FK866 (APO866)
(Synonyms: 达珀利奈; FK866; APO866) 目录号 : GC14308
Inhibitor of nicotinamide phosphoribosyltransferase
Cas No.:658084-64-1
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >99.50%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
| Kinase experiment [1]: | |
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Preparation Method |
To determine the IC50 of inhibitors, 5 μl FK866 (APO866) solutions (containing 10% DMSO) with various concentrations were added into 96-well plate. The plate was incubated at 37 ℃ for 5 min after addition of 16.5 μl reaction buffer containing NAMPT. The enzyme reactions were initiated by 4.5 μl NAM (1.11 μM) following NMN measurement as described above. The IC50 values were determined by non-linear fitting of the concentration-dependent curves with the four-parameter IC50 logistic equation. |
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Reaction Conditions |
37℃ for 5 min |
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Applications |
IC50 of FK866 (APO866) on NAMPT activity is 1.60±0.32 nmol/L |
| Cell experiment [1]: | |
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Cell lines |
HepG2 |
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Preparation Method |
Cells were seeded in 96-well plate and starved for over 12 h with serum-free DMEM at 60~70% confluency, then treated with FK866 (APO866) or vehicle for 24 h to 72 h according to experiment requirements.10 μl CCK-8 solution was added to the culture medium and incubated at 37 °C for 1 h. The absorbance at 450 nm (A450) was detected by a plate reader. |
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Reaction Conditions |
48 h |
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Applications |
IC50 of FK866 (APO866) on HepG2 cells is 2.21±0.21 nmol/L. |
| Cell experiment [2]: | |
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Cell lines |
RAW 264.7 and MODE-K cells |
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Preparation Method |
Cells were stimulated with ultrapure 100ng/mL lipopolysaccharide (LPS) or 1µg/mL flagellin followed by incubation with or without 200nM FK866 (APO866) overnight. Supernatants were harvested and protein was extracted with NE-PER containing protease and phosphatase inhibitors and stored at -80℃ until further workup. |
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Reaction Conditions |
200nM FK866 (APO866) overnight |
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Applications |
FK866 (APO866)treatment strongly reduced NF-κB phosphorylation consequent to LPS treatment. |
| Animal experiment [2]: | |
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Animal models |
8-week-old female wild-type (WT) or Rag1tm1Mom/J (Rag1−/−) mice |
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Preparation method |
Acute colitis was induced in mice with 3.5% or 3% dextran sulfate sodium ad libitum for 5 consecutive days, followed by a tap water period until end of experiments. Control mice received tap water during the study period. Mice were injected intraperitoneally with 10mg/kg bodyweight FK866 (APO866) or vehicle control twice daily until termination of experiments. |
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Dosage form |
Intraperitoneally with 10mg/kg FK866 (APO866) |
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Applications |
FK866 (APO866) significantly ameliorated all features of DSS-induced colitis in Rag1−/− mice. |
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References: [1]. Zhang SL, Xu TY, Yang ZL, Han S, Zhao Q, Miao CY. Crystal structure-based comparison of two NAMPT inhibitors. Acta Pharmacol Sin. 2018 Feb;39(2):294-301. [2]. Gerner RR, Klepsch V, Macheiner S, Arnhard K, Adolph TE, Grander C, Wieser V, Pfister A, Moser P, Hermann-Kleiter N, Baier G, Oberacher H, Tilg H, Moschen AR. NAD metabolism fuels human and mouse intestinal inflammation. Gut. 2018 Oct;67(10):1813-1823. |
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FK866 (APO866) is an inhibitor of nicotinamide phosphoribosyltransferase (NMPRTase). FK866 (APO866) protects against experimental colitis and colitis−associated tumorigenesis by suppression of activated leukocytes particularly macrophages, inflammatory monocytes and T cells. FK866(APO866) also reduced inflammatory responses of lamina propria mononuclear cells (LPMNC) from colonic biopsies of patients with IBD to a comparable extent as dexamethasone [1].
