CX-6258 HCl(known also as CX-6258 hydrochloride) is a novel, highly potent and orally bioactive pan-Pim kinase inhibitor with potential anticancer activity. It inhibits Pim1, Pim2, and Pim3 with IC50s of 5 nM, 25 nM and 16 nM, respectively. At CX-6258 showed robust antiproliferative potencies against all cell lines tested derived from human solid tumors and hematological malignancies. In mechanistic cellular assays with MV-4-11 human AML cells, caused dose-dependent inhibition of the phosphorylation of 2 pro-survival proteins, Bad and 4E-BP1, at the Pim kinase specific sites S112 and S65 and T37/46, respectively.

Physicochemical Properties

Molecular Formula	C26H24CLN3O3.HCL
Molecular Weight	498.40
Exact Mass	497.127
CAS #	1353859-00-3
Related CAS #	1202916-90-2;1353858-99-7 (hydrochloride hydrate)
PubChem CID	72201040
Appearance	Orange to red solid powder
LogP	5.686
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	4
Rotatable Bond Count	3
Heavy Atom Count	34
Complexity	774
Defined Atom Stereocenter Count	0
SMILES	CN1CCCN(CC1)C(=O)C2=CC=CC(=C2)C3=CC=C(O3)/C=C/4\C5=C(C=CC(=C5)Cl)NC4=O.Cl
InChi Key	YYIMMVXTWBIEAG-YHLMHSEJSA-N
InChi Code	InChI=1S/C26H24ClN3O3.ClH/c1-29-10-3-11-30(13-12-29)26(32)18-5-2-4-17(14-18)24-9-7-20(33-24)16-22-21-15-19(27)6-8-23(21)28-25(22)31;/h2,4-9,14-16H,3,10-13H2,1H3,(H,28,31);1H/b22-16+
Chemical Name	(3E)-5-Chloro-3-[[5-[3-[(hexahydro-4-methyl-1H-1,4-diazepin-1-yl)carbonyl]phenyl]-2-furanyl]methylene]-1,3-dihydro-2H-indol-2-one hydrochloride
Synonyms	CX 6258 HCl; CX6258 HCl; CX-6258 HCl; CX 6258 Hydrochloride; CX6258 Hydrochloride; CX-6258 Hydrochloride;
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	Two pro-survival proteins, Bad and 4E-BP1, have their phosphorylation inhibited in a dose-dependent manner by CX-6258 hydrochloride at Pim kinase specific sites S112, S65, and T37/46, respectively [1]. In PC3 cells, treatment with 12 mM CX-6258 hydrochloride for three hours decreased the steady-state amounts of ectopic NKX3.1 [2]. The half-life of NKX3.1 was significantly reduced by CX-6258 hydrochloride treatment [2].
ln Vivo	In two Pim kinase-driven tumor models, CX-6258 hydrochloride (50–100 mg/kg; oral; once daily; 21 days) showed significant in vivo effectiveness [1].
Cell Assay	Western Blot Analysis[1] Cell Types: MV-4-11 human AML cells. Tested Concentrations: 0.1 μM, 1 μM, 10 μM. Incubation Duration: 2 hrs (hours). Experimental Results: Caused dose dependent inhibition of the phosphorylation of two pro-survival proteins, Bad and 4E-BP1, at the Pim kinase specific sites S112 and S65 and T37/46, respectively.
Animal Protocol	Animal/Disease Models: Nude mice, MV-4-11 xenograft models[1] Doses: 50 mg/kg, 100 mg/kg. Route of Administration: Oral administration; one time/day; over a period of 21 days. Experimental Results: demonstrated dose dependent efficacy, with a 50 mg/kg dose producing 45% tumor growth inhibition (TGI) and a 100 mg/kg dose producing 75% TGI.
References	[1]. Mustapha Haddach, Jerome Michaux, Michael K, Discovery of CX-6258. A Potent, Selective, and Orally Efficacious pan-Pim Kinases Inhibitor. ACS Med. Chem. Lett., 2012, 3 (2), pp 135-139. [2]. Padmanabhan A, Gosc EB, Bieberich CJ. Stabilization of the prostate-specific tumor suppressor NKX3.1 by the oncogenic protein kinase Pim-1 in prostate cancer cells. J Cell Biochem. 2013 May;114(5):1050-7.

Solubility Data

Solubility (In Vitro)

DMSO: 89 mg/mL (178.6 mM) Water:<1 mg/mL Ethanol:<1 mg/mL

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.02 mM) (saturation unknown) in 10% DMSO + 40% PEG300 +5% Tween-80 + 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.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.0064 mL	10.0321 mL	20.0642 mL
	5 mM	0.4013 mL	2.0064 mL	4.0128 mL
	10 mM	0.2006 mL	1.0032 mL	2.0064 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.