HG-9-91-01
(Synonyms: N-(2,4-二甲氧基苯基)-N'-(2,6-二甲基苯基)-N-[6-[[4-(4-甲基-1-哌嗪基)苯基]氨基]-4-嘧啶基]脲,SIK inhibitor 1) 目录号 : GC11719
An inhibitor of salt-inducible kinases
Cas No.:1456858-58-4
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >99.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
| Cell experiment [1]: | |
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Cell lines |
Bone marrow-derived macrophages |
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Preparation Method |
Powder was dissolved in DMSO as 10 mM stock solutions and stored at -20℃ until use. BMM cells were plated at concentration of 160 x 103 cells/well in 24 wells osteo assay plates coated with bone biomimetic synthetic surface and cultured with rmM-CSF (30ng/ml) and rmRANKL (50ng/ml) for 3 days in absence or presence of different concentrations of HG-9-91-01. |
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Reaction Conditions |
0, 0.1, 0.3, 0.5µM for 72 days |
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Applications |
No significant toxicity was observed at the concentrations tested. Pre-treatment with 0.5 µM HG-9-91-01 significantly reduced the formation of resorption lacunae compared to RANKL alone. |
| Animal experiment [2]: | |
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Animal models |
TNBS-induced colitis in female BALB/c mice, C57BL/6J mice |
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Preparation Method |
The TNBS-challenged mice were treated with different doses of HG-9-91-01 (3, 10, 30 mg/kg/day intraperitoneally (i.p.), dissolved in 100 µL olive oil). |
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Dosage form |
Intraperitoneal injection, 3, 10, 30 mg/kg/day for 8 days. |
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Applications |
TNBS-treated mice given HG-9-91-01 at 3, 10, or 30 mg/kg rapidly recovered their lost body weight. The macroscopic inflammatory signs (gross bleeding, ulceration) and the shortening of colon length typically caused by TNBS were also improved by treatment with 3, 10, or 30 mg/kg HG-9-91-01 |
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References: [1]: Lombardi M S, GilliÉron C, Berkelaar M, et al. Salt-inducible kinases (SIK) inhibition reduces RANKL-induced osteoclastogenesis[J]. PloS one, 2017, 12(10): e0185426. |
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HG-9-91-01 is a salt-inducible kinases (SIK) inhibitor, inhibited SIK1, SIK2, SIK3 with IC50 values of 0.92nM, 6.5nM and 19.4nM, respectively [1].
HG-9-91-01 potently inhibited the SIKs and did not inhibit any other member of the AMPK-related kinase subfamily. HG-9-91-01 increased LPS-stimulated IL-10 production and suppressed proinflammatory cytokine secretion [2]. 0.5 µM HG-9-91-01 pretreated RAW264.7 for 30 min before RANKL stimulation resulted in reduction of multinucleated cell formation and of TRAP staining, and significantly reduced the mRNA of osteoclast differentiation markers in a concentration dependent manner. Pre-treatment with 0.5 µM HG-9-91-01 significantly reduced the formation of resorption lacunae compared to RANKL alone [3].
HG-9-91-01 (3, 10, or 30 mg/kg) enhanced the expression of IL-10 whereas downregulated the levels of IL-12 and TNF-α in the colon tissue of the TNBS mice [4].
References:
[1]. Sundberg T B, Choi H G, Song J H, et al. Small-molecule screening identifies inhibition of salt-inducible kinases as a therapeutic strategy to enhance immunoregulatory functions of dendritic cells[J]. Proceedings of the National Academy of Sciences, 2014, 111(34): 12468-12473.
[2]. Clark K, MacKenzie K F, Petkevicius K, et al. Phosphorylation of CRTC3 by the salt-inducible kinases controls the interconversion of classically activated and regulatory macrophages[J]. Proceedings of the National Academy of Sciences, 2012, 109(42): 16986-16991.
[3]. Lombardi M S, GilliÉron C, Berkelaar M, et al. Salt-inducible kinases (SIK) inhibition reduces RANKL-induced osteoclastogenesis[J]. PloS one, 2017, 12(10): e0185426.
