UNC1215

UNC1215 A potent, selective antagonist of L3MBTL3 with cellular activity

This probe is available from Cayman Chemical, Sigma and Tocris
Its control compound UNC1079 is available from Cayman Chemical

  • overview
  • properties
  • selectivity profile
  • cell based assay data
  • references
  • co-crystal structures
overview

Methyl-lysine (Kme) recognition domains play a central role in epigenetic regulation during cellular differentiation, development, and gene transcription with more than 200 known “reader” domains in the human proteome. In collaboration with the Center for Integrative Chemical Biology and Drug Discovery (CICBDD) at The University of North Carolina at Chapel Hill, we have developed the first chemical probe for a Kme-binding protein.  UNC1215 is a potent and selective chemical probe for the Kme reading function of L3MBTL3, a member of the malignant brain tumor (MBT) family of chromatin interacting transcriptional repressors. UNC1215 binds the MBT domains of L3MBTL3 with a Kd of 120 nM, competitively displacing mono- or dimethyl-lysine containing peptides.  This probe is greater than 50-fold selective versus other members of the human MBT family and also demonstrates selectivity against more than 200 other Kme reader domains examined. UNC1079 is a structurally similar but substantially less potent antagonist and negative control.

properties

N-phenyl-2,5-bis[4-(pyrrolidin-1-yl)piperidine-1-carbonyl]aniline
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Physical and chemical properties
Molecular weight529.3
Molecular formulaC32H43N5O2
IUPAC NameN-phenyl-2,5-bis[4-(pyrrolidin-1-yl)piperidine-1-carbonyl]aniline
logP4.17
PSA59.13
SMILES:

C1CCN(C1)C1CCN(CC1)C(c1ccc(C(N2CCC(CC2)N2CCCC2)=O)c(c1)Nc1ccccc1)=O

InChI:

InChI=1S/C32H43N5O2/c38-31(36-20-12-27(13-21-36)34-16-4-5-17-34)25-10-11-29(30(24-25)33-26-8-2-1-3-9-26)32(39)37-22-14-28(

InChIKey:

PQOOIERVZAXHBP-UHFFFAOYSA-N

selectivity profile

Selectivity of UNC1215 against a panel of Kme reader proteins.

Alphascreen
IC50 [µM]a		MBT domainsChromodomainTudor DomainsPHD Fingers
L3MBTL1L3MBTL3L3MBTL4SFMBTMBTD1CBX753BP1UHRF1PHF23JARID1A
UNC121520.0411>306>304>30>30>30
(R2 = 0.93,
n = 17)		(R2 = 0.93,
n = 17)		(R2 = 0.63,
n = 21)		(n = 18)(R2 = 0.71,
n = 11)		(n = 19)(R2 = 0.90,
n = 12)		(n = 18)(n = 15)(n = 15)

a - The data for the IC50 values was calculated from n replicate runs; datapoints for each compound concentration were averaged and plotted using 4-parameters curve fitting. R2 is a statistical estimate of goodness-of-fit.

Molecular profiling of UNC1215

Target CategoryTarget Activity

Within methyl-lysine

reader target family

TargetIC50 (µM)aKd (µM)b
L3MBTL30.040.12
L3MBTL129.4
53BP14> 31
PHF20 TudorNT5.6
SPF30 (S.p.) TudorNTWeak binding
PHF20L1 TudorNT> 13
Histone methyltransferases< 50% inhibition at 250 mM versus 10 targetsc
BromodomainsTm shift < 0.5 °C at 10 mM versus 12 targetsd
Histone demethylasesTm shift < 0.1 °C at 50 mM versus 15 targetsd

NIMH Psychoactive Drug

Screening Program

Selectivity Panel  

< 50% inhibition at 10 mM versus 35 targets

> 50% inhibition at 10 mM versus 8 targets

TargetKi (µM)eIC50 (µM)
Alpha 2C0.86NT
DAT> 10NT
KOR4.0NT
M10.0973.6f
M20.07230% at 30 µMg
M30.89NT
M40.40NT
M54.3NT
Kinase Selectivity Panel< 15% inhibition at 10 mM versus 49 kinases
Target% Inhibition at 10 mM
FLT364%

aAlphascreen assays results. bITC results. cRadioactive SAM methyl transfer assay results. dDifferential scanning fluorimetry results. eRadioligand binding assay results. fCa2+ mobilization assay results. gcAMP biosensor assay results.

in vitro potency
cell based assay data

UNC1215 potently antagonizes 3MBT localization in cells.  (a) Recovery time of a photobleached area in GFP-3MBT expressing cells is reduced upon treatment with UNC1215 in a dose response manner, whereas inactive compound UNC1079 shows no effect.  Inset shows time lapse images of photobleached nuclei.  (b, c) GFP fusions of 3MBT and FLMBT localize to the nucleus and form distinct foci in HEK293 cells. UNC1215 inhibits the foci formation of GFP-3MBT in a dose response fashion, whereas UNC1079 has no effect on the foci. In contrast, UNC1215 is relatively ineffective at inhibiting foci formation of GFP-FLMBT (scale bar, 10 µm). (d) The GFP-3MBT D274A MBT1 mutant shows a reduction in foci formation, while the GFP-3MBT D381A MBT2 mutant does not form nuclear foci.

references

Discovery of a chemical probe for a methyl-lysine reader domain: L3MBTL3

James, Lindsey I.; Barsyte-Lovejoy, Dalia; Zhong, Nan; Krichevsky, Liubov; Korboukh, Victoria K.; Herold, J. Martin; MacNevin, Christopher J.; Norris, Jacqueline L.; Sagum, Cari A.; Tempel, Wolfram; Marcon, Edyta; Guo, Hongbo; Gao, Cen; Huang, Xi-Ping; Duan, Shili; Emili, Andrew; Greenblatt, Jack F.; Kireev, Dmitri B.; Jin, Jian; Janzen, William P.; Brown, Peter J.; Bedford, Mark T.; *Arrowsmith, Cheryl H. *Frye, Stephen V. Nature Chemical Biology 9, 184–191 (2013) -  DOI 10.1038/nchembio.1157

pk properties
co-crystal structures

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Main features

  • L3MBTL3 Dimer: UNC1215 binds to L3MBTL3 with a 2:2 stoichiometry. Each "arm" of UNC1215 binds to a different molecule of L3MBTL3.
  • Pocket Key interactions of pyrrolidine A with YYF aromatic cage and pyrrolidine B with YFW aromatic cage. Both interactions are augmented by a hydrogen bond with an Asp.
synthetic schemes
materials and methods
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