Characterization of Degradation Products of Selpercatinib by Mass Spectrometry: Optimization of Stability-Indicating HPLC Method for Separation and Quantification of Process Related Impurities of Selpercatinib

Abstract

The study aimed to investigate a novel approach by utilizing liquid chromatography in coupled with mass spectrometry (LC-MS) to resolve, analyze and characterize significantly less quantity of stress degradation products (DPs) of selpercatinib along with its process related impurities. The analytes resolved on ZORBAX Eclipse (250 mm) stationary phase that was maintained employing 0.5 M sodium perchlorate at pH 5.4 methanol and acetonitrile in 50:20:30 (v/v) pumped at 0.8 mL/min isocratic flow and 241 nm wavelength. The method produces very high correlate linear curve in 30-240 μg/mL for selpercatinib and 0.03-0.24 μg/mL for its impurities with significantly low detection limit of 0.01 μg/mL for impurities. Selpercatinib pure compound was subjected to stress studies and chromatographic results confirm the formation of four DPs, which were characterized with the interpretation of their mass fragmentation pattern. The DPs were identified as 6-hydroxy-4-(6-(6-((6-methoxypyridin3-yl)methyl)-3,6-diazabicyclo[3.1.1]heptan-3-yl)pyridin-3-yl)pyrazolo[1,5-a]pyridine-3-carbonitrile (DP 1), 6′-(6-((6-methoxypyridin-3-yl)methyl)-3,6-diazabicyclo[3.1.1]heptan-3-yl)-1,2-dihydro-[3,3′-bipyridin]-5-ol (DP 2), (6-hydroxy-4-(6-(6-(pyridin-3-ylmethyl)-3,6-diazabicyclo[3.1.1]heptan-3-yl)pyridin-3-yl)pyrazolo[1,5-a]pyridine-3-carbonitrile (DP 3), 1-amino-6′-(6-((6-methoxypyridin-3-yl)methyl)-3,6-diazabicyclo[3.1.1]heptan-3-yl)-1,2-dihydro-[3,3′-bipyridin]-5-ol (DP 4), 6-(2-hydroxy-2-methyl-propoxy)-4-(6-(3-methylpiperazin-1-yl)pyridin-3-yl)pyrazolo[1,5-a]pyridine-3-carbonitrile (DP 5) and 6-(2-hydroxy-2-methyl-propoxy)-4-(6-(3-methylpiperazin-1-yl)pyridin-3-yl)pyrazolo[1,5-a]pyridine-3-carbonitrile (DP 6). Based on findings, it was concluded that this method was effectively applied for the regular quantification of impurities of selpercatinib in formulations. Additionally, it demonstrated applicability for the identification of both known and unknown impurities of selpercatinib.