Development and implementation of a high-throughput AlphaLISA assay for identifying inhibitors of EZH2 methyltransferase

Assay and Drug Development Technologies

Volumn 11, Issue 3, 2013, Pages 152-162

  Simard, Jeffrey R ^a   Plant, Matthew ^a   Emkey, Renee ^a   Yu, Violeta ^a  

^a AMGEN INC   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

BUFFER; EMBRYONIC ECTODERM DEVELOPMENT PROTEIN; ENZYME INHIBITOR; HISTONE H3; HISTONE H3 LYSINE 27; POLYLYSINE; TRANSCRIPTION FACTOR EZH2; UNCLASSIFIED DRUG;

ARTICLE; ENZYMATIC ASSAY; ENZYME ACTIVITY; ENZYME INHIBITION; ENZYME MECHANISM; HIGH THROUGHPUT SCREENING; HUMAN; METHYLATION; PERFORMANCE; PROCESS OPTIMIZATION; SENSITIVITY ANALYSIS; SIGNAL DETECTION; VALIDATION PROCESS;

ALGORITHMS; ANTIBODIES; BUFFERS; DATA INTERPRETATION, STATISTICAL; ENZYME INHIBITORS; FLUORESCENCE POLARIZATION IMMUNOASSAY; HIGH-THROUGHPUT SCREENING ASSAYS; HUMANS; INDICATORS AND REAGENTS; METHYLATION; PEPTIDES; POLYCOMB REPRESSIVE COMPLEX 2; RECOMBINANT PROTEINS; REFERENCE STANDARDS; REPRODUCIBILITY OF RESULTS; STREPTAVIDIN;

EID: 84875978605     PISSN: 1540658X     EISSN: 15578127     Source Type: Journal    
DOI: 10.1089/adt.2012.481     Document Type: Article

References (42)

