Quantitation of 2-hydroxyglutarate in human plasma via LC–MS/MS using a surrogate analyte approach
Abstract
Aim: 2-Hydroxyglutarate (2-HG) is a target engagement biomarker in patients after treatment with inhibitors of mutated isocitrate dehydrogenase (mIDH). Accurate measurement of 2-HG is critical for monitoring the inhibition effectiveness of the inhibitors. Materials & methods: Human plasma samples were spiked with stable isotope labelled internal standard, processed by protein precipitation, and analyzed using LC–MS/MS. This method was validated following regulatory guidance and has been successfully applied in a clinical study for mIDH inhibition. Results: An LC–MS/MS method with a surrogate analyte approach was developed and validated to measure 2-HG in human plasma with acceptable intra- and inter-assay accuracy and precision. Conclusion: A sensitive and robust LC–MS/MS method was developed and validated for measuring 2-HG in human plasma.
Papers of special note have been highlighted as: • of interest; •• of considerable interest
References
- 1. . The human PICD gene encodes a cytoplasmic and peroxisomal NADP(+)-dependent isocitrate dehydrogenase. J. Biol. Chem. 274(43), 30527–30533 (1999).
- 2. Targeted inhibition of mutant IDH2 in leukemia cells induces cellular differentiation. Science 340(6132), 622–626 (2013).
- 3. Localization of cytosolic NADP-dependent isocitrate dehydrogenase in the peroxisomes of rat liver cells: biochemical and immunocytochemical studies. J. Histochem. Cytochem. 49(9), 1123–1131 (2001).
- 4. Mutant IDH inhibits HNF-4α to block hepatocyte differentiation and promote biliary cancer. Nature 513(7516), 110–114 (2014).
- 5. IDH mutation impairs histone demethylation and results in a block to cell differentiation. Nature 483(7390), 474–478 (2012).
- 6. Oncometabolite 2-hydroxyglutarate is a competitive inhibitor of α-ketoglutarate-dependent dioxygenases. Cancer Cell 19(1), 17–30 (2011).
- 7. A comparison of azacitidine and decitabine activities in acute myeloid leukemia cell lines. PLoS ONE 5(2), e9001 (2010).
- 8. Cancer-associated IDH1 mutations produce 2-hydroxyglutarate. Nature 462(7274), 739–744 (2009).
- 9. Cancer-associated metabolite 2-hydroxyglutarate accumulates in acute myelogenous leukemia with isocitrate dehydrogenase 1 and 2 mutations. J. Exp. Med. 207(2), 339–344 (2010).
- 10. The common feature of leukemia-associated IDH1 and IDH2 mutations is a neomorphic enzyme activity converting alpha-ketoglutarate to 2-hydroxyglutarate. Cancer Cell 17(3), 225–234 (2010).
- 11. Prospective serial evaluation of 2-hydroxyglutarate, during treatment of newly diagnosed acute myeloid leukemia, to assess disease activity and therapeutic response. Blood 120(23), 4649–4652 (2012).
- 12. Discovery of AG-120 (Ivosidenib): a first in class mutant IDH1 inhibitor for the treatment of IDH1 mutant cancers. ACS Med. Chem. Lett. 9(4), 300–305 (2018).
- 13. The importance of plasma D-2HG measurement in screening for IDH mutations in acute myeloid leukaemia. Br. J. Haematol. 173(2), 314–328 (2016). • A reference to report (D)-2-HG is the major oncometabolite from IDH mutations.
- 14. Clinical pharmacokinetics and pharmacodynamics of ivosidenib, an oral, targeted inhibitor of mutant IDH1, in patients with advanced solid tumors. Invest. New Drug 38(2), 433–444 (2020).
- 15. Circulating oncometabolite D-2-hydroxyglutarate enantiomer is a surrogate marker of isocitrate dehydrogenase mutated intrahepatic cholangiocarcinomas. Eur. J. Cancer 90, 83–91 (2018).
- 16. US FDA. Biomarker Qualification: Evidentiary Framework (2018). https://www.fda.gov/media/122319/download
- 17. Surrogate matrix and surrogate analyte approaches for definitive quantitation of endogenous biomolecules. Bioanalysis 4(19), 2343–2356 (2012). • An overview of biomarker quantitation using surrogate matrix or surrogate analyte approaches.
- 18. A surrogate analyte-based liquid chromatography–tandem mass spectrometry method for the determination of endogenous cyclic nucleotides in rat brain. J. Pharm. Biomed. Anal. 146, 361–368 (2017).
- 19. A validated LC–MS/MS assay for quantification of 24(S)-hydroxycholesterol in plasma and cerebrospinal fluid. J. Lipid Res. 56(6), 1222–1233 (2015).
- 20. A double surrogate approach for the quantitation of 2-hydroxyglutarate – an oncometabolite in human brain tumors via LC–MS/MS. J. Pharm. Biomed. Anal. 179, 112916 (2020). • The first paper to report a bioanalytical method using surrogate analyte in surrogate matrix approach.
- 21. US FDA. Bioanalytical Method Validation Guidance for Industry (2018). https://www.fda.gov/media/70858/download •• New guidance from US FDA for bioanalytical method validation.