[4]. Fu Y, Ma G, Zhang Y, et al. HG-9-91-01 Attenuates murine experimental colitis by promoting interleukin-10 production in colonic macrophages through the SIK/CRTC3 pathway[J]. Inflammatory Bowel Diseases, 2021, 27(11): 1821-1831.
HG-9-91-01 是一种盐诱导激酶 (SIK) 抑制剂，可抑制 SIK1、SIK2、SIK3，IC50 值分别为 0.92nM、6.5nM 和 19.4nM[1].
HG-9-91-01 有效抑制 SIK，但不抑制 AMPK 相关激酶亚家族的任何其他成员。 HG-9-91-01 增加 LPS 刺激的 IL-10 产生并抑制促炎细胞因子分泌[2]。 0.5 µM HG-9-91-01 在 RANKL 刺激之前将 RAW264.7 预处理 30 分钟导致多核细胞形成和 TRAP 染色减少，并以浓度依赖性方式显着减少破骨细胞分化标志物的 mRNA。与单独使用 RANKL 相比，使用 0.5 µM HG-9-91-01 进行预处理可显着减少吸收陷窝的形成[3]。
HG-9-91- 01（3、10 或 30 mg/kg）增强 IL-10 的表达，同时下调 IL-12 和 TNF-α 的水平；在TNBS小鼠结肠组织中[4]。
| Cas No. | 1456858-58-4 | SDF | |
| 别名 | N-(2,4-二甲氧基苯基)-N'-(2,6-二甲基苯基)-N-[6-[[4-(4-甲基-1-哌嗪基)苯基]氨基]-4-嘧啶基]脲,SIK inhibitor 1 | ||
| 化学名 | (Z)-N-(2,4-dimethoxyphenyl)-N'-(2,6-dimethylphenyl)-N-(6-((4-(4-methylpiperazin-1-yl)phenyl)amino)pyrimidin-4-yl)carbamimidic acid | ||
| Canonical SMILES | CC1=C(/N=C(O)/N(C2=NC=NC(NC3=CC=C(N4CCN(CC4)C)C=C3)=C2)C5=C(OC)C=C(OC)C=C5)C(C)=CC=C1 | ||
| 分子式 | C32H37N7O3 | 分子量 | 567.68 |
| 溶解度 | ≥ 56.8mg/mL in DMSO | 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 | 1.7616 mL | 8.8078 mL | 17.6156 mL |
| 5 mM | 0.3523 mL | 1.7616 mL | 3.5231 mL |
| 10 mM | 0.1762 mL | 0.8808 mL | 1.7616 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。
HG-9-91-01 Attenuates Murine Experimental Colitis by Promoting Interleukin-10 Production in Colonic Macrophages Through the SIK/CRTC3 Pathway
Inflamm Bowel Dis2021 Oct 20;27(11):1821-1831.PMID: 33988718DOI: 10.1093/ibd/izab072
Background: Interleukin-10 (IL-10) is a potent immunoregulatory cytokine that plays a pivotal role in maintaining mucosal immune homeostasis. As a novel synthetic inhibitor of salt-inducible kinases (SIKs), HG-9-91-01 can effectively enhance IL-10 secretion at the cellular level, but its in vivo immunoregulatory effects remain unclear. In this study, we investigated the effects and underlying mechanism of HG-9-91-01 in murine colitis models. Methods: The anti-inflammatory effects of HG-9-91-01 were evaluated on 2, 4, 6-trinitrobenzene sulfonic acid (TNBS)-, dextran sulfate sodium-induced colitis mice, and IL-10 knockout chronic colitis mice. The in vivo effector cell of HG-9-91-01 was identified by fluorescence-activated cell sorting and quantitative real-time polymerase chain reaction. The underlying mechanism of HG-9-91-01 was investigated via overexpressing SIKs in ANA-1 macrophages and TNBS colitis mice. Results: Treatment with HG-9-91-01 showed favorable anticolitis effects in both TNBS- and DSS-treated mice through significantly promoting IL-10 expression in colonic macrophages but failed to protect against IL-10 KO murine colitis. Further study indicated that HG-9-91-01 markedly enhanced the nuclear level of cAMP response element-binding protein (CREB)-regulated transcription coactivator 3 (CRTC3), whereas treatment with lentiviruses encoding SIK protein markedly decreased the nuclear CRTC3 level in HG-9-91-01-treated ANA-1 macrophages. In addition, intracolonic administration with lentiviruses encoding SIK protein significantly decreased the nuclear CRTC3 level in the lamina propria mononuclear cells and ended the anti-inflammatory activities of HG-9-91-01. Conclusions: We found that HG-9-91-01 promoted the IL-10 expression of colonic macrophages and exhibited its anticolitis activity through the SIK/CRTC3 axis, and thus it may represent a promising strategy for inflammatory bowel disease therapy.
