KW-2449

製品コード：S2158 純度：99.77%

KW-2449 is a multiple-targeted inhibitor, mostly for Flt3 with IC50 of 6.6 nM, modestly potent to FGFR1, Bcr-Abl and Aurora A; little effect on PDGFRβ, IGF-1R, EGFR. Phase 1.

CAS No. 1000669-72-6

サイズ	価格(税別)	在庫状況
10mg	JPY 16900	国内在庫あり
50mg	JPY 63400	国内在庫あり

代表番号: 045-509-1970\|電子メール：[email protected] よく尋ねられる質問

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製品安全説明書

現在のバッチを見る：純度： 99.77%

99.77

KW-2449関連製品

関連ターゲット	FLT3-WT FLT3-ITD FLT3-D835Y	もっと見る
関連製品	Dovitinib (TKI-258) Dovitinib (TKI258) Lactate monohydrate Tandutinib (MLN518) ENMD-2076 UNC2025 AST-487 (NVP-AST487) TCS 359 G-749 SKLB4771 (FLT3-IN-1) AMG 925	もっと見る
関連化合物ライブラリー	Tyrosine Kinase Inhibitor Library PI3K/Akt Inhibitor Library Angiogenesis Related compound Library HIF-1 Signaling Pathway Compound Library FDA-approved Anticancer Drug Library	もっと見る

シグナル伝達経路

FLT3シグナル伝達経路

FLT3阻害剤の選択性比較

生物活性

製品説明

KW-2449 is a multiple-targeted inhibitor, mostly for Flt3 with IC50 of 6.6 nM, modestly potent to FGFR1, Bcr-Abl and Aurora A; little effect on PDGFRβ, IGF-1R, EGFR. Phase 1.

特性

Investigated as a FLT3 inhibitor in clinical trials, with others in early development.

Targets

FLT3 (D835Y) ^[1] (Cell-free assay)	Abl (T315I) ^[1] (Cell-free assay)	FLT3 ^[1] (Cell-free assay)	Abl ^[1] (Cell-free assay)	FGFR1 ^[1] (Cell-free assay)	もっとクリックする
1 nM	4 nM	6.6 nM	14 nM	36 nM	もっとクリックする

