DEVELOPMENT AND VALIDATION OF AN LC-MS/MS METHOD FOR THE QUANTIFICATION OF REPOTRECTINIB IN HUMAN PLASMA
DOI:
https://doi.org/10.22159/ijap.2026v18i1.56285Keywords:
Repotrectinib, Cancer, LC-MS/MS, Validation, AccuracyAbstract
Objective: The primary aim of this study is to develop and verify a linear, exact, and specific LC-ESI-MS/MS technique for the measurement of Repotrectinib.
Methods: Chromatographic resolution was attained using an ODS Zorbax C18 column (50 mm × 4.6 mm, 2.1 µm) and a mobile phase comprising methanol, 0.1% formic acid, and acetonitrile in a ratio of 50:15:35, with a flow rate of 0.5 ml/min from the stationary phase. The procedure was conducted by observing the defined ionic transitions of m/z 356.15/318.13 for Repotrectinib and 478.09/451.08 for the Apalutamide internal standard in multiple reaction monitoring.
Results: The linear regression equation was y = 0.0001x + 0.0012, with a coefficient of determination (r²) of 0.9997. The percentage coefficient of variation results for the matrix effect for Low-QC and High-QC levels were 2.90% and 3.41%, respectively. The average recovery percentages for Repotrectinib at High-QC (11.25 µg/ml), MQC (7.50 µg/ml), and Low-QC (1.05 µg/ml) were 103.27%, 97.26%, and 96.34%, respectively. The measured values ranged from 2.00% to 4.03% for the QC samples at concentrations of 0.375, 1.05, 7.50, and 11.25 µg/ml.
Conclusion: The created approach underwent validation in accordance with FDA guidelines and is applicable for the assessment of Repotrectinib in biological samples for quality control, forensic analysis, and bioavailability investigations.
References
1. Yun MR, Kim DH, Kim SY, Joo HS, Lee YW, Choi HM, Park CW, Heo SG, Kang HN, Lee SS, Schoenfeld AJ, Drilon A, Kang SG, Shim HS, Hong MH, Cui JJ, Kim HR, Cho BC. Repotrectinib exhibits potent antitumor activity in treatment-naive and solvent-front-mutant ROS1-rearranged non-small cell lung cancer. Clin Cancer Res. 2020 Jul 1;26(13):3287-95. doi:10.1158/1078-0432.CCR-19-2777.
2. Murray BW, Rogers E, Zhai D, Deng W, Chen X, Sprengeler PA, Zhang X, Graber A, Reich SH, Stopatschinskaja S, Solomon B, Besse B, Drilon A. Molecular characteristics of repotrectinib that enable potent inhibition of TRK fusion proteins and resistant mutations. Mol Cancer Ther. 2021 Dec;20(12):2446-56. doi:10.1158/1535-7163.MCT-21-0632.
3. Keddy C, Shinde P, Jones K, Kaech S, Somwar R, Shinde U, Davare MA. Resistance profile and structural modeling of next-generation ROS1 tyrosine kinase inhibitors. Mol Cancer Ther. 2022 Feb;21(2):336-46. doi:10.1158/1535-7163.MCT-21-0395.
4. Bristol Myers Squibb. AUGTYRO™ (repotrectinib) capsules, for oral use. Prescribing information [Internet]. 2023 Nov [cited 2025 Sep 1]. Available from: https://packageinserts.bms.com/pi/pi_augtyro.pdf
5. Bristol Myers Squibb. U.S. Food and Drug Administration approves Augtyro™ (repotrectinib), a next-generation tyrosine kinase inhibitor (TKI), for the treatment of locally advanced or metastatic ROS1-positive non-small cell lung cancer (NSCLC) [Internet]. 2023 [cited 2025 Sep 1]. Available from: https://news.bms.com/news/corporate-financial/2023/US-Food-and-Drug-Administration-Approves-Augtyro/default.aspx
6. Li W, Perpinioti N, Schinkel AH, Beijnen JH, Sparidans RW. Bioanalytical assay for the new-generation ROS1/TRK/ALK inhibitor repotrectinib in mouse plasma and tissue homogenate using liquid chromatography–tandem mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci. 2020;1144:122098. doi:10.1016/j.jchromb.2020.122098.
7. Li, W., Perpinioti, N., Schinkel, A. H., Beijnen, J. H., & Sparidans, R. W. . Bioanalytical assay for the new-generation ROS1/TRK/ALK inhibitor repotrectinib in mouse plasma and tissue homogenate using liquid chromatography-tandem mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci., 2020;1144, Article 122098. https://doi.org/10.1016/j.jchromb.2020.122098.
8. U.S. Food and Drug Administration. Guidance for industry: bioanalytical method validation. Rockville (MD): FDA; 2001 May.
9. European Medicines Agency. Guideline on bioanalytical method validation. London: EMA; 2011.
10. International Conference on Harmonisation. ICH Q2B: validation of analytical procedures—methodology. Geneva: ICH; 1996.