1. Daub H, Specht K, Ullrich A: Strategies to overcome resistance to targeted protein kinase inhibitors. Nat Rev Drug Discov 2004;3:1001-1010.
2. Zhang JF, Kim JS: Small-molecule fluorescent chemosensors for Hg2+ ion. Anal Sci 2009;25:1271-1281.
3. Comley J: Epigentics: an emerging target class for drug screening. Drug Discov World 2011;12(Spring 2011):40-55.
4. Loyola A, Almouzni G: Marking histone H3 variants: how, when and why? Trends Biochem Sci 2007;32:425-433.
5. Arrowsmith CH, Bountra C, Fish PV, Lee K, Schapira M: Epigenetic protein families: a new frontier for drug discovery. Nat Rev Drug Discov 2012;11:384-400.
6. Cao R, Zhang Y: SUZ12 is required for both the histone methyltransferase activity and the silencing function of the EED-EZH2 complex. Mol Cell 2004;15:57-67.
7. Cao R, Zhang Y: The functions of E(Z)/EZH2-mediated methylation of lysine 27 in histone H3. Curr Opin Genet Dev 2004;14:155-164.
8. Czermin B, Melfi R, McCabe D, Seitz V, Imhof A, Pirrotta V: Drosophila enhancer of Zeste/ESC complexes have a histone H3 methyltransferase activity that marks chromosomal polycomb sites. Cell 2002;111:185-196.
9. Kirmizis A, Bartley SM, Kuzmichev A, et al.: Silencing of human polycomb target genes is associated with methylation of histone H3 Lys 27. Genes Dev 2004;18:1592-1605.
10. Kuzmichev A, Nishioka K, Erdjument-Bromage H, Tempst P, Reinberg D: Histone methyltransferase activity associated with a human multiprotein complex containing the enhancer of Zeste protein. Genes Dev 2002;16:2893-2905.
11. Muller J, Hart CM, Francis NJ, et al.: Histone methyltransferase activity of a Drosophila polycomb group repressor complex. Cell 2002;111:197-208.
12. Fussbroich B, Wagener N, Macher-Goeppinger S, et al.: EZH2 depletion blocks the proliferation of colon cancer cells. PLoS One 2011;6:e21651.
13. Kleer CG, Cao Q, Varambally S, et al.: EZH2 is a marker of aggressive breast cancer and promotes neoplastic transformation of breast epithelial cells. Proc Natl Acad Sci USA 2003;100:11606-11611.
14. Simon JA, Kingston RE: Mechanisms of polycomb gene silencing: knowns and unknowns. Nat Rev Mol Cell Biol 2009;10:697-708.
15. Simon JA, Lange CA: Roles of the EZH2 histone methyltransferase in cancer epigenetics. Mutat Res 2008;647:21-29.
16. Varambally S, Dhanasekaran SM, Zhou M, et al.: The polycomb group protein EZH2 is involved in progression of prostate cancer. Nature 2002;419: 624-629.
17. Morin RD, Johnson NA, Severson TM, et al.: Somatic mutations altering EZH2 (Tyr641) in follicular and diffuse large B-cell lymphomas of germinal-center origin. Nat Genet 2010;42:181-185.
18. Sneeringer CJ, Scott MP, Kuntz KW, et al.: Coordinated activities of wild-type plus mutant EZH2 drive tumor-associated hypertrimethylation of lysine 27 on histone H3 (H3K27) in human B-cell lymphomas. Proc Natl Acad Sci USA 2010;107:20980-20985.
19. Margueron R, Justin N, Ohno K, et al.: Role of the polycomb protein EED in the propagation of repressive histone marks. Nature 2009;461:762-767.
20. Martinez-Garcia E, Licht JD: Deregulation of H3K27 methylation in cancer. Nat Genet 2010;42:100-101.
21. Tsang DP, Cheng AS: Epigenetic regulation of signaling pathways in cancer: role of the histone methyltransferase EZH2. J Gastroenterol Hepatol 2011;26: 19-27.
22. Xu C, Bian C, Yang W, et al.: Binding of different histone marks differentially regulates the activity and specificity of polycomb repressive complex 2 (PRC2). Proc Natl Acad Sci USA 2010;107:19266-19271.
23. Dorgan KM, Wooderchak WL, Wynn DP, et al.: An enzyme-coupled continuous spectrophotometric assay for S-adenosylmethionine-dependent methyltransferases. Anal Biochem 2006;350:249-255.
24. Hendricks CL, Ross JR, Pichersky E, Noel JP, Zhou ZS: An enzyme-coupled colorimetric assay for S-adenosylmethionine-dependent methyltransferases. Anal Biochem 2004;326:100-105.
25. Gauthier N, Caron M, Pedro L, et al.: Development of homogeneous nonradioactive methyltransferase and demethylase assays targeting histone H3 lysine 4. J Biomol Screen 2012;17:49-58.
26. Quinn AM, Allali-Hassani A, Vedadi M, Simeonov A: A chemiluminescence- based method for identification of histone lysine methyltransferase inhibitors. Mol Biosyst 2010;6:782-788.
27. Kruidenier L, Chung CW, Cheng Z, et al.: A selective jumonji H3K27 demethylase inhibitor modulates the proinflammatory macrophage response. Nature 2012; 488:404-408.
28. Wigle TJ, Herold JM, Senisterra GA, et al.: Screening for inhibitors of low-affinity epigenetic peptide-protein interactions: an AlphaScreen-based assay for antagonists of methyl-lysine binding proteins. J Biomol Screen 2010;15: 62-71.
29. Collazo E, Couture JF, Bulfer S, Trievel RC: A coupled fluorescent assay for histone methyltransferases. Anal Biochem 2005;342:86-92.
30. Wang C, Leffler S, Thompson DH, Hrycyna CA: A general fluorescence-based coupled assay for S-adenosylmethionine-dependent methyltransferases. Biochem Biophys Res Commun 2005;331:351-356.
31. Han Z, Xing X, Hu M, Zhang Y, Liu P, Chai J: Structural basis of EZH2 recognition by EED. Structure 2007;15:1306-1315.
32. Smith TF, Gaitatzes C, Saxena K, Neer EJ: The WD repeat: a common architecture for diverse functions. Trends Biochem Sci 1999;24:181-185.
33. Choi Y, Seeliger MA, Panjarian SB, et al.: N-myristoylated c-Abl tyrosine kinase localizes to the endoplasmic reticulum upon binding to an allosteric inhibitor. J Biol Chem 2009;284:29005-29014.
34. Habig M, Blechschmidt A, Dressler S, et al.: Efficient elimination of nonstoichiometric enzyme inhibitors from HTS hit lists. J Biomol Screen 2009;14:679-689.
35. Li H, Huang J, Jiang X, Seefeld M, McQueney M, Macarron R: The effect of triton concentration on the activity of undecaprenyl pyrophosphate synthase inhibitors. J Biomol Screen 2003;8:712-715.
36. PerkinElmer Reagents Technical Support Wiki: The hook effect. Available at: https://perkinelmerreagents.onconfluence.com/display/ts/The + hook + effect (last accessed on May 23, 2012).
37. Richon VM, Johnston D, Sneeringer CJ, et al.: Chemogenetic analysis of human protein methyltransferases. Chem Biol Drug Des 2011;78:199-210.
38. Yu V, Fisch T, Long AM, et al.: High-throughput TR-FRET assays for identifying inhibitors of LSD1 and JMJD2C histone lysine demethylases. J Biomol Screen 2012;17:27-38.
39. Daylight Chemical Information Systems, Inc. Available at: www.daylight.com (last accessed on May 23, 2012).
40. PerkinElmer Reagents Technical Support Wiki: Buffer selection for Alpha assays. Available at: https://perkinelmerreagents.onconfluence.com/display/ts/ Buffer+selection+for+Alpha+assays (last accessed on May 23, 2012).
41. Baell JB, Holloway GA: New substructure filters for removal of pan assay interference compounds (PAINS) from screening libraries and for their exclusion in bioassays. J Med Chem 2010;53:2719-2740.
42. Diaz E, Machutta CA, Chen S, et al.: Development and validation of reagents and assays for EZH2 peptide and nucleosome high-throughput screens. J Biomol Screen 2012;17:1279-1292.