Futscher B W, Erickson L C
Department of Pharmacology, Loyola University of Chicago, Stritch School of Medicine, Maywood, Illinois 60153.
Cancer Res. 1990 Jan 1;50(1):62-6.
This study was initiated to determine if DNA-damaging chemotherapeutic agents can suppress the expression of oncogenes. The effects of three structurally related bifunctional alkylating agents on the steady state mRNA levels of c-myc, c-fos, N-ras, and beta-actin in the human colon carcinoma cell line Colo320HSR were examined. Colo320HSR has an amplified c-myc oncogene, which is highly overexpressed, and is assumed to be one of the transforming genes of this cell line. Two concentrations of mechlorethamine, L-phenylalanine mustard, and 4-hydroperoxycyclophosphamide, which produced 1 or 3 log cell kills were used to examine the effects of drug exposure on the expression of specific genes. Steady state mRNA levels were measured by Northern blot analysis. Following a 1-h drug exposure, RNA was isolated from cells at 0, 6, 12, and 24 h following drug removal. The agents used produced changes in the expression of specific genes, and all three did so in a similar fashion. Immediately following drug removal, the steady state expression of c-myc in treated cells was increased 2- to 3-fold compared to control. At 6 and 12 h following drug removal, c-myc levels were depressed 2.5- to 5-fold. By 24 h, c-myc expression approached, but remained below, control levels. Immediately following drug removal, c-fos levels were increased 3- to 4-fold, and from 6 to 24 h following drug removal, c-fos levels gradually return to, or fell below low basal levels. During the 24-h time course, drug treatment had little or no effect on the steady state levels of N-ras or beta-actin. These data support the hypothesis that alkylating agents may suppress the expression of specific transforming genes.
开展这项研究是为了确定DNA损伤性化疗药物是否能够抑制癌基因的表达。研究了三种结构相关的双功能烷化剂对人结肠癌细胞系Colo320HSR中c-myc、c-fos、N-ras和β-肌动蛋白的稳态mRNA水平的影响。Colo320HSR具有扩增的、高度过表达的c-myc癌基因，被认为是该细胞系的转化基因之一。使用两种浓度的氮芥、L-苯丙氨酸氮芥和4-氢过氧环磷酰胺，它们可导致1或3个对数级的细胞杀伤，以研究药物暴露对特定基因表达的影响。通过Northern印迹分析测量稳态mRNA水平。在1小时的药物暴露后，在去除药物后的0、6、12和24小时从细胞中分离RNA。所用药物导致特定基因表达发生变化，并且所有三种药物的作用方式相似。在去除药物后立即，与对照相比，处理细胞中c-myc的稳态表达增加了2至3倍。在去除药物后的6和12小时，c-myc水平降低了2.5至5倍。到24小时时，c-myc表达接近但仍低于对照水平。在去除药物后立即，c-fos水平增加了3至4倍，并且在去除药物后的6至24小时，c-fos水平逐渐恢复到或降至低基础水平以下。在24小时的时间进程中，药物处理对N-ras或β-肌动蛋白的稳态水平几乎没有影响。这些数据支持烷化剂可能抑制特定转化基因表达的假说。
