IP Library Granted Patent US 12,576,055
Granted Patent B2
US 12,576,055 · App. 16/965,629 · Granted Mar 17, 2026

Disease detection and treatment based on trimethyl-lysine levels

Inventors: Stanley L. Hazen (Pepper Pike, OH); Zeneng Wang (Shaker Heights, OH); Xinmin S. Li (Cleveland, OH)
Assignee: The Cleveland Clinic Foundation
A61K31/198A61K31/133A61K31/4164A61K31/675A61P9/04A61P13/12G01N21/33G01N24/08A61K45/06G01N2800/34
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Quick Facts
Patent No.
US 12,576,055
App. No.
16/965,629
Granted
Mar 17, 2026
Kind
B2
Abstract

The present invention relates to systems, kits, and methods for identifying subjects with increased levels of N6-trimethyl-lysine (TML) or the combination of TML and trimethylamine-n-oxide (TMAO), as well as methods of determining risk of disease (e.g., kidney disease or heart failure) based on such levels. In certain embodiments, subjects with elevated TML, or TML and TMAO are treated with a therapeutic (e.g., one that inhibits the TMA/FMO3/TMAO pathway).

Claims (5)

1 . A method of treating chronic kidney disease comprising:

a) identifying a subject as having chronic kidney disease based on: i) having increased levels of N6-trimethyl-lysine (TML) compared to a control level, or having increased levels of TML and trimethylamine-n-oxide (TMAO) compared to control levels, and

b) treating said subject with a first agent or first procedure that inhibits the TMA/FMO3/TMAO pathway.

2 . The method of claim 1 , wherein said first agent that inhibits the TMA/FMO3/TMAO pathway is selected from a compound comprising: a halomethyl betaine, a halomethyl betaine salt, a halomethyl betaine amide, and a halomethyl dimethyl amino alcohol.

3 . The method of claim 1 , wherein said first agent that inhibits the TMA/FMO3/TMAO pathway is a compound comprising a halomethyl choline.

