Overexpression of folate binding protein α is one of the mechanism explaining the adaptation of HT29 cells to high concentration of methotrexate

Abstract

The human colon adenocarcinoma cell line HT29 can be adapted to 10 −7- 10 −4 M concentrations of methotrexate (MTX). Cells adapted to 10 −4 M MTX have an enterocyte-like phenotype with DHFR gene amplification. Presently, we hypothetized that an increased expression of folate binding protein (FBP) may participate to the MTX resistance of 10 −4 MTX HT29 cells. The cDNA FBPα/β-actin ratio of amplified transcripts was 4.8- and 1.5- fold higher in 10 −4 and in 10 −7 M MTX HT29 respectively, than in standard type HT29 cells. An increase of transcript level was observed when decreasing folic acid concentration. PI-PLC cleaved 7.7 times more membrane FBP in 10 −4 M than in 10 −7 M MTX and wild type HT29 cells. In contrast to 10 −7 M MTX cells, growth of 10 −4 M MTX cells was dependent on folic acid concentration and abolished at a concentration lower than 0.9 μM. In conclusion, the adaptive mechanism of HT29 cells resistant to 10 −4 M MTX is the result of the synergistic overexpression of both DHFR and FBPα. Overexpression of FBPα may be related to the enterocyte-like phenotype of the cells.