AT7519 844442-38-2

.cp_wz table {border-top: 1px solid #ccc; border-left: 1px solid #ccc; } .cp_wz table td {border-right: 1px solid #ccc; borda inferior: 1px sólido #ccc; preenchimento: 5px 0px 0px 5px;} .cp_wz tabela th {border-right: 1px solid #ccc; border-bottom: 1px solid #ccc; preenchimento: 5px 0px 0px 5px;} \ n Peso molecular: \ n 382,24 AT7519 é um inibidor multi-CDK para CDK1, 2, 4, 6 e 9 com IC50 de 10-210 nM. É menos potente para CDK3 e pouco ativo para CDK7. Fase 2. \ n Atividade biológica AT7519 é um inibidor de CDK competitivo de ATP com um valor Ki de 38 nM para CDK1. AT7519 é inativo contra todas as quinases não CDK com exceção de GSK3β (IC50 = 89 nM). AT7519 mostra atividade antiproliferativa potente em uma variedade de linhas de células tumorais humanas com valores de IC50 variando de 40 nM para MCF-7 a 940 nM para SW620 consistente com a inibição de CDK1 e CDK2. AT7519 induz citotoxicidade dependente da dose em linhas de células de mieloma múltiplo (MM) com valores de IC50 variando de 0,5 a 2 μM em 48 horas, com as linhas de células mais sensíveis sendo MM.1S (0,5 μM) e U266 (0,5 μM) e o mais resistente MM.1R (> 2 μM). Não induz citotoxicidade em células mononucleares do sangue periférico (PBMNC). AT7519 supera parcialmente a vantagem proliferativa conferida por IL6 e IGF-1, bem como o efeito protetor das células do estroma da medula óssea (BMSCs). AT7519 induz a rápida desfosforilação de CTD de RNA pol II nos locais da serina 2 e serina 5 e leva à inibição da transcrição, contribuindo parcialmente para a citotoxicidade induzida pelo AT7519 das células MM. AT7519 induz a ativação de GSK-3β por regulação negativa da fosforilação de GSK-3β, que também contribui para a apoptose induzida por AT7519 independente da inibição da transcrição. Uma dosagem duas vezes ao dia de AT7519 (9,1 mg / kg) causa a regressão do tumor de tumores sc em estágio inicial e avançado nos modelos de xenoenxerto de câncer de cólon HCT116 e HT29. O tratamento com AT7519 (15 mg / kg) inibe o crescimento do tumor e prolonga a sobrevida global mediana dos camundongos no modelo de camundongo com xenoenxerto MM humano em associação com o aumento da ativação da caspase 3. Ensaio de cinase de protocolo (apenas para referência): [1]

In vitro Kinase Assays

Kinase assays for CDK1, CDK2 and GSK3-β are all carried out in a radiometric filter binding format. Assays for CDK5 are in DELFIA format and for CDKs 4 and 6 in ELISA format. For CDKs 1 and 2, the relevant CDK and 0.12 μg/mL Histone H1 are incubated in 20 mM MOPS, pH 7.2, 25 mM β-glycerophosphate, 5 mM EDTA, 15 mM MgCl2, 1 mM sodium orthovanadate, 1 mM DTT, 0.1 mg/mL BSA, 45 μM ATP (0.78 Ci/mmol) and different concentrations of AT7519 for 2 or 4 hours respectively. For GSK3-β, the relevant enzyme and 5 μM glycogen synthase peptide 2 along with 10 mM MOPS pH 7.0, 0.1 mg/mL BSA, 0.001% Brij-35, 0.5% glycerol, 0.2 mM EDTA, 10 mM MgCl2, 0.01% β-mercaptoethanol, 15 μM ATP (2.31 Ci/mmol) and different concentrations of AT7519 are incubated for 3 hours. Assay reactions are stopped by adding an excess of orthophosphoric acid and filtered using Millipore MAPH filter plates. The plates are then washed, scintillant added and radioactivity measured by scintillation counting on a Packard TopCount. For CDK5, CDK5/p35 and 1μM of a biotinylated Histone H1 peptide (Biotin-PKTPKKAKKL) are incubated in 25 mM Tris-HCl, pH 7.5, 2.5 mM MgCl2, 0.025% Brij-35, 0.1 mg/mL BSA, 1 mM DTT, 15 μM ATP and different concentrations of AT7519 for 30 minutes. Assay reactions are stopped using EDTA, transferred to Neutravidin-coated plates and phosphorylated peptide quantified by means of a rabbit phospho-cdk1 substrate polyclonal antibody and DELFIA europium-labelled anti-rabbit IgG secondary antibody using time-resolved fluorescence at λex=335nm, λem=620nm. For CDK 4 and 6 assays, plates are coated with GST- pRb769-921 and blocked with Superblock. CDK4 or 6 is incubated with 15 mM MgCl2, 50 mM HEPES, pH 7.4, 1 mM DTT, 1 mM EGTA, pH 8.0, 0.02% Triton X-100, 2.5% DMSO and different concentrations of AT7519; the reaction is initiated by addition of ATP. After 30 minutes, reactions are stopped by the addition of 0.5 M EDTA pH 8.0. Plates are then washed and incubated for one hour with the primary antibody (anti- p-Rb Serine 780) diluted in Superblock followed by secondary antibody (alkaline phosphatase linked anti-rabbit) for a further hour. Plates are developed using the Attophos system and fluorescence read on a Spectramax Gemini plate reader at excitation 450 nm and emission 580 nm. In all cases, IC50 values are calculated from replicate curves, using GraphPad Prism software.

