GNF-5 (GNF 5; GNF5), a GNF-2 analog with better pharmacokinetic profiles, is a potent, selective and allosteric/non-ATP competitive Bcr-Abl inhibitor with potential anticancer activity. It inhibits Bcr-Abl (wild-type Abl) with an IC50 of 220 nM. It exhibits excellent in vivo anticancer efficacy in Ba/F3.p210 xenograft mouse model.

Physicochemical Properties

Molecular Formula	C20H17F3N4O3
Molecular Weight	418.37
Exact Mass	418.125
CAS #	778277-15-9
Related CAS #	778277-15-9
PubChem CID	44129660
Appearance	White to off-white solid powder
Density	1.4±0.1 g/cm3
Boiling Point	612.4±55.0 °C at 760 mmHg
Flash Point	324.2±31.5 °C
Vapour Pressure	0.0±1.9 mmHg at 25°C
Index of Refraction	1.600
LogP	3.31
Hydrogen Bond Donor Count	3
Hydrogen Bond Acceptor Count	9
Rotatable Bond Count	7
Heavy Atom Count	30
Complexity	544
Defined Atom Stereocenter Count	0
InChi Key	IIQUYGWWHIHOCF-UHFFFAOYSA-N
InChi Code	InChI=1S/C20H17F3N4O3/c21-20(22,23)30-16-6-4-15(5-7-16)27-18-11-17(25-12-26-18)13-2-1-3-14(10-13)19(29)24-8-9-28/h1-7,10-12,28H,8-9H2,(H,24,29)(H,25,26,27)
Chemical Name	N-(2-Hydroxyethyl)-3-[6-[[4-(trifluoromethoxy)phenyl]amino]-4-pyrimidinyl]benzamide
Synonyms	GNF 5; GNF5; GNF-5;
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

ln Vitro	GNF-5 inhibits wild-type Abl with an IC50 of 0.22 µM, but not myristate site mutant E505K (IC50 ＞10 µM) [1].
ln Vivo	GNF-5 has appropriate pharmacokinetic properties (5 mg/kg IV or 20 mg/kg oral)[1]. GNF-5 is effective in vivo when taken orally, twice daily for seven days at 50 or 100 mg/kg, however relapses are possible[1]. In vivo, nilotinib with T315I Bcr-Abl combo is inhibited by GNF-5 (75 mg/kg, bid)[1].
Cell Assay	Cell Viability Assay[1] Cell Types: wild type and mutant Bcr- Abl expressing Ba/F3 cells Tested Concentrations: 0.2, 0.8 and 1.6 μM Incubation Duration: 48 h Experimental Results: Inhibited wild-type Abl in a non-ATP competitive fashion.
Animal Protocol	Animal/Disease Models: Male balb/c (Bagg ALBino) mouse[1] Doses: 5 mg/kg, 20 mg/kg Route of Administration: 5 mg/kg intravenously (iv)or 20 mg/kg orally Experimental Results: AUC_inf (min*ug/mL) 292.37 AUC_inf (hrs*nM 11647 Cmax(nM) 4386.08 Tmax(hrs) 0.50 Clast (nM) 636.16 T1 /2(hrs) 2.30 Vss(L/kg) 9.18 F (%) 44.82 Animal/Disease Models: p210 xenograft model[1] Doses: 50 or 100 mg/kg Route of Administration: po (oral gavage) twice (two times) daily, for 7 days Experimental Results: Could normalize blood counts and spleen size. Animal/Disease Models: Bone marrow transduction/transplantation model[1] Doses: 75 mg/kg Route of Administration: twice (two times) daily Experimental Results: demonstrated no significant response (alone). demonstrated no toxicity and had a strong and sustained inhibition of Bcr- Abl-mediated signaling combination with nilotinib.
References	[1]. Targeting Bcr-Abl by combining allosteric with ATP-binding-site inhibitors. Nature. 2010 Jan 28;463(7280):501-6. [2]. Targeting invadopodia-mediated breast cancer metastasis by using ABL kinase inhibitors. Oncotarget. 2018 Apr 24;9(31):22158-22183.

Solubility Data

Solubility (In Vitro)

DMSO: 83 mg/mL (198.4 mM) Water:<1 mg/mL Ethanol: 20 mg/mL warmed (47.8 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.98 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 25.0 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: ≥ 2.5 mg/mL (5.98 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 25.0 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. Solubility in Formulation 3: ≥ 2.5 mg/mL (5.98 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.3902 mL	11.9511 mL	23.9023 mL
	5 mM	0.4780 mL	2.3902 mL	4.7805 mL
	10 mM	0.2390 mL	1.1951 mL	2.3902 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.