Epigenetics
times than other traditional label-free technologies (6-10 second cycles), greatly increasing the efficien- cy of screening programmes for epigenetic and other enzyme targets (Figures 11 and 12).
Figure 12:The Biocius RapidFire® RF300 Mass Spectrometry System processes epigenetic samples in 6-8 seconds each
A range of target assays are available at BioFocus (
www.biofocus.com) with a focus on histone methyltransferases (HMTs) and HDACs, with his- tone demethylases, ubiquitin E3 ligases and deu- biquitnases also available. For HMT hit finding campaigns, a subset of its 900,000 diverse com- pound library is screened typically with a radio- metric assay format. Using this approach BioFocus has recently identified a number of novel chemo- types which were progressed to optimisation. For further characterisation of hits, orthogonal assay technologies, such as Caliper lab-on-a-chip, fluo- rescence lifetime technology and binding assays have successfully been used. BioFocus has success- fully employed protein family structural superposi- tion followed by sequence alignment to generate chemogenomic models that have been developed into protein family ‘roadmaps’. Key residues are identified, aiding the understanding of pocket- based family homology and selectivity. Using the information of such models enables the under- standing of protein targets for which structural information is not available. BioFocus uses this ensemble of information to design new target-fam- ily focused libraries, to prioritise compounds for specific assays (eg hit expansion) and/or to predict potential selectivity targets. It has used this approach to analyse the histone methyl transferase (HMT) sub-family, protein lysine methyltransferas- es (PKMTs). Structural superposition reveals high conservation in the primary and secondary struc- ture around the S-adenosyl methionine (SAM) pocket. This observation can help prioritise com- pounds for screening; for example, adenosine is present in both ATP and SAM, therefore screening minimally functionalised compounds designed for the ATP site in kinases may provide a useful com- ponent of a screening deck for hit identification against this novel protein class (Figure 13).
Figure 13: Post-translational modifications of histone protein that can influence gene transcription. BioFocus supports drug discovery for various epigenetic target families including histone methyltransferases, histone deacetylases, histone demethylases, ubiquitin E3 ligases and deubiquitnases. Abbreviations: Me, methyl; Ac, acetyl; Ub, ubiquitin; PO4-, phosphate
46
BPS Bioscience (
www.bpsbioscience.com) manu- factures a broad portfolio of sensitive, specific epi- genetic assay kits designed to determine enzyme activity or for inhibitor screening. Its portfolio includes both chemiluminescent 96- or 384-well plate assays and homogeneous AlphELISA/ AlphaScreen assays. Many epigenetic enzymes are present in cells as complexes of multiple regulatory subunits, so they can be difficult to express as func- tional enzymes. For example, its EZH2 complex is
Drug Discovery World Spring 2011
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