The IC50 of FK866 (APO866) on NAMPT activity is 1.60±0.32 nmol/L [2].. IC50 of FK866 (APO866) on HepG2 cells is 2.21±0.21 nmol/L. FK866 (APO866) treatment strongly reduced NF-κB phosphorylation consequent to LPS treatment. Inhibition of NAMPT by FK866, or inhibition of SIRT by nicotinamide decreased proliferation and triggered death of 293T cells involving the p53 acetylation pathway [3].. FK866 (APO866) potently inhibited NAMPT activity as demonstrated by reduced mucosal NAD, resulting in reduced abundances and activities of NAD-dependent enzymes including PARP1, Sirt6 and CD38, reduced nuclear factor kappa B activation, and decreased cellular infiltration by inflammatory monocytes, macrophages and activated T cells [1].
FK866 (APO866) significantly ameliorated all features of DSS-induced colitis in Rag1−mice and effectively suppresses inflammatory innate immune responses in the absence of adaptive immunity. FK866 (APO866) significantly reduced chemokine and cytokine release, many of those which are macrophage/monocyte derived. Remarkably, the observed suppression was in the range or even superior to well-established anti-inflammatory compounds such as dexamethasone and infliximab [1].
References:
[1].Gerner RR, Klepsch V, Macheiner S, Arnhard K, Adolph TE, Grander C, Wieser V, Pfister A, Moser P, Hermann-Kleiter N, Baier G, Oberacher H, Tilg H, Moschen AR. NAD metabolism fuels human and mouse intestinal inflammation. Gut. 2018 Oct;67(10):1813-1823.
[2].Zhang SL, Xu TY, Yang ZL, Han S, Zhao Q, Miao CY. Crystal structure-based comparison of two NAMPT inhibitors. Acta Pharmacol Sin. 2018 Feb;39(2):294-301.
[3].Thakur BK, Dittrich T, Chandra P, Becker A, Lippka Y, Selvakumar D, Klusmann JH, Reinhardt D, Welte K. Inhibition of NAMPT pathway by FK866 activates the function of p53 in HEK293T cells. Biochem Biophys Res Commun. 2012 Aug 3;424(3):371-7.
FK866 (APO866) 是烟酰胺磷酸核糖转移酶 (NMPRTase) 的抑制剂。 FK866 (APO866) 通过抑制活化的白细胞特别是巨噬细胞、炎性单核细胞和 T 细胞来防止实验性结肠炎和结肠炎相关的肿瘤发生。 FK866(APO866) 还可降低 IBD 患者结肠活检组织固有层单核细胞 (LPMNC) 的炎症反应，其程度与地塞米松相当[1]。
FK866 (APO866) 对 NAMPT 活性的 IC50 为 1.60±0.32 nmol/L [2].. FK866 (APO866) 对 HepG2 细胞的 IC50 为 2.21±0.21 nmol/L。 FK866 (APO866) 处理强烈降低 LPS 处理后的 NF-κB 磷酸化。 FK866 对 NAMPT 的抑制或烟酰胺对 SIRT 的抑制减少了涉及 p53 乙酰化途径的 293T 细胞的增殖并触发了其死亡[3]..FK866 (APO866) 有效抑制了 NAMPT 活性，如粘膜减少所证明的NAD，导致包括 PARP1、Sirt6 和 CD38 在内的 NAD 依赖性酶的丰度和活性降低，核因子 kappa B 活化减少，炎症单核细胞、巨噬细胞和活化 T 细胞的细胞浸润减少[1] .
