P-glycoprotein (Pgp) and cytochrome P450 3A4 (CYP3A4) are co-expressed in small intestinal enterocytes. Together, they are thought to modulate the oral bioavailability of a wide range of compounds that are substrates for both systems (Hunter & Hirst, 1997; Watkins, 1997).

Caco-2 cells exposed to 0.25 µM 1,25-(OH)₂D₃ for 2 weeks post-confluence (Schmiedlin-Ren et al. 1997) showed expression of CYP3A4 mRNA, whereas this transcript was undetectable in uninduced cells. Expression of Pgp and CYP3A5 mRNA were unaffected by 1,25-(OH)₂D₃ treatment. CYP3A4 activity, as determined by 6β-hydroxy-testosterone production, was below the limits of detection in uninduced cells. In 1,25-(OH)₂D₃-treated cells, 6β-hydroxy-testosterone production was 0.1 ± 0.0 and 0.5 ± 0.0 nmol cm^-2 h^-1 (mean ± S.E.M., n = 4) from 50 and 500 µM testosterone, respectively, comparable to published data (Hochman et al. 2000). Net secretory fluxes (basal-apical; B-A) of vinblastine and digoxin were not significantly different between induced and uninduced cells. Apparent K_m and V_max for vinblastine was 8.0 ± 4.0 µM, 3.6 ± 0.3 nmol cm^-2 h^-1 (n = 6), respectively, for induced cells, and 12.0 ± 5.0 µM, 2.7 ± 0.3 nmol cm^-2 h^-1 (n = 6), respectively, for un-induced cells. Rates of transport of 50 µM testosterone were similar in both the A-B and B-A directions (3.9 ± 0.1 and 3.4 ± 0.1 nmol cm^-2 h^-1; n = 3), indicating no net secretion in uninduced Caco-2 cells, and similarly in induced cells. However, testosterone (50 µM) inhibited vinblastine (10 µM) and digoxin (10 µM) secretory fluxes from 1.1 ± 0.05 to 0.6 ± 0.03 nmol cm^-2 h^-1 (n = 6, P < 0.05, Student’s t test) and from 0.9 ± 0.1 to 0.3 ± 0.03 nmol cm^-2 h^-1 (n = 3, P < 0.05), respectively across uninduced Caco-2 cell monolayers.

The finding that testosterone interacts with Pgp substrates, but is not itself directionally transported, suggests its role as a modulator of Pgp. Alternatively, a high passive permeability of Caco-2 cells for testosterone may mask any secretory influence of Pgp. The results support the use of 1,25-(OH)₂D₃ for CYP3A4 induction in Caco-2 cells to investigate interactions between metabolism and efflux.

L.C. is a BBSRC-CASE student sponsored by AstraZeneca.

Hochman, J.H., Chiba, M., Nishime, J., Yamazaki, M. & Lin, J.H. (2000). J. Pharm. Exp. Ther. 292, 310-318.

Hunter, J. & Hirst, B.H. (1997). Adv. Drug Del. Rev. 25, 129-157.

Schmiedlin-Ren, P., Thummel, K.E., Fisher, J.M., Paine, M.F., Lown, K.S. & Watkins, P.B. (1997). J. Pharmacol. Exp. Ther. 51, 741-754.

Watkins, P.B. (1997). Adv. Drug Deliv. Rev. 27, 161-170.