Assignments (2)
CONFIRMATORY LICENSE Recorded Oct 13, 2023
From: CLEVELAND CLINIC FOUNDATION
To: NATIONAL INSTITUTES OF HEALTH (NIH), U.S. DEPT. OF HEALTH AND HUMAN SERVICES (DHHS), U.S. GOVERNMENT
Reel/Frame 065238/0135 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jan 13, 2021
From: HAZEN, STANLEY L.; WANG, ZENENG; LI, XINMIN S.
To: THE CLEVELAND CLINIC FOUNDATION
Reel/Frame 054900/0337 →
Continuity (2)
Provisional Application 62624969 · Feb 1, 2018
Related Publication 20210038550A1 · Feb 11, 2021
References Cited (96)
US 9168233B2 · Hazen et al. · 2015 [cited by applicant]
US 9694020B2 · Hazen · 2017 [cited by examiner]
US 20100285517A1 · Hazen et al. · 2010 [cited by applicant]
US 20120088259A1 · Pinet et al. · 2012 [cited by applicant]
US 20120157397A1 · Hazen et al. · 2012 [cited by applicant]
US 20160074440A1 · Brugere et al. · 2016 [cited by applicant]
US 20160089386A1 · Hazen · 2016 [cited by applicant]
US 20170152222A1 · Garcia-Garcia et al. · 2017 [cited by applicant]
US 20180000754A1 · Hazen et al. · 2018 [cited by applicant]
WO WO2018236899 · 2018 [cited by applicant]
International Search Report and Written Opinion for PCT/US19/16361. Mailed Apr. 23, 2019. 15 pages. [cited by applicant]
Afshinnia et al., Impaired beta-Oxidation and Altered Complex Lipid Fatty Acid Partitioning with Advancing CKD. J Am Soc Nephrol. Jan. 2018;29(1):295-306. [cited by applicant]
Albert et al., Myocardial lipidomics. Developments in myocardial nuclear lipidomics. Front Biosci. Jan. 1, 2007;12:2750-60. [cited by applicant]
Aldous et al., Comparison of high sensitivity and contemporary troponin assays for the early detection of acute myocardial infarction in the emergency department. Ann Clin Biochem. May 2011;48(Pt 3):241-8. [cited by applicant]
Austin et al., Introduction to the Analysis of Survival Data in the Presence of Competing Risks. Circulation. Feb. 9, 2016;133(6):601-9. [cited by applicant]
Beger et al., Metabolomics enables precision medicine: “A White Paper, Community Perspective”. Metabolomics. 2016;12(10):149. 15 pages. [cited by applicant]
Body et al., One Shot to Rule Out: Does the Limit of Detection of a High-Sensitivity Troponin Assay Hit the Mark? Clin Chem. Jan. 2017;63(1):21-23. [cited by applicant]
Brennan et al., Prognostic value of myeloperoxidase in patients with chest pain. N Engl J Med. Oct. 23, 2003;349(17):1595-604. [cited by applicant]
Chapman et al., Association of High-Sensitivity Cardiac Troponin I Concentration With Cardiac Outcomes in Patients With Suspected Acute Coronary Syndrome. JAMA. Nov. 21, 2017;318(19):1913-1924. [cited by applicant]
Chapman et al., Comparison of the Efficacy and Safety of Early Rule-Out Pathways for Acute Myocardial Infarction. Circulation. Apr. 25, 2017;135(17):1586-1596. [cited by applicant]
Chen et al., A new method of quaternizing aminies and its use in amino acid and peptide chemistry. Can J Chem. 1976;54:3310-3311. [cited by applicant]
Chen et al., A specific CD36-dependent signaling pathway is required for platelet activation by oxidized low-density lipoprotein. Circ Res. Jun. 20, 2008;102(12):1512-9. [cited by applicant]
Craciun et al., Microbial conversion of choline to trimethylamine requires a glycyl radical enzyme. Proc Natl Acad Sci U S A. Dec. 26, 2012;109(52):21307-12. [cited by applicant]
Davis et al., Excess dietary lysine increases skeletal muscle and plasma trimethyllysine in rats. J Nutr. Jun. 1993;123(6):1109-16. [cited by applicant]
Delong et al., Comparing the areas under two or more correlated receiver operating characteristic curves: a nonparametric approach. Biometrics. Sep. 1988;44(3):837-45. [cited by applicant]
Dunn et al., Carnitine biosynthesis by the perfused rat liver from exogenous protein-bound trimethyllysine. Metabolism of methylated lysine derivatives arising from the degradation of 6-N-[methyl-3H]lysine-labeled glyco… [cited by applicant]
Dunn et al., Carnitine biosynthesis from gamma-butyrobetaine and from exogenous protein-bound 6-N-trimethyl-L-lysine by the perfused guinea pig liver. Effect of ascorbate deficiency on the in situ activity of gamma-buty… [cited by applicant]