In Vitro
In vitro	KW-2449, a multikinase inhibitor of FLT3, ABL, ABL-T315I, and Aurora kinase, is under investigation to treat leukemia patients. KW-2449 shows the potent growth inhibitory effects on leukemia cells with FLT3 mutations by inhibition of the FLT3 kinase, resulting in the down-regulation of phosphorylated-FLT3/STAT5, G1 arrest, and apoptosis. Oral administration of KW-2449 shows dose-dependent and significant tumor growth inhibition in FLT3-mutated xenograft model with minimum bone marrow suppression. In FLT3 wild-type human leukemia, KW-2449 induces the reduction of phosphorylated histone H3, G2/M arrest, and apoptosis. In Imatinib-resistant leukemia, KW-2449 contributes to release of the resistance by the simultaneous down-regulation of BCR/ABL and Aurora kinases. The inhibitory activity of KW-2449 is not affected by the presence of human plasma protein, such as α1-acid glycoprotein. KW-2449 has potent growth inhibitory activity against various types of leukemia by several mechanisms of action. KW-2449 has significant activity and warrants clinical study in leukemia patients with FLT3 mutations as well as Imatinib-resistant mutations. Phosphorylation levels of FLT3 and STAT5 are decreased by KW-2449 in a dose-dependent manner. In addition, it potently inhibits ABL-T315I, which is associated with IM resistance, with IC50 of 4 nM. On the other hand, KW-2449 has little effect on PDGFRβ, IGF-1R, EGFR, and various serine/threonine kinases even at a concentration of 1 μM. KW-2449 has the potent growth inhibitory activities against not only FLT3/ITD-expressing leukemia cells but also FLT3/KDM-activated and wild-type FLT3-overexpressing leukemia cells. In accordance with growth inhibitory effect, KW-2449 suppresses the phosphorylations of FLT3 (P-FLT3) and its downstream molecule phospho-STAT5 (P-STAT5) in MOLM-13 cells in a dose-dependent manner. Furthermore, KW-2449 increases the percentage of cells in the G1 phase of the cell cycle and reciprocally reduces the percentage of cells in the S phase, resulting in the increase of apoptotic cell population. KW-2449 can dephosphorylate constitutively active WT-FLT3 kinase but not inhibit the proliferation of leukemia cells if they are not mainly addicted to FLT3 the kinase. ^[1] KW-2449 is rapidly absorbed and converted to a major metabolite M1.Preclinical studies reveal that KW-2449 is converted by monoamine oxidase-B (MAO-B) and aldehyde oxidase into its major metabolite M1.KW-2449 mediates cytotoxicity thru inhibition of FLT3/ITD.KW-2449 is a direct inhibitor of FLT3 and induces inhibition of its downstream target STAT5. ^[2] KW2449 interacts synergistically with HDACIs to induce apoptosis in Ph+ CML cells in a time- and concentration-dependent manner. KW2449 synergistically enhances the lethality of vorinostat/SNDX275 in CML cells.KW-2449 regimens are active against additional IM-resistant Bcr/Abl⁺ leukemia cells. KW2449 moderately reduces phosphorylation of histone H3, an indicator of Aurora B activity, in nocodozole-treated K562 cells. ^[3]
Kinase Assay	FLT3 phosphorylation
Kinase Assay	Leukemia cells are washed in phosphate-buffered saline (PBS), then lysed by resuspending the cells in lysis buffer (20 mM Tris pH 7.4, 100 mM NaCl, 1% Igepal, 1 mM EDTA, 2 mM NaVO₄, plus Complete protease inhibitor KW-2449 for 30 minutes while rocking. The extract is clarified by centrifugation at 1.6 × 10⁴ g and the supernatant is assayed for protein (Bio-Rad). A 50-μg aliquot is removed as a whole-cell lysate for analysis of STAT5, and the remainder is used for immunoprecipitation with anti-FLT3. Anti-FLT3 antibody is added to the extract for overnight incubation, then protein A sepharose is added for 2 additional hours. Separate sodium dodecyl sulfate–polyacrylamide electrophoresis (SDS-PAGE) gels for whole-cell lysate and immunoprecipates are run in parallel. After transfer to Immobilon membranes, immunoblotting is performed with antiphosphotyrosine antibody (4G10) to detect phosphorylated FLT3 or, for the whole-cell lysate gels, with a rat monoclonal antibody against phosphorylated STAT5 (residue Y694) then stripped and reprobed with anti-FLT3 antibody to measure total FLT3. Proteins are visualized using chemiluminescence, exposed on Kodak BioMax XAR film, developed, and scanned using a Bio-Rad GS800 densitometer. The concentration of KW-2449 for which the phosphorylation of FLT3 or STAT5 is inhibited to 50% of its baseline (IC50) is determined using linear regression analysis of the dose response curves. For direct analysis of FLT3 and STAT5 in circulating blasts, peripheral blood is collected in heparinized tubes and promptly chilled on ice. Samples are centrifuged for 10 minutes at 900 g, at 4 °C. The plasma is removed and stored frozen at −80 °C. The buffy coat is carefully transferred to ice-cold PBS, layered onto chilled Ficoll-Hypaque, and centrifuged for 5 minutes at 600 g, at 4 °C. All subsequent steps are carried out at 4 °C. Mononuclear cells are collected and washed rapidly once in red blood cell lysis buffer (0.155 M NH₄Cl, 0.01 M KHCO₃, 0.1 mM EDTA), then washed once in PBS. Cells are then lysed as described for FLT3 and STAT5 analysis.
細胞実験	細胞株	MOLM-13 and RS4;11 cells
	濃度	33nM, 75nM, 0.1μM, 0.3μM and 0.15μM
	反応時間	24, 48, and 72 hours
	実験の流れ	Cell viability is determined by the sodium 3′-[1-(phenylaminocarbonyl)-3, 4-tetrazolium]-bis (4-methoxy-6-nitro) benzene sulfonic acid hydrate assay after incubation with or without KW-2449 for 72 hours at 37 °C. The number of viable cells is determined using the Cell Proliferation Kit II. For cell-cycle analysis, MOLM-13 and RS4;11 cells are treated with KW-2449. After 24, 48, and 72 hours of incubation at 37 °C, DNA contents are analyzed. Cell cycle distribution of K562, TCC-Y, and TCC/Ysr is analyzed 24 hours after treatment with KW-2449 or imatinib.