Salt-inducible kinases inhibitor HG-9-91-01 targets RIPK3 kinase activity to alleviate necroptosis-mediated inflammatory injury
Cell Death Dis2022 Feb 25;13(2):188.PMID: 35217652DOI: 10.1038/s41419-022-04633-y
Receptor-interacting protein kinase 3 (RIPK3) functions as a central regulator of necroptosis, mediating signaling transduction to activate pseudokinase mixed lineage kinase domain-like protein (MLKL) phosphorylation. Increasing evidences show that RIPK3 contributes to the pathologies of inflammatory diseases including multiple sclerosis, infection and colitis. Here, we identified a novel small molecular compound Salt-inducible Kinases (SIKs) inhibitor HG-9-91-01 inhibiting necroptosis by targeting RIPK3 kinase activity. We found that SIKs inhibitor HG-9-91-01 could block TNF- or Toll-like receptors (TLRs)-mediated necroptosis independent of SIKs. We revealed that HG-9-91-01 dramatically decreased cellular activation of RIPK3 and MLKL. Meanwhile, HG-9-91-01 inhibited the association of RIPK3 with MLKL and oligomerization of downstream MLKL. Interestingly, we found that HG-9-91-01 also trigger RIPK3-RIPK1-caspase 1-caspase 8-dependent apoptosis, which activated cleavage of GSDME leading to its dependent pyroptosis. Mechanistic studies revealed that SIKs inhibitor HG-9-91-01 directly inhibited RIPK3 kinase activity to block necroptosis and interacted with RIPK3 and recruited RIPK1 to activate caspases leading to cleave GSDME. Importantly, mice pretreated with HG-9-91-01 showed resistance to TNF-induced systemic inflammatory response syndrome. Consistently, HG-9-91-01 treatment protected mice against Staphylococcus aureus-mediated lung damage through targeting RIPK3 kinase activity. Overall, our results revealed that SIKs inhibitor HG-9-91-01 is a novel inhibitor of RIPK3 kinase and a potential therapeutic target for the treatment of necroptosis-mediated inflammatory diseases.
In vivo screen identifies a SIK inhibitor that induces ¦ cell proliferation through a transient UPR
Nat Metab2021 May;3(5):682-700.PMID: 34031592DOI: 10.1038/s42255-021-00391-x
It is known that ¦ cell proliferation expands the ¦ cell mass during development and under certain hyperglycemic conditions in the adult, a process that may be used for ¦ cell regeneration in diabetes. Here, through a new high-throughput screen using a luminescence ubiquitination-based cell cycle indicator (LUCCI) in zebrafish, we identify HG-9-91-01 as a driver of proliferation and confirm this effect in mouse and human ¦ cells. HG-9-91-01 is an inhibitor of salt-inducible kinases (SIKs), and overexpression of Sik1 specifically in ¦ cells blocks the effect of HG-9-91-01 on ¦ cell proliferation. Single-cell transcriptomic analyses of mouse ¦ cells demonstrate that HG-9-91-01 induces a wave of activating transcription factor (ATF)6-dependent unfolded protein response (UPR) before cell cycle entry. Importantly, the UPR wave is not associated with an increase in insulin expression. Additional mechanistic studies indicate that HG-9-91-01 induces multiple signalling effectors downstream of SIK inhibition, including CRTC1, CRTC2, ATF6, IRE1 and mTOR, which integrate to collectively drive ¦ cell proliferation.
