopic human pancreatic adenooarcinoma was successfully established subcutaneously in the flank of nude mice. Two methods were applied and the antitumor effect ,the expression of HSV-TK gene ,liver function,and liver HE section were observed. Result The PDC312-TP-TK had the same efficacy as the PSU- CMV-TK on antitumor and their tumor growth inhibition rate was 58.3% and 65.6% respectively and there was no significant difference between them （P 〉 0.05）. But its hepatotoxicity was lower than the PSU-CMV- TK. The ALT,AST and ALP in PD312 - TP - TK group were 54.00 ± 1.97, 147.00 ±12.67 and 33.00 ± 2.59 respectively ,while they were 334.00±9.81,511.00± 11.24 and 98.00 ± 3.12 respectively in PSU- CMV-TK group （P 〈 0.01 ）. Nude mice liver sections were stained with hematoxylineosin （HE） and only those rats treated with PSU-CMV-TK had hepatic necrosis, HSV-TK gene was detected in liver. Conclusion The telomerase specific transfer of the TK gene under the hTERT promoter is a novel targeting approach for the treatment of pancreatic carcinoma and more efficient and safe.

关 键 词：胰腺癌 端粒酶 启动子 基因治疗 

分 类 号：R735.9[医药卫生—肿瘤]