RAPID-DETERMINATION OF ORAL PHARMACOKINETICS AND PLASMA-FREE FRACTIONUSING COCKTAIL APPROACHES - METHODS AND APPLICATION

Purpose. To apply cocktail approaches for protein binding (PB) and pha rmacokinetics (PK) within a discovery program as a means of providing timely systemic exposure (AUG and C-max) data. Methods. For PB data, a procedure of cocktail ultrafiltration, mixed matrix sample preparatio n and single quadrupole atmospheric pressure ionization LC/MS analysis was used. In vivo PK studies consisted of 4 experimental compounds an d a control compound dosed orally at 1 mg/kg (5 mg/kg total dose), wit h plasma samples obtained at 0.5, 1, 2, 4 and 8 h post dose. For PB an d in vivo PK analysis, a control compound was tested within each cockt ail to ensure consistent reproducibility. Results. Approximately 2 wee ks were spent comparing single and cocktail approaches to determine th e feasibility of this method for this project. Comparisons of cocktail data with single compound data revealed no significant differences be tween the approaches. The oral AUC values ranged from 0.01 to 9.28 mu g.hr/ml and the C-max values ranged from 0.04 to 2.17 mu g/ml. Free fr actions of the 44 compounds studied ranged from 0.006 to 0.271. Using the free fraction values to correct for free AUC and C-max results in ranges of 0.001 to 0.473 mu g.hr/ml, and 0.001 to 0.119 mu g/ml, respe ctively. Conclusions. All 44 compounds tested had similar potencies in vivo. Thus, these results suggest that a respective 400 and 100-fold range in AUC and C-max corrected for free fraction exist in the presen ce of comparable in vivo activity. The ability to generate this type o f data in a timely manner allowed the selection of a candidate with lo w peripheral exposure relative to the effective dose. The free fractio n and PK data on the 44 compounds described was collected within three work days by 2 lab scientists.