TABLE 2

Increase in CYP3A activity in microsomes from rat hepatocytes dosed once daily for 3 days with N-hydroxyformamide TACE/MMP inhibitors

Microsomes were prepared from hepatocytes collected 24 h after the last daily dose of vehicle or compound. Production of the CYP3A-specific metabolite, 6β-hydroxy testosterone, was measured by LC/MS-MS after incubation of microsomes with testosterone as described under Materials and Methods. CYP3A activity is reported as the rate of 6β-hydroxy testosterone formed per milligram of protein. A smooth spline curve was fit to the fold induction data for each TACE/MMP inhibitor and a significant elevation in CYP3A activity was assessed as described under Materials and Methods. The minimum concentrations that caused a CYP3A fold induction different from control were 1.2, 0.8, and 3.9 μM for GW3333, GW6495, and GI4023, respectively.


Treatment

6β-Hydroxy Testosterone Formation Rate

Rat Liver 1
Rat Liver 2
pmol/min/mg protein fold induction pmol/min/mg protein fold induction
Controla 251 1.0 673 1.0
PCN, 0.2 μM 273 1.1 789 1.2
PCN, 1.0 μM 619 2.5 1240 1.8
Phenobarbital, 10 μM 254 1.0 759 1.1
Phenobarbital, 100 μM 386 1.5 1196 1.8
GW3333, 1.0 μM 373 1.5 958 1.4
GW3333, 2.5 μM 708 2.8 1522 2.3
GW3333, 5.0 μM* 1433 5.7 2453 3.6
GW3333, 10.0 μM* 1635 6.5 3875 5.8
GW3333, 25.0 μM 1628 6.5 4448 6.6
GW3333, 50.0 μM 1659 6.6 4988 7.4
GW6495, 1.0 μM 369 1.5 903 1.3
GW6495, 5.0 μM 511 2.0 1162 1.7
GW6495, 25.0 μM 721 2.9 1903 2.8
GW6495, 100 μM 733 2.9 1668 2.5
GI4023, 1.0 μM 328 1.3 696 1.0
GI4023, 5.0 μM 632 2.5 809 1.2
GI4023, 25.0 μM 893 3.6 1234 1.8
GI4023, 100 μM
1282
5.1
3118
4.6
  • * Significantly different (p < 0.05) CYP3A fold induction by GW3333 relative to GI4023 and GW6495 at the same concentration using a Welch two-sample t test.

  • a Control hepatocytes treated with media containing 0.1%DMSO.