Relationship between protein levels and gene expression of dihydropyrimidine dehydrogenase in human tumor cells during growth in culture and in nude mice.

Protein levels and gene expression of dihydropyrimidine dehydrogenase (DPD), the rate-limiting enzyme for degradation of 5-fluorouracil, were studied in two human tumor cell lines (fibrosarcoma HT-1080 and pancreatic carcinoma MIAPaCa-2) in various growth phases of the cultured cells and of tumor xenografts implanted into nude mice. DPD catalytic activity and DPD protein content in cytosolic preparations were determined by means of radioenzymatic assay and western blot analysis, respectively. Relative DPD mRNA expression was determined by using a semi-quantitative reverse transcription-polymerase chain reaction in which glyceraldehyde-3-phosphate dehydrogenase mRNA was used as an internal standard. DPD activity and protein content in cultures of both cell lines increased in proportion to cell density (DPD activities ranged from undetectable to 84 pmol/min/mg protein in the HT-1080 cells and from undetectable to 335 pmol/min/mg protein in the MIAPaCa-2 cells). DPD mRNA levels, on the other hand, tended to decrease slightly during cell growth. DPD activity and protein content in HT-1080 tumor xenografts increased during growth in proportion to tumor weight (DPD activities ranged from 7 to 131 pmol/min/mg protein), but DPD mRNA levels did not correlate with tumor weight. DPD activity and protein content in MIAPaCa-2 tumor xenografts did not change much, and seemed to have already plateaued, since the tumors were small (weighing about 30 mg). These findings suggest that DPD protein expression during tumor growth is controlled at the post-transcriptional level.