AZD8797
(Synonyms: KAND567) 目录号 : GC31653
AZD8797是非竞争的人CX3CR1变构调节剂，Ki分别为3.9，2800nM。
Cas No.:911715-90-7
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >98.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
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Kinase experiment: |
CHO-hCX3CR1 membranes together with different concentrations of AZD8797 are incubated in 50 mM HEPES, 100 mM NaCl, 5 mM MgCl2, 10 μM GDP and 0.01% gelatin in a MicroWell 96-well plate. 0.56 μCi/mL [35S]GTPγS and EC80 of CX3CL1 are then added. The plate is incubated at 30°C for 1 h and subsequently unbound [35S]GTPγS is separated from bound by vacuum filtration to a Printed Filtermat B. The different AZD8797 concentrations are achieved by stepwise dilution in DMSO to achieve a final DMSO concentration of 1% in all wells after addition of assay buffer, regardless of AZD8797 concentration[1]. |
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Animal experiment: |
Rats: AZD8797 is formulated in 30–35% (wt/wt) hydroxy-propyl-beta-cyklodextrin and administered s.c. through osmotic minipumps. Treatment is blinded to the operator. The plasma concentration of AZD8797 is analyzed twice from each rat[2]. |
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References: [1]. Cederblad L, et al. AZD8797 is an allosteric non-competitive modulator of the human CX3CR1 receptor. Biochem J. 2016 Mar 1;473(5):641-9. |
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AZD8797 is an allosteric non-competitive modulator of the human CX3CR1 receptor; antagonizes CX3CR1 and CXCR2 with Kis of 3.9 and 2800 nM, respectively.
In a flow adhesion assay, AZD8797 antagonizes the natural ligand, fractalkine (CX3CL1), in both human whole blood (hWB) and in a B-lymphocyte cell line with IC50 values of 300 and 6 nM respectively. AZD8797 also prevents G-protein activation in a [35S]GTPγS accumulation assay. AZD8797 positively modulates the CX3CL1 response at sub-micromolar concentrations in a β-arrestin recruitment assay. In equilibrium saturation binding experiments, AZD8797 reduces the maximal binding of 125I-CX3CL1 without affecting Kd[1]. AZD8797 binds selectively with high affinity to human and rat CX3CR1 (Ki of hCX3CR1, 4 nM; Ki of rCX3CR1, 7 nM, respectively). The equilibrium dissociation constant, KB, demonstrates that AZD8797 is a very potent inhibitor for human CX3CR1 (10 nM). The potency is threefold lower for rat CX3CR1 (29 nM) and decreases even further at mouse CX3CR1 (54 nM)[2].
AZD8797 treatment in Dark Agouti rats with myelin oligodendrocyte glycoprotein-induced EAE results in reduced paralysis, CNS pathology, and incidence of relapses. The compound is effective when starting treatment before onset, as well as after the acute phase[2].
[1]. Cederblad L, et al. AZD8797 is an allosteric non-competitive modulator of the human CX3CR1 receptor. Biochem J. 2016 Mar 1;473(5):641-9. [2]. Ridderstad Wollberg A, et al. Pharmacological inhibition of the chemokine receptor CX3CR1 attenuates disease in a chronic-relapsing rat model for multiple sclerosis. Proc Natl Acad Sci U S A. 2014 Apr 8;111(14):5409-14. [3]. Sofia Karlstro?, et al. Substituted 7-amino-5-thio-thiazolo[4,5-d]pyrimidines as potent and selective antagonists of the fractalkine receptor (CX3CR1). J Med Chem. 2013 Apr 25;56(8):3177-90.
