UNC3866 : Antagonist of CBX7, CBX4



  • CBX7
  • CBX4
  • Antagonist
  • 30 uM

In Vitro Validations

Uniprot ID: O95931
Target Class: Epigenetic
Target SubClass: Methyl-lysine reader
Potency: Kd
Potency Value: 97 nM
Potency Assay: Isothermal titration calorimetry (ITC)
PDB ID for probe-target interaction (3D structure): 5EPJ
Structure-activity relationship: Detailed SAR data are available in a J. Med. Chem. publication that should be available shortly. SAR data were determined using an AlphaScreen assay, which is a bead-based competition assay that utilizes His-tagged proteins and biotinylated-peptide bait ligands. The SAR study primarily explored ligand length, unnatural methyl-lysine mimetics, and N-terminal modifications, as well as modifications to the phenylalanine side chain, alanine side chain, and C terminus. We found that certain tertiary amine mimetics of trimethyl-lysine are able to maintain similar affinities for chromodomains as Kme3 and Kme3 derivatives are not active in cells. A thorough examination of the N terminus of our inhibitors has also begun to establish the requirements for optimizing potency and selectivity for the Pc, HP1 and CDY families of chromodomains. CBX4/7 show a strong preference for alkyl substituents located at the para-position of the N-terminal benzoyl cap.
Target aliases:
Chromobox protein homolog 7, CBX7, CBX7_HUMAN

DOI Reference: 10.1038/nchembio.2007

In Cell Validations

In Vivo Data

No in Vivo Validations

Off-Target Selectivity Assesments

Potency end-point : KD CBX2: 1.8 uM, CBX6: 0.610 uM, CBX8: 1.2 uM, CDY1: 6.3 uM, CDYL1b: 0.91 uM, CDYL2: 0.85 uM
Potency assay (off target): Isothermal titration calorimetry (ITC). We also profiled the compound on a microarray containing 96 epigenetic reader proteins across multiple domain types.
Probe Selectivity in Vitro:

UNC3866 is selective over a broad range of chromatin-associated proteins (profiled versus 100+ Kme readers, 52 bromodomains, 33 lysine methyltransferases, and 7 demethylase enzymes). Additionally, UNC3866 was profiled against 49 GPCRs, 5 ion channels and 3 transporter proteins. In the Nature Chemical Biology publication, see Figures 2-3, Supplementary Figures 3-8, and Supplementary Tables 2-5.

Potency in cells, off target :
Potency assay, off target (cells): Assays to assess the cellular consequences of engaging the other non-PRC1 chromodomain targets (CDYs) of UNC3866 have not been performed since these are relatively uncharacterized targets.
Probe Selectivity in Cell:
Assays to assess the cellular consequences of engaging the other non-PRC1 chromodomain targets (CDYs) of UNC3866 have not been performed since these are relatively uncharacterized targets.
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SERP ratings and comments

SERP Ratings

In Cell Rating

SERP Comments:

The probe UNC3866 has been well characterized for its interaction with the desired target CBX7 using ITC, a biotinylated analog, and co-crystal stuctures to provide strong evidence of interactions. Cellular penetration is low (5% of external concentration), meaning high concentrations are needed. A closely related, inactive analog provides evidence that the probe's biological effects are target related. However, the probe is active against other members of the CBX chromodomains (equipotent at CDX4) and also CDY choromodomains so attributing effects to a specific interaction in cells is not possible. The probe has been assessed for pharmacokinetics in mice (IP injection) and shows evidence of moderate clearance that may allow for use in vivo. The high concentrations required for cellular assays would be difficult to achieve in vivo, but the authors propose to examine this possible use in the future.

(last updated: 23 Oct 2016 )

SERP Ratings

In Cell Rating

SERP Comments:

This probe is well-characterized and somewhat selective for CBX4 and CBX7 in biochemical assays, though it is only 6-fold less active against CBX6 and about 10-fold less active against CDYL1b and CDYL2. It also has low cell permeability, necessitating high concentrations to be used in cell assays. This means that any biological effects observed in cell assays can not be attributed solely to inhibition of CBX4 and CBX7.  

(last updated: 12 Jan 2017 )

SERP Ratings

In Cell Rating

SERP Comments:

UNC3866 is a first in class inhibitor for CBX7 and CBX4, as well as other related family members. It's highly potent in vitro and represents a great advancement in the CBX protein biology field. The compound was shown to inhibit cell proliferation at 7.6 uM, and at 30 uM, to induce a cell senescence phenotype that was linked to the target by data published elsewhere. A biotinylated version of the compound can pull down the PRC1 complex components, an effect that is inhibited if the PC3 cells were incubated with 30 uM of UNC3866 for 24 h prior to lysis and pulldown. Data on the cellular compound pulldowns was focused on the known targets instead of a broader proteomic profiling of the targets bound to UNC3866. Additional data on UNC3866's effect on more specific cellular activity readout such as nanoBRET or ChIP would be very informative.

(last updated: 21 Jan 2017 )