Characterizing the in vitro species differences in N-glucuronidation of a potent pan-PIM inhibitor GNE-924 containing a 3,5-substituted 6-azaindazole

1. Glucuronidation of amines has been shown to exhibit large species differences, where the activity is typically more pronounced in human than in many preclinical species such as rat, mouse, dog and monkey. The purpose of this work was to characterize the in vitro glucuronidation of GNE-924, a potent pan-PIM inhibitor, to form M1 using liver microsomes (LM) and intestinal microsomes (IM).

2. M1 formation kinetics varied highly across species and between liver and intestinal microsomes. In LM incubations, rat exhibited the highest rate of M1 formation (CL_int,app) at 140 ± 10 µL/min/mg protein, which was approximately 30-fold higher than human. In IM incubations, mouse exhibited the highest CL_int,app at 484 ± 40 µL/min/mg protein, which was >1000-fold higher than human. In addition, CL_int,app in LM was markedly higher than IM in human and monkey. In contrast, CL_int,app in IM was markedly higher than LM in dog and mouse.

3. Reaction phenotyping indicated that UGT1A1, UGT1A3, UGT1A9, UGT2B4 and the intestine-specific UGT1A10 contributed to the formation of M1.

4. This is one of the first reports showing that N-glucuronidation activity is significantly greater in multiple preclinical species than in humans, and suggests that extensive intestinal N-glucuronidation may limit the oral exposure of GNE-924.