Characterization of stress degradation products of nintedanib by UPLC, UHPLC-Q-TOF/MS/MS and NMR: Evidence of a degradation product with a structure alert for mutagenicity

Publication: J Pharm Biomed Anal
Software: ADMET Predictor®

Abstract

Nintedanib is an anti-cancer drug used for the treatment of idiopathic pulmonary fibrosis and non-small cell lung cancer. The purpose of this study was to explore its degradation chemistry under various stress conditions recommended in ICH guidelines Q1A R(2). The drug was subjected to hydrolytic, photolytic, thermal and oxidative (H2O2, AIBN, FeCl3 and FeSO4) stress conditions. The degradation products formed in stressed solutions were successfully separated on an ACQUITY UPLC CSH C18 (2.1 × 100 mm, 1.7 μm) column, using a gradient UPLC-PDA method, developed with acetonitrile:methanol (90:10) and 0.1 % formic acid (pH 3.0) as the mobile phase. The drug proved to be labile to acidic, neutral and alkaline hydrolytic, and H2O2/AIBN oxidative conditions. It was stable to photolytic and thermal stress conditions, and even in oxidative reaction solutions containing FeCl3 or FeSO4. Additionally, the drug exhibited instability when its powder with added sodium bicarbonate was stored at 40 °C/75 % RH for 3 months. In total, nine degradation products (DPs 1–9) were formed. To characterize them, a comprehensive mass fragmentation pathway of the drug was first established using UHPLC-Q-TOF/MS/MS data. Similarly, the mass studies were then carried out on the stressed samples using the developed UPLC method. All the degradation products were primarily characterized through comparison of their mass fragmentation profiles with that of the drug. To confirm the structure in one case (DP 3), additional nuclear magnetic resonance (NMR) studies were carried out on the isolated product. Subsequently, mechanisms for their formation were laid down. A significant finding was the formation of a degradation product upon acid hydrolysis having a free aromatic amine moiety, which is considered as a structural alert for mutagenicity. Furthermore, the physicochemical and ADMET properties of the drug and its degradation products were predicted using ADMET predictor™ software.

By Vivek Dhiman, Ankit Balhara, Saranjit Singh, Shristy Tiwari, Samanthula Gananadhamu, M.V.N. Kumar Talluri