FK866 (APO866) 显着改善 Rag1-/- 小鼠中 DSS 诱导的结肠炎的所有特征，并在缺乏适应性免疫的情况下有效抑制炎症性先天免疫反应。 FK866 (APO866) 显着减少趋化因子和细胞因子的释放，其中许多是巨噬细胞/单核细胞衍生的。值得注意的是，观察到的抑制作用在范围内甚至优于地塞米松和英夫利昔单抗等公认的抗炎化合物 [1]。
| Cas No. | 658084-64-1 | SDF | |
| 别名 | 达珀利奈; FK866; APO866 | ||
| 化学名 | (E)-N-[4-(1-benzoylpiperidin-4-yl)butyl]-3-pyridin-3-ylprop-2-enamide | ||
| Canonical SMILES | C1CN(CCC1CCCCNC(=O)C=CC2=CN=CC=C2)C(=O)C3=CC=CC=C3 | ||
| 分子式 | C24H29N3O2 | 分子量 | 391.51 |
| 溶解度 | ≥ 19.6mg/mL in DMSO, ≥ 49.6 mg/mL in EtOH | 储存条件 | Store at -20°C |
| General tips | 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；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。 储备液的保存方式和期限：-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。 为了提高溶解度，请将管子加热至37℃，然后在超声波浴中震荡一段时间。 |
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| Shipping Condition | 评估样品解决方案：配备蓝冰进行发货。所有其他可用尺寸：配备RT，或根据请求配备蓝冰。 | ||
| 制备储备液 | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 2.5542 mL | 12.7711 mL | 25.5421 mL |
| 5 mM | 0.5108 mL | 2.5542 mL | 5.1084 mL |
| 10 mM | 0.2554 mL | 1.2771 mL | 2.5542 mL |
| 第一步：请输入基本实验信息（考虑到实验过程中的损耗，建议多配一只动物的药量） | ||||||||||
| 给药剂量 | mg/kg |  |
动物平均体重 | g | |
每只动物给药体积 | ul | |
动物数量 | 只 |
| 第二步：请输入动物体内配方组成（配方适用于不溶于水的药物；不同批次药物配方比例不同，请联系GLPBIO为您提供正确的澄清溶液配方） | ||||||||||
| % DMSO % % Tween 80 % saline | ||||||||||
| 计算重置 | ||||||||||
计算结果:
工作液浓度： mg/ml；
DMSO母液配制方法： mg 药物溶于 μL DMSO溶液（母液浓度 mg/mL，
体内配方配制方法：取 μL DMSO母液，加入 μL PEG300，混匀澄清后加入μL Tween 80，混匀澄清后加入 μL saline，混匀澄清。
1. 首先保证母液是澄清的；
2.
一定要按照顺序依次将溶剂加入，进行下一步操作之前必须保证上一步操作得到的是澄清的溶液，可采用涡旋、超声或水浴加热等物理方法助溶。
3. 以上所有助溶剂都可在 GlpBio 网站选购。
NAD metabolism fuels human and mouse intestinal inflammation
Gut2018 Oct;67(10):1813-1823.PMID: 28877980DOI: 10.1136/gutjnl-2017-314241
Objective: Nicotinamide phosphoribosyltransferase (NAMPT, also referred to as pre-B cell colony-enhancing factor or visfatin) is critically required for the maintenance of cellular nicotinamide adenine dinucleotide (NAD) supply catalysing the rate-limiting step of the NAD salvage pathway. NAMPT is strongly upregulated in inflammation including IBD and counteracts an increased cellular NAD turnover mediated by NAD-depleting enzymes. These constitute an important mechanistic link between inflammatory, metabolic and transcriptional pathways and NAD metabolism.
Design: We investigated the impact of NAMPT inhibition by the small-molecule inhibitor FK866 in the dextran sulfate sodium (DSS) model of colitis and the azoxymethane/DSS model of colitis-associated cancer. The impact of NAD depletion on differentiation of mouse and human primary monocytes/macrophages was studied in vitro. Finally, we tested the efficacy of FK866 compared with dexamethasone and infliximab in lamina propria mononuclear cells (LPMNC) isolated from patients with IBD.
Results: FK866 ameliorated DSS-induced colitis and suppressed inflammation-associated tumorigenesis in mice. FK866 potently inhibited NAMPT activity as demonstrated by reduced mucosal NAD, resulting in reduced abundances and activities of NAD-dependent enzymes including PARP1, Sirt6 and CD38, reduced nuclear factor kappa B activation, and decreased cellular infiltration by inflammatory monocytes, macrophages and activated T cells. Remarkably, FK866 effectively supressed cytokine release from LPMNCs of patients with IBD. As FK866 was also effective in Rag1-⁄- mice, we mechanistically linked FK866 treatment with altered monocyte/macrophage biology and skewed macrophage polarisation by reducing CD86, CD38, MHC-II and interleukin (IL)-6 and promoting CD206, Egr2 and IL-10.