SIK2 maintains breast cancer stemness by phosphorylating LRP6 and activating Wnt/¦catenin signaling
Oncogene2022 Apr;41(16):2390-2403.PMID: 35277657DOI: 10.1038/s41388-022-02259-0
Breast cancer stem cells (BCSCs) are the main drivers of recurrence and metastasis. However, commonly used drugs rarely target BCSCs. Via screenings, we found that Salt-inducible kinase 2 (SIK2) participated in breast cancer (BC) stemness maintenance and zebrafish embryos development. SIK2 was upregulated in recurrence samples. Knockdown of SIK2 expression reduced the proportion of BCSCs and the tumor initiation of BC cells. Mechanistically, SIK2, phosphorylated by CK1¦Á, directly phosphorylated LRP6 in a SIK2 kinase activity-dependent manner, leading to Wnt/¦catenin signaling pathway activation. ARN-3236 and HG-9-91-01, inhibitors of SIK2, inhibited LRP6 phosphorylation and ¦catenin accumulation and disturbed stemness maintenance. In addition, the SIK2-activated Wnt/¦catenin signaling led to induction of IDH1 expression, causing metabolic reprogramming in BC cells. These findings demonstrate a novel mechanism whereby Wnt/¦catenin signaling pathway is regulated by different kinases in response to metabolic requirement of CSCs, and suggest that SIK2 inhibition may potentially be a strategy for eliminating BCSCs.
SIK1 Regulates CRTC2-Mediated Gluconeogenesis Signaling Pathway in Human and Mouse Liver Cells
Front Endocrinol (Lausanne)2020 Sep 2;11:580.PMID: 33013689DOI: 10.3389/fendo.2020.00580
The regulation of hepatic gluconeogenesis is of great significance to improve insulin resistance and benefit diabetes therapy. cAMP-Regulated Transcriptional Co-activator 2 (CRTC2) plays a key role in regulating hepatic gluconeogenesis through controlling the expression of gluconeogenic rate-limiting enzymes such as glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase (PEPCK). Recently, salt-inducible kinase 1 (SIK1) has been identified to play an important role in glucose metabolism disorders, but whether and how SIK1 regulates the CTRC2 signaling in liver cells under high glucose conditions has rarely been intensively elucidated. Here, we show that high glucose stimulation resulted in time-dependent down-regulated expression of SIK1, phosphorylated SIK1 at T182 site, and phosphorylated CRTC2 at S171 site, as well as upregulated expression of total CRTC2 and its downstream targets G6Pase and PEPCK in the human liver cell line HepG2. The nuclear expression levels of SIK1 and CRTC2 were time-dependently upregulated upon high glucose challenge, which was accompanied by enhanced cytoplasm-to-nucleus translocation of SIK1. Manipulation of SIK1 activity using plasmid-mediated SIK1 over-expression and the use of the SIKs inhibitor HG-9-91-01 confirmed that SIK1 regulated the CRTC2 signaling pathway in HepG2 cells. Furthermore, in mouse primary hepatocytes, high glucose exposure down-regulated SIK1 expression, and promoted SIK1 nuclear accumulation. While HG-9-91-01 treatment suppressed SIK1 expression and released the inhibitory effects of SIK1 on the expressions of key molecules involved in the CRTC2 signaling pathway, additional ectopic expression of SIK1 using adenovirus infection reversed the impacts of HG-9-91-01 on the expressions of these molecules in mouse hepatocytes. Therefore, SIK1 regulates CRTC2-mediated gluconeogenesis signaling pathway under both physiological and high glucose-induced pathological conditions. The modulation of the SIK1-CRTC2 signaling axis could provide an attractive means for treating diabetes.