GSK484 hydrochloride, Inhibitor | GSK484, inhibitor

GSK484 hydrochloride

Cat. Num. AAA5764324

• Purity: 99.36%
• Synonyms: GSK484 hydrochloride; GTPL8577; GSK484; inhibitor

• Purity/Purification: 99.36%
• Form/Format: Solid. Powder

• Solubility: DMSO: 60mg/mL (117.64mM) Need ultrasonic; (<1mg/ml refers to the product slightly soluble or insoluble)
• Formula: C27H30ClN5O3
• SMILES: Cl.COC1=CC(=CC2=C1N(C)C(=N2)C1=CC2=CC=CC=C2N1CC1CC1)C(=O)N1CC[C@@H](O)[C@@H](N)C1
• CAS Number: 1652591-81-5
• Cell Research: HEK293 cells stably expressing N-terminal DYKDDDDK-tagged PAD1, PAD2, PAD3 or PAD4 are engineered by retroviral transduction. Cells are grown in 15cm diameter plates to subconfluency in DMEM supplemented with 10% Foetal Bovine Serum, harvested by centrifugation and washed once in PBS/2mM EGTA. Cells are lysed in 50mM Tris-Cl, pH 7.4, 1.5mM MgCl2, 5% glycerol, 150mM NaCl, 25mM NaF, 1mM Na3VO4, 0.4% NP40, 1mM DTT with protease inhibitors. Lysates are pre-incubated for 20 minutes at 4 degree C with DMSO alone (2%), 100uM of GSK199, GSK484, GSK106 or 200uM Cl-amidine. Citrullination reactions are performed for 30 minutes at 37 degree C in the presence of 2mM calcium. Extracts are loaded on to gels, proteins separated by SDS-PAGE and transferred to PVDF membranes. Citrullinated proteins are then chemically modified and detected using an anti-modified citrulline antibody. DYKDDDDK-PAD constructs are detected using the anti-DYKDDDDK antibody [1].
• Kinase Assay: PAD4 is serially diluted in the presence of 10nM GSK215 in assay buffer (100mM HEPES, pH 8, 50mM NaCl, 5% glycerol, 1mM CHAPS, 1mM DTT) at varying concentrations of calcium (0, 0.2, 2 and 10mM). Following incubation for 50 minutes, apparent Kds for each calcium concentration are determined using a single site saturation curve. For IC50 determination, GSK484 is serially diluted in DMSO (1% final assay concentration) and tested at the same range of calcium concentrations in the presence of PAD4 (at the calculated Kd for each calcium condition) and 10nM GSK215 in the same assay buffer and volume. Reactions are incubated for 50 min after which IC50 values are calculated using a four-parameter logistic equation [1].
• In Vitro: To address whether PAD4 inhibition can suppress cancer-associated kidney injury,mMTV-PyMT mice are treated with the PAD4 inhibitor GSK484 at 4mg/kg daily for one week. This dose suppresses the elevated number of neutrophils undergoing NETosis in peripheral blood in mice with cancer. In parallel, the total protein level in urine frommMTV-PyMT mice is significantly reduced compared with untreated tumor-bearing mice. Administration of GSK484 at a dose of 4mg/kg daily during one week reverts signs of kidney dysfunction in tumor-bearing mice to the same extent as DNase I treatment without any detectable signs of toxicity [2].
• Preparation and Storage: Store at -20 degree C for 3 years powder; -80 degree C for 2 years in solvent

Related Product Information for GSK484, inhibitor

GSK484 hydrochloride (GTPL8577) is a specifc and reversible peptidyl arginine deiminase 4 (PAD4) inhibitor. It demonstrates high-affinity binding to PAD4 with IC50s of 50nM in the absence of Calcium.

Product Categories/Family for GSK484, inhibitor

Epigenetics

References

Lewis HD, et al. Inhibition of PAD4 activity is sufficient to disrupt mouse and human NET formation. Nat Chem Biol. 2015 Mar;11(3):189-91.
Lewis HD, et al. Inhibition of PAD4 activity is sufficient to disrupt mouse and human NET formation. Nat Chem Biol. 2015 Mar;11(3):189-91.
Cedervall J, et al. Pharmacological targeting of peptidylarginine deiminase 4 prevents cancer-associated kidney injury in mice. Oncoimmunology. 2017 Apr 20;6(8):e1320009.
Cedervall J, et al. Pharmacological targeting of peptidylarginine deiminase 4 prevents cancer-associated kidney injury in mice. Oncoimmunology. 2017 Apr 20;6(8):e1320009.