Bioactive indanes: Development and validation of an LC-MS/MS bioanalytical method for the determination of PH46A, a new potential anti-inflammatory agent, in dog and rat plasma and its application to a pharmacokinetic study in dog

A new chemical entity, which is a chiral indane dimer, PH46A, has been developed by our research group. As a clinical candidate. PH46A has recently completed Phase I clinical studies in man. Previously, during its pre-clinical development, in in vivo pre-clinical studies PH46A showed potent anti-inflammatory properties, which can be targeted at a range of diseases, including inflammatory bowel disease (IBD). To support the pre-clinical development of this drug candidate, we developed a LC—MS/MS method for determining PH46 (the acid form of PH46A salt) in both dog and rat plasma using Compound 1 as internal standard (IS). Those species were selected for safety pharmacology and toxicology, as well as pharmacokinetics studies. The method was validated over the range 10−10000 ng/mL for both matrices and the linearity, accuracy, precision and specificity over this range were demonstrated to be acceptable. No significant matrix effects or carryover were observed for both PH46 and IS and recovery was consistent. PH46 was found to be stable in both dog and rat plasma under the test conditions, such as at room temperature for >24 h, through 3 freeze/thaw cycles, and at -20 °C for >1 month. PH46 and IS in dog and rat plasma extracts were also found to be stable in the autosampler against fresh standard extracts on re-injection after 143.5 h and 243.5 h, respectively at 4 °C. 10- and 100-fold dilutions with control matrix were found not to affect the performance of the assay. This method was successfully applied to a pharmacokinetic study in the dog. With the exception of one dog, 003 M, oral administration of PH46A in gelatine capsules was well tolerated at a dose level of 100 mg/kg. The highest C(max) was observed in animal 003 M. The rapid absorption and high plasma concentration observed for animal 003 M compared to the data for animals 001 M and 002 M may account for the sickness observed in this animal; however, the reasons for this have not been investigated.