Eagle et al., Identifying patients at high risk of a cardiovascular event in the near future: current status and future directions: report of a national heart, lung, and blood institute working group. Circulation. Mar. … [cited by applicant]
Fischer et al., A moderate excess of dietary lysine lowers plasma and tissue carnitine concentrations in pigs. Br J Nutr. Jan. 2009;101(2):190-6. [cited by applicant]
Giannitsis et al., Analytical validation of a high-sensitivity cardiac troponin T assay. Clin Chem. Feb. 2010;56(2):254-61. [cited by applicant]
Greer et al., Members of the H3K4 trimethylation complex regulate lifespan in a germline-dependent manner in C. elegans. Nature. Jul. 15, 2010;466(7304):383-7. [cited by applicant]
Gregory et al., Transmission of atherosclerosis susceptibility with gut microbial transplantation. J Biol Chem. Feb. 27, 2015;290(9):5647-60. [cited by applicant]
Gross. The evolution of lipidomics through space and time. Biochim Biophys Acta Mol Cell Biol Lipids. Aug. 2017;1862(8):731-739. [cited by applicant]
Handy et al., Epigenetic modifications: basic mechanisms and role in cardiovascular disease. Circulation. May 17, 2011;123(19):2145-56. [cited by applicant]
Hollander et al., State-of-the-Art Evaluation of Emergency Department Patients Presenting With Potential Acute Coronary Syndromes. Circulation. Aug. 16, 2016;134(7):547-64. [cited by applicant]
Hoppel et al., N6-Trimethyl-lysine metabolism. 3-Hydroxy-N6-trimethyl-lysine and carnitine biosynthesis. Biochem J. May 15, 1980;188(2):509-19. [cited by applicant]
Hulse et al., Carnitine biosynthesis. beta-Hydroxylation of trimethyllysine by an alpha-ketoglutarate-dependent mitochondrial dioxygenase. J Biol Chem. Mar. 10, 1978;253(5):1654-9. [cited by applicant]
Huszar. Tissue-specific biosynthesis of epsilon-N-monomethyllysine and epsilon-N-trimethyllysine in skeletal and cardiac muscle myosin: a model for the cell-free study of post-translational amino acid modifications in p… [cited by applicant]
Kind et al., Identification of small molecules using accurate mass MS/MS search. Mass Spectrom Rev. Jul. 2018;37(4):513-532. [cited by applicant]
Klingenberg et al., Improved risk stratification of patients with acute coronary syndromes using a combination of hsTnT, NT-proBNP and hsCRP with the Grace score. Eur Heart J Acute Cardiovasc Care. Mar. 2018;7(2):129-13… [cited by applicant]
Koeth et al., gamma-Butyrobetaine is a proatherogenic intermediate in gut microbial metabolism of L-carnitine to TMAO. Cell Metab. Nov. 4, 2014;20(5):799-812. [cited by applicant]
Koeth et al., Intestinal microbiota metabolism of L-carnitine, a nutrient in red meat, promotes atherosclerosis. Nat Med. May 2013;19(5):576-85. [cited by applicant]
Korley et al., Preparing the United States for high-sensitivity cardiac troponin assays. J Am Coll Cardiol. Apr. 30, 2013;61(17):1753-8. [cited by applicant]
Kouzarides. Chromatin modifications and their function. Cell. Feb. 23, 2007;128(4):693-705. [cited by applicant]
Labadie et al., Hepatic synthesis of carnitine from protein-bound trimethyl-lysine. Lysosomal digestion of methyl-lysine-labelled asialo-fetuin. Biochem J. Oct. 15, 1976;160(1):85-95. [cited by applicant]
Leening et al., Net reclassification improvement: computation, interpretation, and controversies: a literature review and clinician's guide. Ann Intern Med. Jan. 21, 2014;160(2):122-31. [cited by applicant]
Levey et al., A new equation to estimate glomerular filtration rate. Ann Intern Med. May 5, 2009;150(9):604-12. [cited by applicant]
Li et al., Gut microbiota-dependent trimethylamine N-oxide in acute coronary syndromes: a prognostic marker for incident cardiovascular events beyond traditional risk factors. Eur Heart J. Mar. 14, 2017;38(11):814-824. [cited by applicant]
Li et al., Untargeted metabolomics identifies trimethyllysine, a TMAO-producing nutrient precursor, as a predictor of incident cardiovascular disease risk. JCI Insight 2018;3(6):e99096(6). 18 pages. [cited by applicant]
Loland et al., The association between progression of atherosclerosis and the methylated amino acids asymmetric dimethylarginine and trimethyllysine. PLoS One. May 29, 2013;8(5):e64774. 9 pages. [cited by applicant]
