Name: Cl-amidine
Brand: Selleck Chemicals
SKU: S8141
Rating: 5 (28 reviews)

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Quality Control

Batch: Purity: 99.89%

99.89

Related Targets	PD-1/PD-L1 CXCR STING AhR CD markers Interleukins Anti-infection Antioxidant COX Histamine Receptor
Other Immunology & Inflammation related Inhibitors	Bestatin (Ubenimex) Bindarit (AF 2838) Tranilast Tempol Sinomenine GI254023X (GI4023) ATP Geniposidic acid CORM-3 Oxymatrine

Cell Culture, Treatment & Working Concentration

Cell Lines	Assay Type	Concentration	Incubation Time	Activity Description	PMID
MDA-MB-231	Cytotoxicity assay	400 uM	96 hrs	Cytotoxicity against human MDA-MB-231 cells assessed as decrease in cell number at 400 uM after 96 hrs by trypan blue assay	23420624
MDA-MB-231	Cytotoxicity assay	200 to 400 uM	96 hrs	Cytotoxicity against human MDA-MB-231 cells assessed as decrease in cell viability at 200 to 400 uM after 96 hrs by trypan blue assay	23420624
HL60	Function assay	5 to 10 uM	15 mins	Inhibition of PAD4 in human HL60 cells assessed as reduction in A23187-induced citrullinated H4 level at 5 to 10 uM preincubated for 15 mins followed by A23187 addition measured after 15 mins by Western blot analysis	30344909
HEK293T	Function assay	10 nM to 100 uM		Inhibition of human recombinant PAD3 expressed in HEK293T cells assessed as conversion of BAEE to sodium benzoyl-L-citrulline at 10 nM to 100 uM by colorimetric analysis	ChEMBL
HEK293T	Function assay	100 uM	24 hrs	Inhibition of thapsigargin-induced cell death in human HEK293T cells overexpressing human recombinant PAD3 assessed as increase in cell survival at 100 uM treated 15 mins prior to thapsigargin addition measured after 24 hrs drug treatment by methylene blu	ChEMBL
Click to View More Cell Line Experimental Data

Chemical Information, Storage & Stability

Molecular Weight	424.8	Formula	C₁₄H₁₉ClN₄O₂.C₂HF₃O₂	Storage (From the date of receipt)	3 years-20°Cpowder
CAS No.	1043444-18-3	--		Storage of Stock Solutions	1 year-80°Cin solvent 1 month-20°Cin solvent
Synonyms	N/A			Smiles	C1=CC=C(C=C1)C(=O)NC(CCCN=C(CCl)N)C(=O)N.C(=O)(C(F)(F)F)O

Solubility

In vitro
Batch:

DMSO : 84 mg/mL (197.74 mM)
(Moisture-contaminated DMSO may reduce solubility. Use fresh, anhydrous DMSO.)

Ethanol : 84 mg/mL

Water : 35 mg/mL (Ultrasonic and heating.)

DMSO : 85 mg/mL (200.09 mM)
(Moisture-contaminated DMSO may reduce solubility. Use fresh, anhydrous DMSO.)

Water : 70 mg/mL (超声加热十分钟，60℃)

Ethanol : Insoluble

Molarity Calculator

Mass	Concentration	Volume	Molecular Weight
=	×	×

Dilution Calculator Molecular Weight Calculator

In vivo Batch:
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Clear solution	5%DMSO 1 40%PEG300 2 5%Tween80 3 50%ddH2O Validated by Selleck labs. Should you need adjustments to this formulation, contact our sales team for custom testing.	4.250mg/ml (10.00mM)	Taking the 1 mL working solution as an example, add 50 μL of 85 mg/ml clarified DMSO stock solution to 400 μL of PEG300, mix evenly to clarify it; add 50 μL of Tween80 to the above system, mix evenly to clarify; then continue to add 500 μL of ddH2O to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.

In vivo Formulation Calculator (Clear solution)

Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)

Dosage: mg/kg Average weight of animals: g Dosing volume per animal:μL Number of animals:

Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)

% DMSO + % + % Tween 80 + % ddH₂O

%DMSO+ %

Calculation results:

Working concentration: mg/ml;

Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )

Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH₂O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.

Mechanism of Action

Targets/IC₅₀/Ki	PAD1 (Cell-free assay) 0.8 μM PAD4 (Cell-free assay) 5.9 μM PAD3 (Cell-free assay) 6.2 μM
In vitro	Cl-amidine antagonizes the PAD4-mediated enhancement of the the p300GBD-GRIP1 interaction in a dose-dependent manner. The inhibitory effect of this compound is not a nonspecific one but is targeted at the active PAD4 enzyme. This compound increases p53 expression in CD45 positive immune cells. It triggers the differentiation and apoptosis of multiple cancer cell lines that are p53+/+ and p53 (e.g., HL60, HT29, TK6, and U2-OS cells). This chemical induces the expression of p53 and several downstream target genes including the cyclin dependent kinase inhibitor p21, GADD45, and the proapoptotic protein PUMA in U2-OS osteosarcoma cells.
In vivo	Cl-amidine treatment inhibits NZM(New Zealand mixed 2328) NET(neutrophil extracellular trap) formation in vivo and significantly alters circulating autoantibody profiles and complement levels while reducing glomerular IgG deposition. Further, this compound increases the differentiation capacity of bone marrow endothelial progenitor cells, improves endothelium-dependent vasorelaxation, and markedly delays time to arterial thrombosis induced by photochemical injury. It delays thrombosis development in NZM mice. This chemical inhibits PADs in mice without significant toxicity and improves disease phenotypes in animal models of inflammatory arthritis and inflammatory bowel disease. And it is shown to reduce disease severity in mouse models of ulcerative colitis and RA.
References	[1] https://pubmed.ncbi.nlm.nih.gov/17002273/ [2] https://pubmed.ncbi.nlm.nih.gov/20469888/ [3] https://pubmed.ncbi.nlm.nih.gov/23722903/ [4] https://pubmed.ncbi.nlm.nih.gov/21415415/

Applications

Methods	Biomarkers	Images	PMID
Western blot	PAD4 / H3R17Me / H3Cit / p21 / p53 / p-p53 iNOS MMP-2 / MMP-9 p-Elk-1 / Elk-1		18505818

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Handling Instructions

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Cl-amidine PAD inhibitor

Chemical Structure

Molecular Weight: 424.8