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AMD 3465 hexahydrobromide
本产品不向个人销售,仅用作科学研究,不用于任何人体实验及非科研性质的动物实验。
AMD 3465 hexahydrobromide图片
CAS NO:185991-07-5
规格:≥98%
包装与价格:
包装价格(元)
5mg电议
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25mg电议
50mg电议
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产品介绍
理化性质和储存条件
Molecular Weight (MW) 896.07
Formula C24H38N6.6HBr
CAS No. 185991-07-5 (6HBr)
Storage-20℃ for 3 years in powder form
-80℃ for 2 years in solvent
Solubility (In vitro)DMSO: 2 mg/mL (2.23 mM)
Water: 98 mg/mL (109.36 mM)
Ethanol: Insoluble
SMILES C1(CNCC2=CC=C(CN3CCNCCCNCCNCCC3)C=C2)=NC=CC=C1.[6HBr]
Synonyms AMD3465 hexahydrobromide; AMD-3465 6HBr; AMD 3465; GENZ-644494 hexahydrobromide; GENZ644494 hexahydrobromide; GENZ 644494 hexahydrobromide
实验参考方法
In Vitro

In vitro activity:The affinity of AMD3465 is 8-fold higher compared with AMD3100, in competition against the radiolabeled monoclonal antibody raised against CXCR4, 12G5. The affinity of AMD3465 is decreased>5000-fold in (D262N)-CXCR4 and 1913-fold in (A175F)-CXCR4. AMD3465 appears to interact with HisVII:-02 at the extracellular end of TM-VII (at the interface to extracellular loop 3) and HisIII:05. Both of these His residues are facing right into the main binding pocket of CXCR4. AMD3465 inhibits 125I-SDF-1α ligand binding to CCRF-CEM cells. AMD3465 inhibits CXCR4 activation as measured by GTP binding with an IC50 of 10.38±1.99 nM, and inhibits SDF-1α mediated calcium flux with an IC50 of 12.07±2.42 nM.


Kinase Assay: For the competition binding studies against CXCR4, a concentration range of AMD3465 is incubated for 3 h at 4°C in binding buffer (PBS containing 5 mM MgCl2, 1 mM CaCl2, 0.25% BSA pH 7.4) with 5×105 CCRF-CEM cells and 100 pM 125I-SDF-1α in Millipore DuraporeTM filter plates. Unbound 125I-SDF-1α is removed by washing with cold 50 mM HEPES, 0.5 M NaCl pH 7.4. The competition binding assay against BLT1 is performed on membranes from CHO-S cells expressing recombinant BLT1. The membranes is prepisd by mechanical cell lysis followed by high speed centrifugation, resuspended in 50 mM HEPES, 5 mM MgCl2 buffer and flash frozen. The membrane preparation is incubated with AMD3465 for 1 h at room temperature in an assay mixture containing 50 mM Tris, pH 7.4, 10 mM MgCl2, 10 mM CaCl2, 4 nM LTB4 mixed with 1 nM 3H-LTB4 and 8 μg membrane. The unbound 3H-LTB4 is separated by filtration on Millipore Type GF-C filter plates. The bound radioactivity is counted using a LKB Rackbeta 1209 Liquid Scintillation Counter.


Cell Assay: On the day prior to the experiment, the U87.CD4.CXCR4 transfectants is seeded in 0.1% gelatin-coated 96-well black wall microplates (Costar, Cambridge, MA) at 2×104 cells per well. On the day of the experiment, the cells is loaded with the fluorescent calcium indicator Fluo-3 acetoxymethyl at 4 μM for 45 min at 37°C. After thorough washing with calcium flux assay buffer (Hanks' balanced salt solution with 20 mM Hepes buffer and 0.2% bovine serum albumin, pH 7.4), the cells is preincubated for 15 min at 37°C with AMD3100 or AMD3465 (1 μg/mL) in the same buffer. Then, the intracellular calcium mobilization in response to 2-50 ng/mL CXCL12 is measured at 37°C by monitoring the fluorescence as a function of time simultaneously in all the wells using a Fluorometric Imaging Plate Reader.

In Vivo AMD3465 (5 mg/kg, s.c.) significantly elevates total white blood cells in DBA/2, C57Bl/6 and BALB/c mice between 0.5 and 2 h. AMD3465 significantly increases the specific cell populations in all three strains of mice included neutrophils, lymphocytes, and monocytes.
Animal model Male Swiss Webster mice
Formulation & Dosage PBS; 5, 10, 20, 50 and 100 mg/kg; Subcutaneous injection.
References
J Biol Chem. 2007 Sep 14;282(37):27354-65;Biochem Pharmacol. 2009 Oct 15;78(8):993-1000.