腺病毒介导融合双自杀基因治疗膀胱癌

Adenovirus-mediated double suicide gene therapy for experimental bladder carcinoma

目的探讨腺病毒介导胞嘧啶脱氨酶(CD)和胸苷激酶(TK)融合双自杀基因系统对膀胱癌治疗作用并与单基因系统作比较。方法利用含有CD-TK双自杀融合基因及绿色荧光蛋白(GFP)基因的复制缺陷腺病毒作载体,PCR检测CD、TK及E1基因。建立C57BL/6同系膀胱癌Mb49皮下移植瘤模型,观察肿瘤注射腺病毒联合丙氧鸟苷(GCV)或/和5-氟胞嘧啶(5-FC)治疗后肿瘤体积及组织学变化。结果PCR可检测到腺病毒DNA中含CD及TK基因、无E1基因。腺病毒注射联合GCV、5-FC、GCV+5-FC治疗后,肿瘤体积与对照组相比明显缩小(P=0.00),腺病毒注射联合GCV+5-FC有协同作用(P=0.04),优于腺病毒注射联合GCV以及腺病毒注射5-FC。基因治疗后肿瘤细胞大片坏死,对照组细胞形态无变化。结论腺病毒介导CD-TK融合双自杀基因联合GCV或/和5-FC能有效治疗膀胱癌,融合双自杀基因CD-TK/(GCV+5-FC)系统对膀胱癌治疗有协同作用,其效果优于CD-TK/GCV或CD-TK/5-FC系统。

Objective To evaluate the feasibility and efficacy of cytocine deaminase-thymidine kinase （CD-TK） fusion double suicide gene therapy using adenovirus mediated CD-TK gene and green fluorescent rotein （GFP） gene combined with ganciclovir（GCV） or 5-flourocytosine（5-FC） in a murine subcutaneous bladder carcinoma model. Methods A replication defective adenovirus vector containing CD-TK gene was used. Subcutaneous tumors were established in syngenic C57BL/6 female mice with 1 ×10^6 Mb49 cells. Intratumoral injection of AdCD-TK （1.58×10^8 PFU, qd^days） in combination with GCV （40 mg·kg^-t·d^-1, ip, qd×10 days） or 5-FC （400 mg·kg^-t·d^-1, ip, qd×10 days） was administered in vivo for the determination of treatment efficacy in separate controlled experiments. Results In vivo experiments demonstrated that the mean volume of tumor in the group of AdCD-TK/GCV （326.58 ±109.56 mm^3）, AdCD-TK/5-FC （235.33 ±62.94 mm^3） and AdCD-TK/ （GCV＋5-FC） （23.58±6.78 mm^3） was reduced significantly compared with that of control group （993.51±158.32 mm^3） （P=0. 00）, the mean volume of tumor in the group of AdCD-TK/ （GCV＋5-FC） was significantly less than that in the group of AdCD-TK/GCV or AdCD-TK/5-FC （P=0.04）. Tumor necrosis was revealed by histomorphology compared with control animals. Conclusions Adenovirus mediated CD-TK double suicide gene combining with GCV or 5-FC could provide an effective therapy in an experimental murine bladder carcinoma by significantly inhibiting tumor growth. The treatment efficacy of AdCD-TK combining GCV and 5-FC was superior to that of AdCD-TK combining GCV or AdCD-TK combining 5-FC.