D03012908

The pharmacokinetic / pharmacodynamic (PK/PD) relationship of PTM in vivo was measured by assessing DNL in male, lean C57BL/6 mice (3 months old) fed either a regular chow diet or a high fructose diet (60% kcal; D03012908, Research diets) for 2 weeks, or eDIO mice maintained on high fat diet (60% kcal, Research Diets, D12492) for 16 wks (n = 7–8 per group). Control mice were fasted overnight. DNL was quantified using a stable isotope tracer method and mass spectrometric analysis as described [20 (link)]. PTM was formulated into water and dosed at indicated doses, orally (p.o.) at 5 ml/kg body weight, 30 minutes before dosing of deuterated water (D₂O) given intraperitoneally (i.p.) at 20 mpk body weight. Mice were euthanized by CO₂ asphyxiation 2 hours post D₂O injection. EDTA plasma was collected via cardiac puncture and livers were collected for analysis of PK and DNL.

Lean C57BL/6 and established diet-induced obese (eDIO) mice were purchased from Taconic Biosciences (Germantown, NY). Lean db/+ and db/db mice were purchased from Jackson Laboratory (Bar Harbor, ME). Animals were maintained in a 12/12 hour light-dark cycle with free access to food and water in an environment with temperature maintained at 22°C. Animals were maintained on a regular rodent chow diet (Teklad, 7012, 5% dietary fat; 3.75 kcal/g) (Madison, WI), or a high fructose diet (Research Diets, D03012908, 60% kcal), or a high-fat diet (Research Diets, D12492, 60% kcal% dietary fat) (New Brunswick, NJ) with free access to water. Diet and dosing information of acute and chronic mouse studies are summarized in Table 1. At the end of studies, carbon dioxide (10–30% / min) inhalation was used to euthanize mice at the end of studies. The method to verify the death of mice includes no movement and heartbeat. All mice procedures and experimental procedures used in this paper were approved and performed in accordance with the guidelines of the Institutional Animal Care and Use Committee (IACUC) of Merck.