Conclusion: Our data emphasise the importance of NAD immunometabolism for mucosal immunity and highlight FK866-mediated NAMPT blockade as a promising therapeutic approach in acute intestinal inflammation.
Selective Cytotoxicity of the NAMPT Inhibitor FK866 Toward Gastric Cancer Cells With Markers of the Epithelial-Mesenchymal Transition, Due to Loss of NAPRT
Gastroenterology2018 Sep;155(3):799-814.e13.PMID: 29775598DOI: 10.1053/j.gastro.2018.05.024
Background & aims: Markers of the epithelial-to-mesenchymal transition (EMT) in gastric tumor tissues are associated with poor patient outcomes. We performed a screen to identify pharmacologic compounds that kill gastric cancer cells with EMT-associated gene expression patterns and investigate their mechanisms.
Methods: We identified 29 gastric cancer cell lines with a gene expression signature previously associated with an EMT subtype, based on data from RNA sequence analyses, and confirmed the mesenchymal phenotypes of 7 lines (Hs746T, SNU1750, MKN1, SK4, SNU484, SNU668, and YCC11), based on invasive activity and protein markers. We screened 1,345 compounds for their ability to kill cells with the EMT signature compared with cell lines without this pattern. We tested the effects of identified compounds in BALB/c nude mice bearing GA077 tumors; mice were given intraperitoneal injections of the compound or vehicle (control) twice daily for 24 days and tumor growth was monitored. Proteins associated with the toxicity of the compounds were overexpressed in MKN1 and SNU484 cells or knocked down in MKN45 and SNU719 using small interfering RNAs. We performed immunohistochemical analyses of 942 gastric cancer tissues and investigated associations between EMT markers and protein expression patterns.
Results: The nicotinamide phosphoribosyltransferase inhibitor FK866 killed 6 of 7 gastric cancer cell lines with EMT-associated gene expression signatures but not gastric cancer cells without this signature. The 6 EMT-subtype gastric cell lines expressed significantly low levels of nicotinic acid phosphoribosyltransferase (NAPRT), which makes the cells hypersensitive to nicotinamide phosphoribosyltransferase inhibition. Gastric cell lines that expressed higher levels of NAPRT, regardless of EMT markers, were sensitized to FK866 after knockdown of NAPRT, whereas overexpression of NAPRT in deficient EMT cell lines protected them from FK866-mediated toxicity. Administration of FK866 to nude mice with tumors grown from GA077 cells (human gastric cancer tumors of the EMT subtype) led to tumor regression in 2 weeks; FK866 did not affect tumors grown from MKN45 cells without the EMT expression signature. Loss of NAPRT might promote the EMT, because it stabilizes β-catenin. We correlated the EMT gene expression signature with lower levels of NAPRT in 942 gastric tumors from patients; we also found lower levels of NAPRT mRNA in colorectal, pancreatic, and lung adenocarcinoma tissues with the EMT gene expression signature.
Conclusions: FK866 selectively kills gastric cancer cells with an EMT gene expression signature by inhibiting nicotinamide phosphoribosyltransferase in cells with NAPRT deficiency. Loss of NAPRT expression, frequently through promoter hypermethylation, is observed in many gastric tumors of the EMT subtype. FK866 might be used to treat patients with tumors of this subtype.
Lipopolysaccharide-Induced Microglial Neuroinflammation: Attenuation by FK866
Neurochem Res2021 May;46(5):1291-1304.PMID: 33713324DOI: 10.1007/s11064-021-03267-4
Alleviating microglia-mediated neuroinflammation bears great promise to reduce neurodegeneration. Nicotinamide phosphoribosyltransferase (NAMPT) may exert cytokine-like effect in the brain. However, it remains unclear about role of NAMPT in microglial inflammation. Also, it remains unknown about effect of NAMPT inhibition on microglial inflammation. In the present study, we observed that FK866 (a specific noncompetitive NAMPT inhibitor) dose-dependently inhibited lipopolysaccharide (LPS)-induced proinflammatory mediator (interleukin (IL)-6, IL-1β, inducible nitric oxide synthase, nitric oxide and reactive species) level increase in BV2 microglia cultures. FK866 also significantly inhibited LPS-induced polarization change in microglia. Furthermore, LPS significantly increased NAMPT expression and nuclear factor kappa B (NF-κB) phosphorylation in microglia. FK866 significantly decreased NAMPT expression and NF-κB phosphorylation in LPS-treated microglia. Finally, conditioned medium from microglia cultures co-treated with FK866 and LPS significantly increased SH-SY5Y and PC12 cell viability compared with conditioned medium from microglia cultures treated with LPS alone. Our study strongly indicates that NAMPT may be a promising target for microglia modulation and NAMPT inhibition may attenuate microglial inflammation.
