RGFP966

Reagent Authentication Certificate:

Chemical Probes Portal RGFP966.pdf

Protein target name:

HDAC3

Mechanism of action:

Inhibitor

Recommended Concentration for use in cells:

100 nM - 1 uM

Probe Availability

MedChem Express (1357389-11-7)

Abcam (1396841-57-8)

Cayman (1357389-11-7)

Probe Information

Target and Probe Information

Target Details

Target class:

Epigenetics

Subclass:

HDAC

HUGO ID:

HGNC:4854

Entrez gene ID:

8841

UNIPROT ID:

O15379

Target alias:

Histone deacetylase 3, HD3, RPD3, RPD3-2

Probe Details

Chemical Probes Portal ID:

CPP1484

Probe alias:

RGFP 966; RGFP-966

PubChem CID:

56650312

SMILES string:

C1=CC=C(C=C1)/C=C/CN2C=C(C=N2)/C=C/C(=O)NC3=C(C=C(C=C3)F)N

InChi Key:

BLVQHYHDYFTPDV-VCABWLAWSA-N

Orthogonal probe:

Romidepsin

Validation in Cells and Model Organisms

In vitro validation

On target activity

Potency:

IC50

0.08 uM

Potency assay:

Substrate-dependent biochemical assays (Reaction Biology). Extensive biochemical characterization has been published on a related compound 106 (Chem Biol. publication): HDAD3 Ki= 14 nM; IC50 with recombinant HDAC3=0.79 uM.

Structure-activity relationship:

Yes, several compounds on the pimelic diphenylamide scaffold have been reported with variable activity and specificity for HDAC proteins (see PLoS One report).

Off target activity

Potency (off target):

Off target protein and potency:

HDAC1: 148 nM

Potency assay (off target):

In Reaction Biology biochemical assay, RGFP966 had no activity on HDACs other than HDAC3 up to 15 uM. The related compound (106): is 10-15-fold selective for HDAC3 > HDAC1 and HDAC2, and only weakly active against HDAC8. In experiments with recombinant HDAC1, IC50= 0.24 uM. Using a modified version of 106 for proteomic analysis after photo-crosslinking of recombinant HDACs (HDAC1, 2, 3, 4, 5, and 8), the compound was selective for HDAC3, with low level cross-linking with HDAC1.

In cell validation

On target activity in cells

Potency in cells:

IC50

1.3 uM

Potency assay (cells):

General HDAC inhibition assay in HeLa cells.

Target engagement assay (cells):

Yes, direct but with the related compound (106), which was tested in chemoproteomic assays in nuclear extracts. In these experiments, the 106 derivative only interacted with HDAC3 after photo-crosslinking.

Off target activity in cells

Potency assay, off target (cells):

The related compound (106) was tested in chemoproteomic assays in nuclear extracts. In these experiments, the 106 derivative interacted with only HDAC3 after photo-crosslinking.

In vivo validation

Organism:

C57BL/6J male mice

Dose:

10 mg/mL

Route of delivery:

Subcutaneous

Ratio of probe in organ to plasma (O:P):

0.45

Cmax:

1.25 ug/g

Tmax:

0.5 h

Organ of Interest

Organ (O):

brain

Target engagement assay:

Not done

Primary Reference

Reference (doi):

10.1016/j.chembiol.2009.07.010 10.1073/pnas.1213364110 Patent WO 2012016081 A2

Reference (PMID):

19778726 23297220

Endorsed probe

The Chemical Probes Portal endorses this compound as chemical probe for use in cells and in model organisms.

SAB Rating

Rating for use in Cells

3 review(s)

Rating for use in Model Organisms

3 review(s)

SAB Members

SAB Comments

This is an N-(o-amino-phenyl)carboxamide HDAC3-selective inhibitor with an IC50 of 80 nM and 'no effective inhibition of other HDACs' at concentrations up to 15 uM utilizing a substrate-dependent biochemical assay with recombinant human protein (Malvaez et al PNAS 2013; 110: 2647-2652 - primary data not shown). However, functional data suggests possible inhibition of other class I HDACs at concentrations >1 uM (Matthews et al Blood 2015; 126: 2392-2403) - please refer to 'cellular concentration notes'. For in vivo dosing, 10 mg/kg delivered subcutaneously achieves a maximum tissue concentration (brain) in mice of 3.15 uM at 30 minutes, with concentrations exceeding the IC50 for HDAC3 for at least 2 hours in this tissue.

Jun 1 2017 - 1:56pm

RGFP966 has been demonstrated to have selective activity for HDAC3 in biochemical assays over most other HDAC enzymes. In cells, Matthews et al., (Blood, 2015) observed that RGFP966 treatment phenocopied genetic HDAC3 perturbation up to ~1,000 nM. Thus, cellular studies using RGFP966 for selective pharmacologic HDAC3 perturbation should be limited to concentrations <1,000 nM. In mouse and rat models, RGFP966 has been observed to be brain-penetrant when dosed subcutaneously. When used for in vivo studies, care should be taken to pair phenotypic observations from genetic-perturbation studies of HDACs in order to have higher confidence in HDAC3-selective effects.

Jun 13 2017 - 2:11pm

RGFP966

Probe Availability

Probe Information

Target Details

Histone deacetylase 3, HD3, RPD3, RPD3-2

Probe Details

In vitro validation

In cell validation

In vivo validation

Primary Reference

Supporting Organizations

Interested in supporting the portal?

Search form

RGFP966

Probe Availability

Probe Information

Target Details

Histone deacetylase 3, HD3, RPD3, RPD3-2

Probe Details

In vitro validation

In cell validation

In vivo validation

Primary Reference

Supporting Organizations

Interested in supporting the portal?