KHK-IN-1 HCl (compound 8) is a selective and cell membrane-permeable/penetrable inhibitor of hexulokinase (KHK) (IC50=12 nM; F=34%). KHK-IN-1 HCl can inhibit the production of F1P in HepG2 cell lysate (IC50=400 nM). KHK-IN-1 HCl may be used for studying diabetes and obesity.

Physicochemical Properties

Molecular Formula	C21H27CLN8S
Molecular Weight	459.013
Exact Mass	458.176
CAS #	1303470-48-5
Related CAS #	KHK-IN-1;1303469-70-6
PubChem CID	92044374
Appearance	Light yellow to yellow solid powder
Hydrogen Bond Donor Count	4
Hydrogen Bond Acceptor Count	9
Rotatable Bond Count	7
Heavy Atom Count	31
Complexity	543
Defined Atom Stereocenter Count	0
InChi Key	VKIBPWKARAEIHN-UHFFFAOYSA-N
InChi Code	InChI=1S/C21H26N8S.ClH/c1-30-16-5-3-2-4-15(16)26-20-17-18(19(25-13-24-17)23-12-14-6-7-14)27-21(28-20)29-10-8-22-9-11-29;/h2-5,13-14,22H,6-12H2,1H3,(H,23,24,25)(H,26,27,28);1H
Chemical Name	8-N-(cyclopropylmethyl)-4-N-(2-methylsulfanylphenyl)-2-piperazin-1-ylpyrimido[5,4-d]pyrimidine-4,8-diamine;hydrochloride
Synonyms	INVIVO 0485; INVIVO0485; INVIVO-0485
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 Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.
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	KHK-IN-1 hydrochloride does not significantly inhibit cytochrome P450 (1A2, 2C19, 2D6, 2C9, and 3A4) and is stable in human and rat liver microsomal preparations (88% and 72% residual at 10 minutes)[1]. HepG2 cell lysate generation of F1P is inhibited by KHK-IN-1 hydrochloride (0-10 µM; incubate for 30 minutes, then add 15 mM fructose and incubate for an additional 3 hours) with an IC50 value of 400 nM [1].
ln Vivo	In rats, KHK-IN-1 hydrochloride (10 mg/kg; oral; single dosage) has an oral bioavailability of 34% [1].
Cell Assay	Cell viability assay [1] Cell Types: HepG2 Cell Tested Concentrations: 0-10 µM Incubation Duration: Incubate for 30 minutes, then add 15 mM fructose, and incubate for another 3 hrs (hours). Experimental Results: F1P production in HepG2 cell lysate was inhibited (IC50= 400 nm ).
Animal Protocol	Animal/Disease Models: Male SD (SD (Sprague-Dawley)) rat (approximately 250 g) [1]. Doses: 10 mg/kg Route of Administration: po (oral gavage); single Experimental Results:demonstrated reasonable oral bioavailability in rats (F=34%; oral t1/2=4 h), but had a high volume of distribution ( Vdss=32 L/kg) and high clearance (CL=160 mL/min/kg).
References	[1]. Inhibitors of Ketohexokinase: Discovery of Pyrimidinopyrimidines with Specific Substitution that Complements the ATP-Binding Site. ACS Med Chem Lett. 2011 Apr 18;2(7):538-43.

Solubility Data

Solubility (In Vitro)

DMSO : ~25 mg/mL (~54.47 mM) H2O : ~14.29 mg/mL (~31.13 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.08 mg/mL (4.53 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 20.8 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: ≥ 2.08 mg/mL (4.53 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 20.8 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	2.1786 mL	10.8930 mL	21.7860 mL
	5 mM	0.4357 mL	2.1786 mL	4.3572 mL
	10 mM	0.2179 mL	1.0893 mL	2.1786 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.