McErlean et al., Comparison of troponin T versus creatine kinase-MB in suspected acute coronary syndromes. Am J Cardiol. Feb. 15, 2000;85(4):421-6. [cited by applicant]
Morse et al., Sites of biological methylation of proteins in cultured chick muscle cells. Biochemistry. Sep. 23, 1975;14(19):4316-25. [cited by applicant]
Muller-Bardorff et al., Improved troponin T Elisa specific for cardiac troponin T isoform: assay development and analytical and clinical validation. Clin Chem. Mar. 1997;43(3):458-66. [cited by applicant]
Nikpay et al., A comprehensive 1,000 Genomes-based genome-wide association meta-analysis of coronary artery disease. Nat Genet. Oct. 2015;47(10):1121-1130. [cited by applicant]
Peacock et al., Efficacy of High-Sensitivity Troponin T in Identifying Very-Low-Risk Patients With Possible Acute Coronary Syndrome. JAMA Cardiol. Feb. 1, 2018;3(2):104-111. [cited by applicant]
Pencina et al., Evaluating the added predictive ability of a new marker: from area under the ROC curve to reclassification and beyond. Stat Med. Jan. 30, 2008;27(2):157-72; discussion 207-12. [cited by applicant]
Pencina et al., Extensions of net reclassification improvement calculations to measure usefulness of new biomarkers. Stat Med. Jan. 15, 2011;30(1):11-21. [cited by applicant]
Pluskal et al., MZmine 2: modular framework for processing, visualizing, and analyzing mass spectrometry-based molecular profile data. BMC Bioinformatics. Jul. 23, 2010;11:395. [cited by applicant]
Porras-Yakushi et al., A novel SET domain methyltransferase modifies ribosomal protein Rpl23ab in yeast. J Biol Chem. Oct. 14, 2005;280(41):34590-8. [cited by applicant]
Purcell et al., Plink: a tool set for whole-genome association and population-based linkage analyses. Am J Hum Genet. Sep. 2007;81(3):559-75. [cited by applicant]
Qi et al., Circulating trimethylamine N-oxide and the risk of cardiovascular diseases: a systematic review and meta-analysis of 11 prospective cohort studies. J Cell Mol Med. Jan. 2018;22(1):185-194. [cited by applicant]
Rakoff-Nahoum et al., Recognition of commensal microflora by toll-like receptors is required for intestinal homeostasis. Cell. Jul. 23, 2004;118(2):229-41. [cited by applicant]
Rhein et al., Human METTL20 methylates lysine residues adjacent to the recognition loop of the electron transfer flavoprotein in mitochondria. J Biol Chem. Aug. 29, 2014;289(35):24640-51. [cited by applicant]
Roffi et al., 2015 ESC Guidelines for the management of acute coronary syndromes in patients presenting without persistent ST-segment elevation: Task Force for the Management of Acute Coronary Syndromes in Patients Pres… [cited by applicant]
Romano et al., Intestinal microbiota composition modulates choline bioavailability from diet and accumulation of the proatherogenic metabolite trimethylamine-N-oxide. mBio. Mar. 17, 2015;6(2):e02481. 8 pages. [cited by applicant]
Schiattarella et al., Gut microbe-generated metabolite trimethylamine-N-oxide as cardiovascular risk biomarker: a systematic review and dose-response meta-analysis. Eur Heart J. Oct. 14, 2017;38(39):2948-2956. [cited by applicant]
Scoumanne et al., Protein methylation: a new mechanism of p53 tumor suppressor regulation. Histol Histopathol. Sep. 2008;23(9):1143-9. [cited by applicant]
Seldin et al., Trimethylamine N-Oxide Promotes Vascular Inflammation Through Signaling of Mitogen-Activated Protein Kinase and Nuclear Factor-Kb. J Am Heart Assoc. Feb. 22, 2016;5(2):e002767. 12 pages. [cited by applicant]
Senthong et al., Trimethylamine N-Oxide and Mortality Risk in Patients With Peripheral Artery Disease. J Am Heart Assoc. Oct. 19, 2016;5(10):e004237. 14 pages. [cited by applicant]
Servillo et al., Where does N(ε)-trimethyllysine for the carnitine biosynthesis in mammals come from? PLoS One. Jan. 13, 2014;9(1):e84589. 7 pages. [cited by applicant]
Shi et al., Histone demethylation mediated by the nuclear amine oxidase homolog LSD1. Cell. Dec. 29, 2004;119(7):941-53. [cited by applicant]
Skagen et al., The Carnitine-butyrobetaine-trimethylamine-N-oxide pathway and its association with cardiovascular mortality in patients with carotid atherosclerosis. Atherosclerosis. Apr. 2016;247:64-9. [cited by applicant]
