PDD00017273

Reagent Authentication Certificate:

Chemical Probes Portal PDD00017273.pdf

Protein target name:

PARG

Mechanism of action:

PolyADP ribose (PAR) competitive inhibitor

Recommended Concentration for use in cells:

Up to 10 uM, or 10X the cellular IC50 depending on the cell line

Probe Availability

Tocris (1952247-05-0)

Probe Information

Target and Probe Information

Target Details

Target class:

DNA replication & repair enzyme

HUGO symbol:

PARG

Entrez gene ID:

8505

Target alias:

Poly (ADP-Ribose) Glycohydrolase, PARG99

Probe Details

Chemical Probes Portal ID:

CPP745

Probe alias:

SCHEMBL17825271

PubChem CID:

121398766

SMILES string:

CC1=NN(C(=C1)CN2C3=C(C=C(C=C3)S(=O)(=O)NC4(CC4)C)C(=O)N(C2=O)CC5=CN=C(S5)C)C

InChi Key:

IFWUBRBMMNTBRZ-UHFFFAOYSA-N

Control compound:

N-Methyl-PDD00017273

Physico-chemical properties

Solubility - preferred solvent:

DMSO, aqueous + 1% DMSO

Solubility - concentration:

>200 uM, aqueous >100 uM

Potential reactivities:

None identified to date

NMR:

1H NMR (300MHz, DMSO-d6) δ = 8.44 (d, J=2.3 Hz, 1H), 8.23 (s, 1H), 8.03 (dd, J=2.3, 8.9 Hz, 1H), 7.64 (s, 1H), 7.56 (d, J=8.9 Hz, 1H), 5.76 (s, 1H), 5.38 (s, 2H), 5.26 (s, 2H), 3.82 (s, 3H), 2.58 (s, 3H), 1.99 (s, 3H), 1.07 (s, 3H), 0.63 - 0.55 (m, 2H), 0.42 - 0.36 (m, 2H).

13C NMR (75MHz, CHLOROFORM-d) δ = 159.80, 150.06, 147.82, 142.72, 141.96, 137.85, 136.46, 133.54, 131.27, 128.79, 125.23, 115.75, 115.07, 105.35, 39.63, 37.38, 36.67, 31.81, 24.61, 18.97, 14.07 (2C), 13.32

Storage Recommendations

Storage (dry powder):

Storage (solution):

Desiccate or freeze

Validation in Cells and Model Organisms

In vitro validation

On target activity

Potency:

IC50

26 nM

Potency assay:

Biochemical PAR chain persistence assay

Off target activity

Off target protein and potency:

PARP1 IC50 >30µMARH3 IC50 >30µM

Potency assay (off target):

Biochemical PAR chain persistence assay

Selectivity against non-family targets:

Biochemical selectivity against safety targets (% inhibition at 10 µM, binding assay Eurofins Panlabs): Adenosine A1=1%, Adenosine A2A=-1%, Adrenergic a1A=-5%, Adrenergic a1B=7%, Adrenergic a2A=10%, Adrenergic β1=-9%, Adrenergic β2=-9%, Calcium Channel=L Type=16%, Cannabinoid CB1=21%, Dopamine D1=6%, Dopamine D2S=-9%, GABAA=Flunitrazepam Central=4%, GABAA=Muscimol Central=3%, Glutamate=NMDA=4%, Histamine H1=-6%, Imidazoline I2=Central=-18%, Muscarinic M2 =-1%, Muscarinic M3 =3%, Nicotinic Acetylcholine=3%, Nicotinic Acetylcholine=a1=-3%, Opiate µ(OP3=MOP)=-6%, Phorbol Ester=-11%, Potassium Channel [KATP]=5%, Potassium Channel hERG=12%, Prostanoid EP4=11%, Rolipram=16%, Serotonin (5-Hydroxytryptamine) 5-HT2B=4%, Sigma σ1=15%, Sodium Channel site 2=6%, Transporter=Norepinephrine (NET)=20%, Acetyl Cholinesterase=-1%, Cyclooxegenase COX-2=-2%, Phosphodiesterase PDE4=17%,

In cell validation

On target activity in cells

Potency in cells:

IC50

37 nM

Potency assay (cells):

Whole-cell PAR chain-persistence assay in HeLa cells after MMS challenge.

Target engagement assay (cells):

Antibody-based PAR chain-persistence assay

Toxicity

Cytoxicity assay:

Yes

Notes on cytotoxicity:

On target: Yes, in clonogenic assays, depending on the genetic background (7-21-day assay, dependent upon cell line and replicative rate). Example data: ZR-75-1 cells, IC50=200 nM; MDA-MB436 cells, IC50=800 nM; HCC1937 cells, IC50>10 µM) In the absence of a DNA-damaging agent, the compound is not cytotoxic below 30 µM in HeLa cells, for up to 72 hrs.

Primary Reference

Reference (doi):

10.1021/acschembio.6b00609

Reference (PMID):

27689388

This probe is in the process of SAB review. Please continue to check back for new reviews and commentary.

SAB Rating

Rating for use in Cells

3 review(s)

Rating for use in Model Organisms

3 review(s)

SAB Members

SAB Comments

PD00017273 is an inhibitor of human PARG enzyme with IC50 of 26 nM and good selectivity over the two closest functional homologues, PARP1 and ARH3 (>100 μM). It is suitable for use in cellular assays (cell permeability data not available) and is not cytotoxic in HeLa cells up to 30 μM; however, different sensitivities in different cell lines are possible. Interestingly, the negative control (compound 6) also shows some weak activity in the Promega Glo assay. Off-target activity is not completely ruled out, since the compound was tested on a relatively small panel of 33 receptors and enzymes at 10 μM. PD00017273 is far a better compound compared with the previously reported PARG inhibitor, and it can used as chemical tool to investigate PARG biology. Due to high intrinsic clearance in HLM, the compound is not suitable for use for any in vivo studies.

PDD00017273 is a potent inhibitor of PARG in cells (EC50=37 nM in a cellular PAR chain persistence assay) with little toxicity observed at 10 µM. While some degree of selectivity screening has been performed—for example, PDD00017273 is inactive at the related protein ARH3 and also at PARP1—a larger body of characterization data would be preferred for the informed deconvolution of cellular usage of the compound. The availability of a negative control compound inactive at the primary target increases the utlity of the compound. PDD00017273 shows poor metabolic stability in human liver microsomes and is thus unlikely to be useful as an in vivo probe.

PDD00017273

Probe Availability

Probe Information

Target Details

Poly (ADP-Ribose) Glycohydrolase, PARG99

Probe Details

Physico-chemical properties

In vitro validation

In cell validation

Toxicity

Primary Reference

Supporting Organizations

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PDD00017273

Probe Availability

Probe Information

Target Details

Poly (ADP-Ribose) Glycohydrolase, PARG99

Probe Details

Physico-chemical properties

In vitro validation

In cell validation

Toxicity

Primary Reference

Supporting Organizations

Interested in supporting the portal?