11. Fernandez MMR, Wille SMR, Samyn N. Quantitative method validation for the analysis of 27 antidepressants and metabolites in plasma with ultra-performance liquid chromatography–tandem mass spectrometry. Ther Drug Monit. 2012;34(1):11-24. doi:10.1097/FTD.0b013e31823bf0fd.
12. Wozniakiewicz M, Wietecha-Posłuszny R, Moos A. Development of microextraction by packed sorbent for toxicological analysis of tricyclic antidepressant drugs in human oral fluid. J Chromatogr A. 2014;1337:9-16. doi:10.1016/j.chroma.2014.02.037.
13. Sellappan M, Devakumar D. Development and validation of RP-HPLC method for estimation of escitalopram oxalate and flupentixol dihydrochloride in combined dosage form and plasma. Int J Pharm Pharm Sci. 2021 Feb 1;13(2):61-6. doi:10.22159/ijpps.2021v13i2.30158.
14. Gurav P, Damle M. Bioanalytical method for estimation of teriflunomide in human plasma. Int J Pharm Pharm Sci. 2022 Sep;14(9):19-23. doi:10.22159/ijpps.2022v14i9.45151.
15. Krishnan VS, Bhikshapathi D, Cheruku S. Method development and validation for the quantification of abametapir in biological matrices by LC-ESI-MS/MS. Indian J Pharm Educ Res. 2024 Aug;58(3s):1028-s1033. doi:10.5530/ijper.58.3s.102.
16. SaiUdayKiran G, Sandhya P, Shankar CH, Bhikshapathi DVRN, Mamatha P. An LC–MS/MS quantification method development and validation for dabrafenib in biological matrices. J Appl Pharm Sci. 2023 Jan;13(1):180-6. doi:10.7324/JAPS.2023.130117.
17. Dadhaniya T, Chaudhary K, Mehta P. Development of LC-MS/MS method for determination of iloperidone in rabbit plasma: application to a pharmacokinetic study. Int J Pharm Pharm Sci. 2013 Feb;7(4):294-7. Available from: https://journals.innovareacademics.in/index.php/ijpps/article/download/5121/8609/0
18. Puttagunta SB, Shaik RP, Bannoth CK, Challa BSR, Awen BZS. Bioanalytical method for quantification of solifenacin in rat plasma by LC-MS/MS and its application to pharmacokinetic study. J Anal Sci Technol. 2014 Jul;5(1):35. doi:10.1186/s40543-014-0035-0.
19. Shankar CH, Bhikshapathi D. Method development and validation for the quantification of pexidartinib in biological samples by LC-MS/MS. Int J Pharm Res. 2021;13(1):6522-30.
20. Deepan T, Basaveswara Rao MV, Dhanaraju MD. Bioanalytical method development and validation of canagliflozin in human plasma by liquid chromatography–tandem mass spectrometry. Asian J Pharm Clin Res. 2019;12(8):46-51. doi:10.22159/ajpcr.2019.v12i18.33228.
21. Gurav P, Damle M. Bioanalytical method for estimation of teriflunomide in human plasma. Int J Pharm Pharm Sci. 2022 Sep;14(9):19-23. doi:10.22159/ijpps.2022v14i9.45151.
22. Lolla S, Gubbiyappa KS, Cheruku S, Bhikshapathi DVRN. Validation of an LC-MS/MS method for quantitation of fostemsavir in plasma. J Pharmacol Toxicol Methods. 2023;120:107254. doi:10.1016/j.vascn.2023.107254.
23. Nimmakayala MR, Kolli D, Durga Bhavani PN. Bioanalytical method development and validation of maralixibat in rat plasma by LC-MS/MS detection and its application to a pharmacokinetic study. Int J Appl Pharm. 2023;15(4):166-72. doi:10.22159/ijap.2023v15i4.47768.
24. Yatha R, Bhikshapathi D, Cheruku S, Bigala R. Development of fast and simple LC-ESI-MS/MS technique for the quantification of regorafenib; application to pharmacokinetics in healthy rabbits. Curr Pharm Anal. 2021;17:554-63. doi:10.2174/1573412916666191111144707.
25. Parmar I, Patel YA. Recent method development by analytical techniques of new FDA approved drugs in 2021. Int J Curr Pharm Res. 2022;14(3):17-21. doi:10.22159/ijcpr.2022v14i3.1975.
26. Shankar CH, Bhikshapathi D, Medipalli V, Arjuna RN, Sadasivam RK. Bioanalytical method development and validation for the quantitation of larotrectinib in human plasma: application to pharmacokinetics in healthy rabbits. J Appl Pharm Sci. 2023 Nov;13(11):111-8.
Published
How to Cite
Issue
Section
Copyright (c) 2025 VASUDEVA RAO BOBBA, MOGILI PADMA, VUNDAVILLI JAGADEESH KUMAR
This work is licensed under a Creative Commons Attribution 4.0 International License.
