EXPRESSION OF ENZYMATICALLY ACTIVE CYP3A4 BY CACO-2 CELLS GROWN ON EXTRACELLULAR MATRIX-COATED PERMEABLE SUPPORTS IN THE PRESENCE OF 1-ALPHA,25-DIHYDROXYVITAMIN D-3

The human colon carcinoma cell line, Caco-2, is widely used as a model for oral absorption of xenobiotics. The usefulness of Caco-2 cells ha s been limited, however, because they do not express appreciable quant ities of CYP3A4, the principle cytochrome P450 present in human small bowel epithelial cells. We report that treatment of Caco-2 cells with 1 alpha,25-dihydroxyvitamin D-3, beginning at confluence, results in a dose- and duration-dependent increase in CYP3A4 mRNA and protein, wit h little apparent effect on the expression of CYP3A5 or CYP3A7. This t reatment also results in increases in NADPH cytochrome P450 reductase and P-glycoprotein (the MDR1 gene product) but has no detectable effec t on expression of CYP1A1, CYP2D6, cytochrome b(5), liver or intestina l fatty acid binding proteins, or villin. Maximal expression of CYP3A4 requires an extracellular matrix on a permeable support and the prese nce of serum. In the treated cells, the intrinsic formation clearance of 1'-hydroxymidazolam (a reaction characteristically catalyzed by CYP 3A enzymes) was estimated to be somewhat lower than that of human jeju nal mucosa (1.14 and 3.67 ml/min/g of cells, respectively). The 1'-OH- midazolam/4-OH-midazolam product ratio produced by the cells (similar to 5.3) is comparable to, but somewhat lower than, that observed in hu man jejunal microsomes (7.4-15.4), which may reflect the presence of C YP3A7 in the Caco-2 cells. 25-Hydroxyvitamin D-3 is less efficacious b ut reproduces the effects of the dihydroxy compound, whereas unhydroxy lated vitamin D is without appreciable effect. These observations, tog ether with the time course of response, suggest that the vitamin D rec eptor may be involved in CYP3A4 regulation. The culture model we descr ibe should prove useful in defining the role of CYP3A4 in limiting the oral bioavailability of many xenobiotics.