Stallcup. Role of protein methylation in chromatin remodeling and transcriptional regulation. Oncogene. May 28, 2001;20(24):3014-20. [cited by applicant]
Strand et al., Serum Carnitine Metabolites and Incident Type 2 Diabetes Mellitus in Patients With Suspected Stable Angina Pectoris. J Clin Endocrinol Metab. Mar. 1, 2018;103(3):1033-1041. [cited by applicant]
Suzuki et al., Trimethylamine N-oxide and Risk Stratification after Acute Myocardial Infarction. Clin Chem. Jan. 2017;63(1):420-428. [cited by applicant]
Tang et al., Intestinal microbial metabolism of phosphatidylcholine and cardiovascular risk. N Engl J Med. Apr. 25, 2013;368(17):1575-84. [cited by applicant]
Tang et al., Increased Trimethylamine N-Oxide Portends High Mortality Risk Independent of Glycemic Control in Patients with Type 2 Diabetes Mellitus. Clin Chem. Jan. 2017;63(1):297-306. [cited by applicant]
Thelin et al., Early rule-out of acute coronary syndrome using undetectable levels of high sensitivity troponin T. Eur Heart J Acute Cardiovasc Care. Oct. 2015;4(5):403-9. [cited by applicant]
Tsugawa et al., Hydrogen Rearrangement Rules: Computational MS/MS Fragmentation and Structure Elucidation Using MS-Finder Software. Anal Chem. Aug. 16, 2016;88(16):7946-58. [cited by applicant]
Tsugawa et al., MS-DIAL: data-independent MS/MS deconvolution for comprehensive metabolome analysis. Nat Methods. Jun. 2015;12(6):523-6. [cited by applicant]
Tsukada et al., Histone demethylation by a family of JmjC domain-containing proteins. Nature. Feb. 16, 2006;439(7078):811-6. [cited by applicant]
Ussher et al., The Emerging Role of Metabolomics in the Diagnosis and Prognosis of Cardiovascular Disease. J Am Coll Cardiol. Dec. 27, 2016;68(25):2850-2870. [cited by applicant]
Van De Werf et al., Management of acute myocardial infarction in patients presenting with ST-segment elevation. The Task Force on the Management of Acute Myocardial Infarction of the European Society of Cardiology. Eur … [cited by applicant]
Vaz et al., Carnitine biosynthesis in mammals. Biochem J. Feb. 1, 2002;361(Pt 3):417-29. [cited by applicant]
Wang et al., Gut flora metabolism of phosphatidylcholine promotes cardiovascular disease. Nature. Apr. 7, 2011;472(7341):57-63. [cited by applicant]
Wang et al., Measurement of trimethylamine-N-oxide by stable isotope dilution liquid chromatography tandem mass spectrometry. Anal Biochem. Jun. 15, 2014;455:35-40. [cited by applicant]
Wang et al., Non-lethal Inhibition of Gut Microbial Trimethylamine Production for the Treatment of Atherosclerosis. Cell. Dec. 17, 2015;163(7):1585-95. [cited by applicant]
Wang et al., Prognostic value of choline and betaine depends on intestinal microbiota-generated metabolite trimethylamine-N-oxide. Eur Heart J. Apr. 2014;35(14):904-10. [cited by applicant]
Wang et al., Protein carbamylation links inflammation, smoking, uremia and atherogenesis. Nat Med. Oct. 2007;13(10):1176-84. [cited by applicant]
Wang et al., Sharing and community curation of mass spectrometry data with Global Natural Products Social Molecular Networking. Nat Biotechnol. Aug. 9, 2016;34(8):828-837. [cited by applicant]
Wohlgemuth et al., Splash, a hashed identifier for mass spectra. Nat Biotechnol. Nov. 8, 2016;34(11):1099-1101. [cited by applicant]
Yang et al., Lipidomics: Techniques, Applications, and Outcomes Related to Biomedical Sciences. Trends Biochem Sci. Nov. 2016;41(11):954-969. [cited by applicant]
Zhang et al., Lysine methylation: beyond histones. Acta Biochim Biophys Sin (Shanghai). Jan. 2012;44(1):14-27. [cited by applicant]
Zhelev et al., Diagnostic accuracy of single baseline measurement of Elecsys Troponin T high-sensitive assay for diagnosis of acute myocardial infarction in emergency department: systematic review and meta-analysis. BMJ… [cited by applicant]
Zhu et al., Carnitine metabolism to trimethylamine by an unusual Rieske-type oxygenase from human microbiota. Proc Natl Acad Sci U S A. Mar. 18, 2014;111(11):4268-73. [cited by applicant]
Zhu et al., Gut Microbial Metabolite TMAO Enhances Platelet Hyperreactivity and Thrombosis Risk. Cell. Mar. 24, 2016;165(1):111-124. [cited by applicant]