Antisense down-regulation of 5-FU related enhances cytotoxicity of 5-FU in human lung cancer

胸苷酸合成酶（TS）是胸苷从头合成的关键酶，是5-FU抗肿瘤化疗的重要靶点。我们用反义寡核苷酸（ODN）下调TS的表达。我们制备了5个与TSmRNA互补的反义ODN和1个随机ODN。使用HVJ-脂质体进行ODNs的转染。将HVJ-脂质体与人肺癌细胞株Ma 10和Ma 25及反义ODNs共同孵育后，用RT-PCR方法检测细胞株TS的表达，并测定TS活性。TS表达被抑制50%至20%，通过温育细胞系和反义ODNs和TS表达。TS的反义ODNs不可能抑制TS的表达。Chu等报道了TS蛋白本身在翻译起始位点和编码区都与T mRNA结合，抑制信息的翻译加工。明年，我们研究了脂质体结合的CDDP（抗癌药物）（脂质体-CDDP）是否增强抗癌效果。顺铂对人肺癌细胞株Ma 44 -3的IC_（50）为40 μ g/ML。脂质体-顺铂孵育48小时的IC_（50）为10 μ g/mL。孵育96小时，浓度为1.5 μ g/ML。随着孵育时间的延长，脂质体-顺铂的IC_（50）逐渐降低。罗丹明标记的脂质体-CDDP在体外可与癌细胞胞浆结合。罗丹明标记的脂质体-CDDP显示脂质体-CDDP在体外与癌细胞胞浆结合。罗丹明标记的脂质体-CDDP使用雾化器（通过气道）在SCID小鼠中施用。在小鼠肺中观察到罗丹明标记的脂质体。这些结果表明脂质体-CDDP经气道给药可能成为肺癌的治疗方法。

Thymidylate synthase (TS), the key enzyme in de novo synthesis of thymidine, is an important target for antitumor chemothrapy of 5-FU. We used antisense oligodeoxynucleotide (ODN) to down-regulate the expression of TS. We made 5 antisense ODNs which is complementary to TS Mrna, and one random ODN. Transfection of ODNs was performed using HVJ-liposome. After incubating human lung cancer cell lines (Ma10 and Ma25) and antisense ODNs with HVJ-liposome, the TS expression of cell lines was examined by RT-PCR method and TS activity was measured. TS expression was repressed from 50% to 20% by incubaating cell lines and antisense ODNs and TS expression. It was impossible that antisense ODNs of TS inhibited the expression of TS. Chu e al. reported that the TS protein itself binds to the T Mrna both at the translational start site and in the coding region, inhibiting translational processing of the message.Next year, we examined whether CDDP (anticancer drug) conjugated by liposome (liposome-CDDP) enhance anticancer effect. The IC50 of CDDP was 40ug/ML in human lung cancer cell line Ma44-3 in vitro. Meanwhile, the IC50 of liposome-CDDP was 10ug/ML for 48 hr incubation. And it was 1.5ug/ML for 96 hr incubation. When thecubation time becomes longer, the IC50 of liposome-CDDP was lower. The use of Rhodamine-labeled liposome-CDDP revealed that the lipsome-CDDP attached the cytoplasm of cancer cells in vitro. Rhodamine-labeled liposome-CDDP revealed that the liposome-CDDP attached the cytoplasm of cancer cells in vitro. Rhodamine-labeled liposome-CDDP was administrared in SCID mice using a nebulizer (through airway). Rhodamine-labeled liposome was observed in the lung of the mice. These results demonstrate that the administration of liposome-CDDP through airway may become a therapy for lung cancer.