In Vivo
In Vivo	In the MOLM-13 tumor xenograft model, oral administration of KW-2449 for 14 days shows a potent and significant antitumor effect in a dose-dependent manner. ^[1]
動物実験	動物モデル	CBySmn.CB17-Prkds^scid/J (BALB/C) mice are injected with BV173/E255K/Luc cl4 cells.
	投与量	32 mg/kg/day, 5 days/week
	投与経路	Orally (p.o.) administered

NCT Number	Recruitment	Conditions	Sponsor/Collaborators	Start Date	Phases
臨床試験 (data from https://clinicaltrials.gov, updated on 2024-05-22)
NCT00346632	Terminated	Acute Myelogenous Leukemia\|Acute Lymphoblastic Leukemia\|Myelodysplastic Syndromes\|Chronic Myelogenous Leukemia	Kyowa Kirin Inc.	June 2006	Phase 1

参考
[1] Shiotsu Y, et al. Blood, 2009, 114(8), 1607-17. [2] Pratz KW, et al. Blood, 2009, 113(17), 3938-46. [3] Nguyen T, et al. Clin Cancer Res, 2011, 17(10), 3219-32.

参考

化学情報

分子量	332.4	化学式	C₂₀H₂₀N₄O
CAS No.	1000669-72-6	SDF	Download KW-2449 SDFをダウンロードする
Smiles	C1CN(CCN1)C(=O)C2=CC=C(C=C2)C=CC3=NNC4=CC=CC=C43
保管	3年-20°C粉	溶液の保管冷凍と解凍の繰り返しを避けるため小分けにしてください	1年-80°Cin solvent
保管	3年-20°C粉	溶液の保管冷凍と解凍の繰り返しを避けるため小分けにしてください	１ケ月-20°Cin solvent

In vitro
Batch:

DMSO : 67 mg/mL ( (201.56 mM)；吸湿したDMSOは溶解度を減少させます。新しいDMSOをご使用ください。)

Ethanol : 67 mg/mL

Water : Insoluble

モル濃度計算器

in vivo
Batch:

Add solvents to the product individually and in order.

投与溶液組成計算機

実験計算

モル濃度計算器

質量	濃度	体積	分子量
=	×	×

希釈計算器分子量計算器

投与溶液組成計算機(クリア溶液)

ステップ１：実験データを入力してください。（実験操作によるロスを考慮し、動物数を1匹分多くして計算・調製することを推奨します）

投与量 mg/kg 動物平均体重 g 投与体積（動物毎）μL 動物数匹

ステップ２：投与溶媒の組成を入力してください。（ロット毎に適した溶解組成が異なる場合があります。詳細については弊社までお問い合わせください）

% DMSO + % + % Tween 80 + % ddH₂O

%DMSO+ %

計算結果：

投与溶媒濃度： mg/ml;

DMSOストック溶液調製方法： mg 試薬を μL DMSOに溶解する（濃度 mg/mL, 注：濃度が当該ロットのDMSO溶解度を超える場合はご連絡ください。 )

投与溶媒調製方法：Take μL DMSOストック溶液に μL PEG300，を加え、完全溶解後μL Tween 80，を加えて完全溶解させた後 μL ddH₂O，を加え完全に溶解させます。

投与溶媒調製方法：μL DMSOストック溶液に μL Corn oil，を加え、完全溶解。

注意：１.ストック溶液に沈殿、混濁などがないことをご確認ください；
２.順番通りに溶剤を加えてください。次のステップに進む前に溶液に沈殿、混濁などがないことを確認してから加えてください。ボルテックス、ソニケーション、水浴加熱など物理的な方法で溶解を早めることは可能です。

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