Understanding the determinants of resistance of
5-fluorouracil (
5FU) is of significant value to optimising administration of the drug, and introducing novel agents and treatment strategies. Here, the expression of 92 genes involved in
5FU transport, metabolism, co-factor (
folate) metabolism and downstream effects was measured by real-time PCR low density arrays in 14 patient-derived
colorectal cancer xenografts characterised for
5FU resistance. Candidate gene function was tested by
siRNA and
uridine modulation, and immunoblotting, apoptosis and cell cycle analysis. Predictive significance was tested by immunohistochemistry of tumours from 125 stage III
colorectal cancer patients treated with and without
5FU. Of 8 genes significantly differentially expressed between
5FU sensitive and resistant xenograft tumours, CTPS2 was the gene with the highest probability of differential expression (p=0.008). Reduction of CTPS2 expression by
siRNA increased the resistance of
colorectal cancer cell lines DLD1 and LS174T to
5FU and its analogue,
FUDR. CTPS2
siRNA significantly reduced cell S-phase accumulation and apoptosis following
5FU treatment. Exposure of cells to
uridine, a precursor to the CTPS2 substrate
uridine triphosphate, also increased
5FU resistance. Patients with low CTPS2 did not gain a survival benefit from
5FU treatment (p=0.072), while those with high expression did (p=0.003). Low CTPS2 expression may be a rationally-based determinant of
5FU resistance.