Ensaio de células: [2]

Cell lines	MM.1S, MM.1R, RPMI8226, U266, RPMI8266, RPMI-Dox40, OPM1 cells, primary MM cells and PBMNCs
Concentrations	Dissolved in DMSO at a concentration of 10 mM, final concentrations 0.25-4 μM
Incubation Time	24 or 48 hours
Method	Cells are incubated with different concentrations of AT7519 for 24 or 48 hours at 37 °C. Cell viability is assessed by measuring 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrasodium bromide (MTT) dye absorbance. DNA synthesis is measured by tritiated thymidine uptake (3H-TdR). Apoptosis is assessed by using Annexin V/PI staining. The percentage of cells undergoing apoptosis is defined as the sum of early apoptosis (Annexin V-positive cells) and late apoptosis (Annexin V-positive and PI-positive cells

Estudo Animal: [2]

Animal Models	Male SCID mice inoculated subcutaneously with MM.1S cells
Formulation	Dissolved in 0.9% saline
Dosages	15 mg/kg/day
Administration	Dosed i.p.
Solubility	30% propylene glycol, 5% Tween 80, 65% D5W, 30 mg/mL
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.

Conversão de diferentes modelos de animais com base em BSA (valor com base em dados das diretrizes preliminares da FDA)

Species	Baboon	Dog	Monkey	Rabbit	Guinea pig	Rat	Hamster	Mouse
Weight (kg)	12	10	3	1.8	0.4	0.15	0.08	0.02
Body Surface Area (m2)	0.6	0.5	0.24	0.15	0.05	0.025	0.02	0.007
Km factor	20	20	12	12	8	6	5	3

Animal A (mg/kg) = Animal B (mg/kg) multiplied by	Animal B Km
	Animal A Km

Por exemplo, para modificar a dose de resveratrol usada para um camundongo (22,4 mg / kg) para uma dose baseada na BSA para um rato, multiplique 22,4 mg / kg pelo fator Km para um camundongo e, em seguida, divida pelo fator Km para um rato. Este cálculo resulta em uma dose equivalente de rato para o resveratrol de 11,2 mg / kg.

Rat dose (mg/kg) = mouse dose (22.4 mg/kg) ×	mouse Km(3)	= 11.2 mg/kg
	rat Km(6)

Informação Química

Molecular Weight (MW)	382.24
Formula	C16H17Cl2N5O2
CAS No.	844442-38-2

Storage	3 years -20℃Powder
	6 months-80℃in solvent (DMSO, water, etc.)
Synonyms

Solubility (25°C) *	In vitro	DMSO	10 mg/mL (26.16 mM)
		Water	<1 mg/mL (
		Ethanol	<1 mg/mL (
	In vivo	30% propylene glycol, 5% Tween 80, 65% D5W	30 mg/mL
* <1 mg/ml means slightly soluble or insoluble. * Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.

Chemical Name	4-(2,6-dichlorobenzamido)-N-(piperidin-4-yl)-1H-pyrazole-3-carboxamide

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Grupo de Produto : Ciclo de célula > Inibidor CDK