Kievit E, Nyati M K, Ng E, Stegman L D, Parsels J, Ross B D, Rehemtulla A, Lawrence T S
Department of Radiation Oncology, University of Michigan, Ann Arbor 48109-0010, USA.
Cancer Res. 2000 Dec 1;60(23):6649-55.
The efficacy of cancer gene therapy using bacterial cytosine deaminase (bCD)/5-fluorocytosine (5-FC) enzyme/prodrug strategy is limited by the inefficiency of cytosine deaminase (CD)-catalyzed conversion of 5-FC into 5-fluorouracil (5-FU). We have shown previously that yeast CD (yCD) is more efficient at the conversion of 5-FC than bCD. In the current study, we hypothesized that the increased production of 5-FU by yCD would enhance the efficacy of the CD/5-FC treatment strategy by increasing the bystander effect as well as the efficacy of radiotherapy because of the radiosensitizing capacity of 5-FU. To test this hypothesis, we generated stable HT29 human colon cancer cell lines expressing either bCD (HT29/bCD) or yCD (HT29/yCD). The amount of 5-FU produced in HT29/yCD tumors after a single injection of 5-FC (1000 mg/kg, i.p.) was 15-fold higher than that produced in HT29/bCD tumors. In tumor-bearing nude mice, the average minimum relative tumor size (compared with pretreatment values) of HT29/bCD tumors treated with 5-FC and radiation (500 mg/kg i.p. and 3 Gy, 5 days a week for 2 weeks) was 0.55+/-0.1, compared with 0.01+/-0.01 in HT29/yCD tumors (P = 0.002). Moreover, an increased cytotoxic and radiosensitizing effect of 5-FC on bystander cells was observed in vitro and in vivo when yCD was expressed in HT29 cells instead of bCD. In mice bearing HT29 tumors containing 10% HT29/yCD cells, the combined treatment resulted in a minimum tumor size of 0.20+/-0.07 compared with 0.60+/-0.1 in 10% HT29/bCD cells (P < 0.001). These results demonstrate that the use of yCD in the CD/5-FC strategy has a high potential to improve the therapeutic outcome of combined gene therapy and radiotherapy in cancer patients.
使用细菌胞嘧啶脱氨酶（bCD）/5-氟胞嘧啶（5-FC）酶/前药策略进行癌症基因治疗的疗效受到胞嘧啶脱氨酶（CD）催化5-FC转化为5-氟尿嘧啶（5-FU）效率低下的限制。我们之前已经表明，酵母CD（yCD）在5-FC转化方面比bCD更有效。在当前研究中，我们假设yCD增加5-FU的产生将通过增强旁观者效应以及由于5-FU的放射增敏能力而提高放疗效果，从而增强CD/5-FC治疗策略的疗效。为了验证这一假设，我们构建了稳定表达bCD（HT29/bCD）或yCD（HT29/yCD）的HT29人结肠癌细胞系。单次注射5-FC（1000 mg/kg，腹腔注射）后，HT29/yCD肿瘤中产生的5-FU量比HT29/bCD肿瘤中产生的高15倍。在荷瘤裸鼠中，用5-FC和放疗（500 mg/kg腹腔注射和3 Gy，每周5天，共2周）治疗的HT29/bCD肿瘤的平均最小相对肿瘤大小（与治疗前值相比）为0.55±0.1，而HT29/yCD肿瘤为0.01±0.01（P = 0.002）。此外，当HT29细胞中表达yCD而非bCD时，在体外和体内均观察到5-FC对旁观者细胞的细胞毒性和放射增敏作用增强。在含有10% HT29/yCD细胞的HT29肿瘤荷瘤小鼠中，联合治疗导致最小肿瘤大小为0.20±0.07，而在含有10% HT29/bCD细胞的小鼠中为0.60±0.1（P < 0.001）。这些结果表明，在CD/5-FC策略中使用yCD具有很大潜力来改善癌症患者联合基因治疗和放疗的治疗效果。
