Schmittgen T D, Koolemans-Beynen A, Webb T E, Rosol T J, Au J L
Division of Pharmaceutics and Pharmaceutical Chemistry, College of Pharmacy, Ohio State University, Columbus 43210.
Cancer Chemother Pharmacol. 1992;30(1):25-30. doi: 10.1007/BF00686481.
In a previous study, we showed that 5-fluorouracil (FU) is active against the dimethylhydrazine-induced colon tumor in rats; a 7-day infusion of FU at 30 mg/kg daily produced 85% tumor-free cures. The present study examined the effects of FU alone and in combination with leucovorin (LV) or D-glucarate (GT) using an ex vivo system that maintained the growth of the rat colon-tumor explants on collagen gels. The labeling index (LI) was determined by the incorporation of [3H]-thymidine and autoradiography. The mean LI of the untreated control was 64.8% +/- 19.8%. The IC50, IC90, and IC95 values following a 7-day exposure to FU were 0.36, 0.75, and 1.22 microM, respectively. In comparison, the steady-state FU concentration required to produce 67% tumor-free cures in rats following a 7-day infusion is 1.54 microM. LV alone did not produce any antiproliferative effect at concentrations as high as 10 microM. The addition of LV at concentrations of 0.001-10 microM did not significantly reduce the IC50 of FU. The lack of effect of LV may have been due to tissue saturation with folate provided in the culture medium. GT alone reduced the tumor LI by 20%-30% at concentrations of 0.1-10 microM. GT enhanced the effect of FU. As compared with FU alone, the addition of GT at concentrations of 0.1 and 1.0 microM reduced the IC50 of FU by 47% and 60% to 0.21 and 0.16 microM, respectively. Assessment of the potentiation of the inhibitory effect of FU by GT using two-way analysis of variance and the isobologram method indicated a significant synergistic interaction between FU and GT. This interaction occurred within the FU concentration range of 0.08 and 0.4 microM. In summary, these data indicate that (a) the IC values for FU are comparable in tumor explants and in rats, suggesting that the effects in cultured tumors reflect those in intact animals; (b) GT alone showed antitumor activity, albeit relatively minor as compared with FU; (c) FU and GT exhibited synergistic activity, which was most pronounced at FU concentrations that produced submaximal activity (less than 30% inhibition of tumor LI); and (d) GT and LV had different effects on the growth inhibition by FU, suggesting that GT acts by a mechanism different from the thymidylate synthase-directed effect of FU and LV.
在先前的一项研究中，我们发现5-氟尿嘧啶（FU）对二甲基肼诱导的大鼠结肠肿瘤具有活性；每天以30mg/kg的剂量连续输注FU 7天可使85%的大鼠实现无瘤治愈。本研究使用一种能维持大鼠结肠肿瘤外植体在胶原凝胶上生长的体外系统，检测了FU单独使用以及与亚叶酸（LV）或D-葡萄糖醛酸（GT）联合使用的效果。通过掺入[3H]-胸腺嘧啶核苷及放射自显影来测定标记指数（LI）。未处理对照组的平均LI为64.8%±19.8%。暴露于FU 7天后的IC50、IC90和IC95值分别为0.36、0.75和1.22μM。相比之下，在连续输注7天后，大鼠体内产生67%无瘤治愈率所需的稳态FU浓度为1.54μM。单独使用LV时，即使浓度高达10μM也未产生任何抗增殖作用。添加浓度为0.001 - 10μM的LV并未显著降低FU的IC50。LV无作用可能是由于培养基中提供的叶酸使组织达到饱和。单独使用GT时，在浓度为0.1 - 10μM时可使肿瘤LI降低20% - 30%。GT增强了FU的作用。与单独使用FU相比，添加浓度为0.1和1.0μM的GT可使FU的IC50分别降低47%和60%，至0.21和0.16μM。使用双向方差分析和等效线图法评估GT对FU抑制作用的增强表明，FU与GT之间存在显著的协同相互作用。这种相互作用发生在FU浓度范围为0.08至0.4μM时。总之，这些数据表明：（a）FU在肿瘤外植体和大鼠体内的IC值具有可比性，这表明在培养肿瘤中的作用反映了在完整动物中的作用；（b）单独使用GT显示出抗肿瘤活性，尽管与FU相比相对较小；（c）FU和GT表现出协同活性，在产生次最大活性（对肿瘤LI的抑制小于30%）的FU浓度下最为明显；（d）GT和LV对FU生长抑制的影响不同，这表明GT的作用机制不同于FU和LV的胸苷酸合成酶导向作用。
