b-AP15(Synonyms: NSC 687852)

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b-AP15 (Synonyms: NSC 687852) 纯度: 98.75%

b-AP15 特异性抑制去泛素化酶 (deubiquitinating) UCHL5Usp14活性。

b-AP15(Synonyms: NSC 687852)

b-AP15 Chemical Structure

CAS No. : 1009817-63-3

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b-AP15 相关产品

相关化合物库:

  • Bioactive Compound Library Plus
  • Peptidomimetic Library

生物活性

b-AP15 is a specific inhibitor of the deubiquitinating enzymes UCHL5 and Usp14.

IC50 & Target

UCHL5/Usp14[1]

体外研究
(In Vitro)

Purified 19S proteasomes (5 nM) are treated with indicated concentrations of b-AP15 and DUB activity is determined by detectionof Ub-AMC cleavage. The IC50 value (2.1±0.411 μM) is determined from log concentration curves in Graph Pad Prism using non linear regression analysis. b-AP15 as a previously unidentified class of proteasome inhibitor that abrogates the deubiquitinating activity of the 19S regulatory particle. b-AP15 inhibited the activity of two 19S regulatory-particle-associated deubiquitinases, ubiquitin C-terminal hydrolase 5 (UCHL5) and ubiquitin-specific peptidase 14 (USP14), resulting in accumulation of polyubiquitin. b-AP15 induced tumor cell apoptosis that is insensitive to TP53 status and overexpression of the apoptosis inhibitor BCL2[1]. The ability of b-AP15 is determined to inhibit proteasome deubiquitinase activity using Ub-AMC as the substrate. An IC50 of 16.8±2.8 μM is observed[2]. b-AP15 is a specific USP14 and UCHL5 inhibitor, which blocks growth and induces apoptosis in MM cells[3].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

体内研究
(In Vivo)

b-AP15 (2.5 mg/kg) inhibits tumor growth in syngenic mice models with less frequent administration schedules. We administered b-AP15 to C57BL/6J mice with Lewis lung carcinomas (LLCs) using a 2-d-on, 2-d-off schedule and to BALB/c mice with orthotopic breast carcinoma (4T1) using a 1-d-on, 3-d-off schedule. b-AP15 significantly inhibited tumor growth in both models, with T/C=0.16 (P≤0.01) for the C57BL/6J mice and T/C=0.25 (P≤0.001) for the BALB/c mice. A reduction in the number of pulmonary metastases also is observed in the group of mice with 4T1 breast carcinomas treated with b-AP15[1].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

分子量

419.39

Formula

C22H17N3O6

CAS 号

1009817-63-3

运输条件

Room temperature in continental US; may vary elsewhere.

储存方式
Powder -20°C 3 years
4°C 2 years

*该产品在溶液状态不稳定,建议您现用现配,即刻使用。

溶解性数据
In Vitro: 

DMSO : 20 mg/mL (47.69 mM; Need ultrasonic)

配制储备液
浓度 溶剂体积 质量 1 mg 5 mg 10 mg
1 mM 2.3844 mL 11.9221 mL 23.8442 mL
5 mM 0.4769 mL 2.3844 mL 4.7688 mL
10 mM 0.2384 mL 1.1922 mL 2.3844 mL

*

请根据产品在不同溶剂中的溶解度,选择合适的溶剂配制储备液;该产品在溶液状态不稳定,建议您现用现配,即刻使用

In Vivo:

请根据您的实验动物和给药方式选择适当的溶解方案。以下溶解方案都请先按照 In Vitro 方式配制澄清的储备液,再依次添加助溶剂:

——为保证实验结果的可靠性,澄清的储备液可以根据储存条件,适当保存;体内实验的工作液,建议您现用现配,当天使用; 以下溶剂前显示的百
分比是指该溶剂在您配制终溶液中的体积占比;如在配制过程中出现沉淀、析出现象,可以通过加热和/或超声的方式助溶

  • 1.

