Influence of 5-fluorouracil on ferredoxin reductase mRNA splice variants in colorectal carcinomas

Abstract

5-Fluorouracil (5-FU) is a frequently used antitumor drug. Recently, it has been shown that mRNA and protein levels of the ferredoxin reductase gene (gene, FDXR; protein, FR) increase drastically after 5-FU treatment in various cell lines including colorectal cancer. The induction is mediated by p53 and enhanced reactive oxygen species (ROS)-associated apoptosis. Thus, knowledge about FDXR expression in human tissue and expression of the known splice variants is critical for understanding this finding. A sensitive and specific reverse transcriptase polymerase chain reaction (RT-PCR) assay for quantification of FDXR mRNA levels including the splice variants, a biological active variant (-18 bp) and an inactive variant (+18 bp), was developed and used to measure mRNAs after 5-FU chemotherapy in colorectal tissues of 40 cancer patients prior to and after treatment with 5-FU for 14 days. Before treatment, the great majority of normal tissues expressed the splice variants in a 100:1 ratio in favor of the -18-variant similar to what has been reported for other tissues. In tumors, the mRNA levels of total FDXR and splice variants were approximately 2-fold higher compared to the normal tissue. After 5-FU treatment, levels of the +18-variant increased 17-fold in tumors and 31-fold in normal tissues, clearly shifting the ratio towards the +18-form. 5-FU-mediated -18-variant induction (>1) in normal (12/17) and tumor tissues (12/16) was apparently associated with response, while a balanced ratio (0.1-2) was associated with 5-FU resistance (n=5) based on the histological evaluation of the tissues.