Establishment of MDCKII cell monolayer with metabolic activity by CYP3A4 transduced with recombinant adenovirus.
This study aims to establish an in vitro system that can assess intestinal first-pass metabolism of CYP3A4 substrate drugs using adenoviral transduction. Madin-Darby canine kidney II (MDCKII) cells were used as a model of intestinal epithelial cells. Recombinant adenovirus expressing green fluorescent protein (AdGFP) and CYP3A4 (AdCYP3A4) was used as vectors. On day 2 after seeding MDCKII cells onto a semipermeable membrane, cells were infected with each adenovirus vector at various MOIs (multiplicities of infection) ranging from 0 to 200. On day 5, cell monolayers were used for drug transport study. The expression of GFP in monolayers of MDCKII cells transduced with AdGFP increased MOI-dependently and adenoviral infection showed no effect on the membrane permeability of drugs. The metabolite formation rate of midazolam, a CYP3A4 substrate, in the permeation process of a monolayer linearly increased with an increase in the MOI of AdCYP3A4. When the period that vectors and cells were located adjacent to each other was prolonged, the rate increased 2-fold compared with that calculated from a result with a monolayer obtained from a shorter period of adjacency. This study indicates that monolayers of MDCKII cells transduced with AdCYP3A4 have the potential to enable estimation of the first-pass metabolism by CYP3A4 in the intestinal absorption process.
Faculty of Pharmaceutical Sciences, Setsunan University, Hirakata, Japan. makoto@pharm.setsunan.ac.jp
Makoto
M
Kataoka
Yuki
Y
Terashima
Katsuhiko
K
Mizuno
Yoshie
Y
Masaoka
Shinji
S
Sakuma
Tsuyoshi
T
Yokoi
Shinji
S
Yamashita
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D002461:Cell Line, Transformed; D002463:Cell Membrane Permeability / Q000235:genetics; D051544:Cytochrome P-450 CYP3A / Q000235:genetics* / Q000378:metabolism*; D004285:Dogs; D005822:Genetic Vectors / Q000235:genetics; D061985:Madin Darby Canine Kidney Cells; D008874:Midazolam / Q000493:pharmacokinetics; D014161:Transduction, Genetic
D051544:Cytochrome P-450 CYP3A; D008874:Midazolam