    请依序添加每种溶剂: 10% DMSO    40% PEG300    5% Tween-80    45% saline

    Solubility: 2.08 mg/mL (4.96 mM); Suspended solution; Need ultrasonic

    此方案可获得 2.08 mg/mL (4.96 mM) 的均匀悬浊液,悬浊液可用于口服和腹腔注射。

    以 1 mL 工作液为例,取 100 μL 20.8 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中,混合均匀;向上述体系中加入50 μL Tween-80,混合均匀;然后继续加入 450 μL生理盐水定容至 1 mL。

    将 0.9 g 氯化钠,完全溶解于 100 mL ddH₂O 中,得到澄清透明的生理盐水溶液

*以上所有助溶剂都可在 上海金畔生物科技有限公司 网站选购。
参考文献
  • [1]. D’Arcy P, et al. Inhibition of proteasome deubiquitinating activity as a new cancer therapy. Nat Med. 2011 Nov 6;17(12):1636-40.

    [2]. Wang X, et al. The 19S Deubiquitinase Inhibitor b-AP15 is Enriched in Cells and Elicits Rapid Commitment to Cell Death. Mol Pharmacol. 2014 Jun;85(6):932-45.

    [3]. Tian Z, et al. A novel small molecule inhibitor of deubiquitylating enzyme USP14 and UCHL5 induces apoptosis in multiple myeloma and overcomes bortezomib resistance. Blood. 2014 Jan 30;123(5):706-16.

Kinase Assay
[1]

For deubiquitinase inhibition assays, 19S regulatory particle (5 nM), 26S (5 nM) UCH-L1 (5 nM), UCH-L3 (0.3 nM), USP2CD (5 nM) USP7CD (5 nM) USP8CD (5 nM) or BAP1 (5 nM) is incubated with DMSO or b-AP15 and monitored the cleavage of ubiquitin-AMC (1,000 nM) using a Wallac VICTOR Multilabel counter or a Tecan Infinite M1000 equipped with 380 nm excitation and 460 nm emission filters[1].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

Cell Assay
[2]

Cell viability is monitored by either the fluorometric microculture cytotoxicity assay or the MTT assay. For the MTT assay, cells are seeded into 96-well flat-bottomed plates overnight and exposed to drugs, using DMSO as the control. At the end of incubations, 10 µl of a stock solution of 5 mg/mL MTT is added into each well, and the plates are incubated 4 hours at 37°C. Formazan crystals are dissolved with 100 µL 10% SDS/10 mM HCl solution overnight at 37°C. Absorbance is measured using an enzyme-linked immunosorbent assay (ELISA) plate reader at 590 nm[2].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

Animal Administration
[1]

Mice[1]
For the squamous carcinoma model, 1×106 FaDu cells are subcutaneously injected into the right rear flank of female SCID mice. Tumor growth is measured by the formula length×width2×0.44. When tumors have grown to a size of approximately 200 mm3 (defined as day 0), mice are randomized to receive either vehicle (n=10) or b-AP15 (n=15) at 5 mg per kg of body weight by daily subcutaneous injection. For the colon carcinoma model, we subcutaneously injected 2.5×106 HCT-116 colon carcinoma cells overexpressing Bcl2 into the right flank of female nude mice. We treated mice with 5 mg of b-AP15 per kg of body weight by intraperitoneal injection. For the lung carcinoma model, we subcutaneously injected 2×105 LLC cells into the right rear flank of female C57/B6 mice. When tumors had grown to a size of approximately 50 mm3 (defined as day 0), we randomized mice to receive either vehicle (n=4) or b-AP15 (n=4) at 5 mg per kg of body weight intraperitoneally, with a treatment cycle consisting of 2 d of treatment followed by 2 d of rest (2 d on, 2 d off) for 2 weeks.

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

参考文献
  • [1]. D’Arcy P, et al. Inhibition of proteasome deubiquitinating activity as a new cancer therapy. Nat Med. 2011 Nov 6;17(12):1636-40.

    [2]. Wang X, et al. The 19S Deubiquitinase Inhibitor b-AP15 is Enriched in Cells and Elicits Rapid Commitment to Cell Death. Mol Pharmacol. 2014 Jun;85(6):932-45.

    [3]. Tian Z, et al. A novel small molecule inhibitor of deubiquitylating enzyme USP14 and UCHL5 induces apoptosis in multiple myeloma and overcomes bortezomib resistance. Blood. 2014 Jan 30;123(5):706-16.

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