Zajakina Anna, Vasilevska Jelena, Zhulenkovs Dmitry, Skrastina Dace, Spaks Artjoms, Plotniece Aiva, Kozlovska Tatjana
Department of Cell Biology, Biomedical Research and Study Centre, Ratsupites Str,, 1, Riga LV-1067, Latvia.
BMC Cancer. 2014 Jun 20;14:460. doi: 10.1186/1471-2407-14-460.
The combination of virotherapy and chemotherapy may enable efficient tumor regression that would be unachievable using either therapy alone. In this study, we investigated the efficiency of transgene delivery and the cytotoxic effects of alphaviral vector in combination with 5-fluorouracil (5-FU) in a mouse mammary tumor model (4 T1).
Replication-deficient Semliki Forest virus (SFV) vectors carrying genes encoding fluorescent proteins were used to infect 4 T1 cell cultures treated with different doses of 5-FU. The efficiency of infection was monitored via fluorescence microscopy and quantified by fluorometry. The cytotoxicity of the combined treatment with 5-FU and alphaviral vector was measured using an MTT-based cell viability assay. In vivo experiments were performed in a subcutaneous 4 T1 mouse mammary tumor model with different 5-FU doses and an SFV vector encoding firefly luciferase.
Infection of 4 T1 cells with SFV prior to 5-FU treatment did not produce a synergistic anti-proliferative effect. An alternative treatment strategy, in which 5-FU was used prior to virus infection, strongly inhibited SFV expression. Nevertheless, in vivo experiments showed a significant enhancement in SFV-driven transgene (luciferase) expression upon intratumoral and intraperitoneal vector administration in 4 T1 tumor-bearing mice pretreated with 5-FU: here, we observed a positive correlation between 5-FU dose and the level of luciferase expression.
Although 5-FU inhibited SFV-mediated transgene expression in 4 T1 cells in vitro, application of the drug in a mouse model revealed a significant enhancement of intratumoral transgene synthesis compared with 5-FU untreated mice. These results may have implications for efficient transgene delivery and the development of potent cancer treatment strategies using alphaviral vectors and 5-FU.
病毒疗法与化疗相结合可能实现有效的肿瘤消退，而单独使用任何一种疗法都无法做到这一点。在本研究中，我们在小鼠乳腺肿瘤模型（4T1）中研究了转基因递送效率以及α病毒载体与5-氟尿嘧啶（5-FU）联合使用的细胞毒性作用。
携带编码荧光蛋白基因的复制缺陷型塞姆利基森林病毒（SFV）载体用于感染用不同剂量5-FU处理的4T1细胞培养物。通过荧光显微镜监测感染效率，并通过荧光测定法进行定量。使用基于MTT的细胞活力测定法测量5-FU与α病毒载体联合治疗的细胞毒性。在皮下4T1小鼠乳腺肿瘤模型中进行体内实验，使用不同剂量的5-FU和编码萤火虫荧光素酶的SFV载体。
在5-FU处理之前用SFV感染4T1细胞未产生协同抗增殖作用。另一种治疗策略是在病毒感染之前使用5-FU，这强烈抑制了SFV表达。然而，体内实验表明，在用5-FU预处理的4T1荷瘤小鼠中，瘤内和腹腔内给予载体后，SFV驱动的转基因（荧光素酶）表达显著增强：在此，我们观察到5-FU剂量与荧光素酶表达水平之间呈正相关。
虽然5-FU在体外抑制了4T1细胞中SFV介导的转基因表达，但与未用5-FU处理的小鼠相比，在小鼠模型中应用该药物显示瘤内转基因合成显著增强。这些结果可能对使用α病毒载体和5-FU进行有效的转基因递送以及开发有效的癌症治疗策略具有重要意义。
