Gusacitinib (formerly known as EN-3351; ASN-002) is a novle and potent dual inhibitor of SYK/JAK (spleen tyrosine kinase/janus kinase) with antitumor activity in both hematological and solid tumor xenograft models. In pLAT and pSTAT assays, ASN002 significantly inhibited the JAK family kinase and SYK signaling pathways, with IC50 values ranging from 5 to 46 nM.

Physicochemical Properties

Molecular Formula	C24H28N8O2
Molecular Weight	460.531523704529
Exact Mass	460.233
Elemental Analysis	C, 62.59; H, 6.13; N, 24.33; O, 6.95
CAS #	1425381-60-7
Related CAS #	Gusacitinib hydrochloride;2228989-14-6
PubChem CID	71269142
Appearance	Light yellow to yellow solid powder
LogP	2.6
Hydrogen Bond Donor Count	3
Hydrogen Bond Acceptor Count	9
Rotatable Bond Count	5
Heavy Atom Count	34
Complexity	775
Defined Atom Stereocenter Count	0
InChi Key	NLFLXLJXEIUQDL-UHFFFAOYSA-N
InChi Code	InChI=1S/C24H28N8O2/c25-10-5-16-6-11-32(12-7-16)24-28-20-15-26-30-23(34)21(20)22(29-24)27-17-1-3-18(4-2-17)31-13-8-19(33)9-14-31/h1-4,15-16,19,33H,5-9,11-14H2,(H,30,34)(H,27,28,29)
Chemical Name	2-[1-[4-[4-(4-hydroxypiperidin-1-yl)anilino]-5-oxo-6H-pyrimido[4,5-d]pyridazin-2-yl]piperidin-4-yl]acetonitrile
Synonyms	ASN002; ASN 002; ASN-002; Gusacitinib; EN3351; EN-3351; EN 3351
HS Tariff Code	2934.99.9001
Storage	Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

Targets	SYK, JAK (IC50 = 5-46 nM)
ln Vitro	ASN-002 significantly inhibits the SYK and JAK family kinase signaling pathways, as indicated by pSTAT and pLAT levels, in mechanistic cell-based studies with IgE and cytokine stimulations. In a wide range of human cancer cell lines, such as DHL6, DHL4, OCI-LY10, H929, Pfeiffer, HT-1376, and Lovo, ASN-002 exhibits anti-proliferative activity, indicating activity in both solid and hematological tumor types[1].
ln Vivo	ASN-002 shows notable efficacy in preventing tumor growth (>95%) in a multiple myeloma (H929) xenograft model. In the human erythroleukemia (HEL) mouse model, it also markedly postpones the onset of hind limb paralysis. ASN-002 exhibits little to no inhibition of CYP450 isozymes, good oral bioavailability, metabolic stability, and is not a Pgp substrate. In toxicology studies on rats and dogs, ASN-002 exhibits a good safety profile[1].
References	[1]. Small-Molecule Kinase Inhibitors for the Treatment of Nononcologic Diseases. J Med Chem. 2021 Feb 11;64(3):1283-1345. [2]. Janus Kinase Inhibitors in Dermatology: Part 1 - General Considerations and Applications in Vitiligo and Alopecia Areata. Actas Dermo-Sifiliográficas, 2021, 112(6): 503-515. [3]. Abstract 792: ASN002: A novel dual SYK/JAK inhibitor with strong antitumor activity. AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA.
Additional Infomation	Gusacitinib is under investigation in clinical trial NCT02550678 (A Study of the Efficacy and Safety of ASN-002 in Adult Patients With Low-risk Nodular Basal Cell Carcinoma).

Solubility Data

Solubility (In Vitro)

DMSO: 92~100 mg/mL (199.8~217.1 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 1.67 mg/mL (3.63 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 16.7 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 1.67 mg/mL (3.63 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 16.7 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.1714 mL	10.8571 mL	21.7141 mL
	5 mM	0.4343 mL	2.1714 mL	4.3428 mL
	10 mM	0.2171 mL	1.0857 mL	2.1714 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.