FK866 attenuates acute hepatic failure through c-jun-N-terminal kinase (JNK)-dependent autophagy
Sci Rep2017 May 19;7(1):2206.PMID: 28526886DOI: 10.1038/s41598-017-02318-7
FK866 exhibits a protective effect on D-galactosamine (GaIN)/lipopolysaccharide (LPS) and concanavalin A (ConA)-induced acute liver failure (ALF), but the mechanism by which FK866 affords this benefit has not yet been elucidated. Autophagy has a protective effect on acute liver injury. However, the contribution of autophagy to FK866-conferred hepatoprotection is still unclear. This study aimed to investigate whether FK866 could attenuate GaIN/LPS and ConA-induced ALF through c-jun-N-terminal kinase (JNK)-dependent autophagy. In vivo, Mice were pretreated with FK866 at 24, 12, and 0.5 h before treatment with GaIN/LPS and ConA. 3-methyladenine (3MA) or rapamycin were used to determine the role of autophagy in FK866-conferred hepatoprotection. In primary hepatocytes, autophagy was inhibited by 3MA or autophagy-related protein 7 (Atg7) small interfering RNA (siRNA). JNK was suppressed by SP600125 or Jnk siRNA. FK866 alleviated hepatotoxicity and increased autophagy while decreased JNK activation. Suppression of autophagy abolished the FK866-conferred protection. Inhibition of JNK increased autophagy and exhibited strongly protective effect. Collectively, FK866 could ameliorate GaIN/LPS and ConA-induced ALF through induction of autophagy while suppression of JNK. These findings suggest that FK866 acts as a simple and applicable preconditioning intervention to protect against ALF; autophagy and JNK may also provide therapeutic targets for ALF treatment.
FK866 attenuates sepsis-induced acute lung injury through c-jun-N-terminal kinase (JNK)-dependent autophagy
Life Sci2020 Jun 1;250:117551.PMID: 32179075DOI: 10.1016/j.lfs.2020.117551
Aims: Increasing evidence indicates that FK866, a specific noncompetitive nicotinamide phosphoribosyl transferase inhibitor, exhibits a protective effect on acute lung injury (ALI). Autophagy plays a pivotal role in sepsis-induced ALI. However, the contribution of autophagy and the underlying mechanism by which FK866-confered lung protection remains elusive. Herein, we aimed to study whether FK866 could alleviate sepsis-induced ALI via the JNK-dependent autophagy.
Main methods: Male C57BL/6 mice were subjected to cecal ligation and puncture (CLP) to establish the polymicrobial sepsis mice model, and treated with FK866 (10 mg/kg) at 24, 12 and 0.5 h before the CLP procedure. The lung protective effects were measured by lung histopathology, tissue edema, vascular leakage, inflammation infiltration, autophagy-related protein expression and JNK activity. A549 cells were stimulated with LPS (1000 ng/ml) to generate the ALI cell model, and pretreated with FK866 or SP600125 for 30 min to measure the autophagy-related protein expression and JNK activity.
Key findings: Our results demonstrated that FK866 reduced lung injury score, tissue edema, vascular leakage, and inflammatory infiltration, and upregulated autophagy. The protective effect of autophagy conferred by FK866 on ALI was further clarified by using 3-methyladenine (3MA) and rapamycin. Additionally, the activity of JNK was suppressed by FK866, and inhibition of JNK promoted autophagy and showed a benefit effect.
Significance: Our study indicates that FK866 protects against sepsis-induced ALI by induction of JNK-dependent autophagy. This may provide new insights into the functional mechanism of NAMPT inhibition in sepsis-induced ALI.