An Increase in the Expression of Ribonucleotide Reductase Large Subunit 1 Is Associated with Gemcitabine Resistance in Non-Small Cell Lung Cancer Cell Lines

Abstract

The mechanisms of resistance to the antimetabolite gemcitabine in non-small cell lung cancer have not been extensively evaluated. In this study, we report the generation of two gemcitabine-selected non-small cell lung cancer cell lines, H358-G200 and H460-G400. Expression profiling results indicated that there was evidence for changes in the expression of 134 genes in H358-G200 cells compared with its parental line, whereas H460-G400 cells exhibited 233 genes that appeared to be under-or overexpressed compared with H460 cells. However, only the increased expression of ribonucleotide reductase subunit 1 (RRM1), which appeared in both resistant cell lines, met predefined analysis criteria for genes to investigate further. Quantitative PCR analysis demonstrated H358-G200 cells had a greater than 125-fold increase in RRM1 RNA expression. Western blot analysis confirmed high levels of RRM1 protein in this line compared with the gemcitabine-sensitive parent. No significant change in the expression of RRM2 was observed in either cell line, although both gemcitabine-resistant cell lines had an approximate 3-fold increase in p53R2 protein. A partial revertant of H358-G200 cells had reduced levels of RRM1 protein (compared with G200 cells), without observed changes in RRM2 or p53R2. In vitro analyses of ribonucleotide reductase activity demonstrated that despite high levels of RRM1 protein, ribonucleotide reductase activity was not increased in H358-G200 cells when compared with parental cells. The cDNA encoding RRM1 from H358-G200 cells was cloned and sequenced but did not reveal the presence of any mutations. The results from this study indicate that the level of RRM1 may affect gemcitabine response. Furthermore, RRM1 may serve as a biomarker for gemcitabine response.

Gemcitabine chemoresistance and molecular markers associated with gemcitabine transport and metabolism in human pancreatic cancer cells

“…This result supports the previous report that RRM1 is the marker predicting resistance to gemcitabine in lung cancer cell lines (Davidson et al, 2004). In this study, KLM1 gemcitabine-resistant cells had no dCK deficiency in contrast to the lack of dCK in PCI43 and PK1 gemcitabine-resistant cells.…”

In vitro synergistic cytotoxicity of gemcitabine and pemetrexed and pharmacogenetic evaluation of response to gemcitabine in bladder cancer patients

“…Furthermore, high expression of the catabolic enzymes 5'-nucleotidase (5'-NT) and cytidine deaminase (CDA) has been found in many cell lines resistant to gemcitabine (Eda et al, 1998;Lotfi et al, 2001). non-small cell lung cancer patients with low expression of the M1 subunit of RR (RRM1) significantly benefited from gemcitabine/ cisplatin neoadjuvant chemotherapy (Rosell et al, 2004), while resistance to gemcitabine was observed both in RRM1 and RRM2 overexpressing cells (Goan et al, 1999;Davidson et al, 2004).…”

Importance of Molecular Features of Non–Small Cell Lung Cancer for Choice of Treatment

“…46 In addition, when reflecting on the potential future of molecular testing in lung cancer, it is important to consider the association of the excision repair cross-complementation group 1 gene and high expression of the ribonucleotide reductase subunit 1 with platinum 47,48 and gemcitabine resistance, respectively. 49 Clearly, the ability to more easily determine mutation status in patients with advanced NSCLC would be invaluable for overcoming the barriers related to the relative or absolute lack of tumor tissue for direct sequencing.…”