| Cas No. | 911715-90-7 | SDF | |
| 别名 | KAND567 | ||
| Canonical SMILES | CC(C)C[C@@H](NC1=NC(S[C@@H](C)C2=CC=CC=C2)=NC3=C1SC(N)=N3)CO | ||
| 分子式 | C19H25N5OS2 | 分子量 | 403.56 |
| 溶解度 | DMSO : ≥ 150 mg/mL (371.69 mM) | 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 | 2.4779 mL | 12.3897 mL | 24.7795 mL |
| 5 mM | 0.4956 mL | 2.4779 mL | 4.9559 mL |
| 10 mM | 0.2478 mL | 1.239 mL | 2.4779 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。
AZD8797 is an allosteric non-competitive modulator of the human CX3CR1 receptor
The chemokine receptor CX3CR1 has been implicated as an attractive therapeutic target in several diseases, including atherosclerosis and diabetes. However, there has been a lack of non-peptide CX3CR1 inhibitors to substantiate these findings. A selective small-molecule inhibitor of CX3CR1, AZD8797, was recently reported and we present here an in-depth in vitro characterization of that molecule. In a flow adhesion assay, AZD8797 antagonized the natural ligand, fractalkine (CX3CL1), in both human whole blood (hWB) and in a B-lymphocyte cell line with IC50 values of 300 and 6 nM respectively. AZD8797 also prevented G-protein activation in a [(35)S]GTP污S (guanosine 5'-[污-thio]triphosphate) accumulation assay. In contrast, dynamic mass redistribution (DMR) experiments revealed a weak G汐i-dependent AZD8797 agonism. Additionally, AZD8797 positively modulated the CX3CL1 response at sub-micromolar concentrations in a 汕-arrestin recruitment assay. In equilibrium saturation binding experiments, AZD8797 reduced the maximal binding of (125)I-CX3CL1 without affecting Kd. Kinetic experiments, determining the kon and koff of AZD8797, demonstrated that this was not an artefact of irreversible or insurmountable binding, thus a true non-competitive mechanism. Finally we show that both AZD8797 and GTP污S increase the rate with which CX3CL1 dissociates from CX3CR1 in a similar manner, indicating a connection between AZD8797 and the CX3CR1-bound G-protein. Collectively, these data show that AZD8797 is a non-competitive allosteric modulator of CX3CL1, binding CX3CR1 and effecting G-protein signalling and 汕-arrestin recruitment in a biased way.
Neuron derived fractalkine promotes microglia to absorb hematoma via CD163/HO-1 after intracerebral hemorrhage
Background: Hematoma leads to progressive neurological deficits and poor outcomes after intracerebral hemorrhage (ICH). Early clearance of hematoma is widely recognized as an essential treatment to limit the damage and improve the clinical prognosis. CD163, alias hemoglobin (Hb) scavenger receptor on microglia, plays a pivotal role in hematoma absorption, but CD163 on neurons permits Hb uptake and results in neurotoxicity. In this study, we focus on how to specially promote microglial but not neuronal CD163 mediated-Hb uptake and hematoma absorption.
Methods: RNA sequencing was used to explore the potential molecules involved in ICH progression, and hematoma was detected by magnetic resonance imaging (MRI). Western blot and immunofluorescence were used to evaluate the expression and location of fractalkine (FKN) after ICH. Erythrophagocytosis assay was performed to study the specific mechanism of action of FKN in hematoma clearance. Small interfering RNA (siRNA) transfection was used to explore the effect of peroxisome proliferator-activated receptor-污 (PPAR-污) on hematoma absorption. Enzyme-linked immunosorbent assay (ELISA) was used to determine the serum FKN concentration in ICH patients.
Results: FKN was found to be significantly increased around the hematoma in a mouse model after ICH. With its unique receptor CX3CR1 in microglia, FKN significantly decreased the hematoma size and Hb content, and improved neurological deficits in vivo. Further, FKN could enhance erythrophagocytosis of microglia in vitro via the CD163/ hemeoxygenase-1 (HO-1) axis, while AZD8797 (a specific CX3CR1 inhibitor) reversed this effect. Moreover, PPAR-污 was found to mediate the increase in the CD163/HO-1 axis expression and erythrophagocytosis induced by FKN in microglia. Of note, a higher serum FKN level was found to be associated with better hematoma resolution in ICH patients.
Conclusions: We systematically identified that FKN may be a potential therapeutic target to improve hematoma absorption and we shed light on ICH treatment.
A novel CX3CR1 inhibitor AZD8797 facilitates early recovery of rat acute spinal cord injury by inhibiting inflammation and apoptosis
The present study aimed to evaluate the effect of the CX3CR1 inhibitor AZD8797 in early recovery after acute SCI and elucidate its potential mechanism in blocking inflammation and apoptosis. Adult rats were sacrificed after 3, 7, 10, or 14 days of SCI. The injured spinal tissues were collected for assessing C?X3?C motif chemokine ligand 1(CX3CL1)/C?X3?C motif chemokine receptor 1 (CX3CR1) expression at each time point via western blotting (WB) and quantitative PCR. The cellular localization of the proteins was detected by immunofluorescence. Another batch of rats (subdivided into sham, injury model, AZD8797 and methylprednisolone groups) were used to evaluate locomotive recovery with a Basso Beattie Bresnahan score. Based on the expression level of CX3CR1, these rats were sacrificed at the most prominent stage of CX3CR1 expression (10 days after SCI), for assessing the serum levels of tumor necrosis factor?汐/interleukin (IL)?6/IL?1汕 and the expression of CX3CL1/CX3CR1/caspase 3/Bcl?2/Bax in the spinal cord tissues through WB and ELISA. Additionally, apoptosis and necrosis in the injured spinal cord were evaluated by terminal deoxynucleotidyl transferase?-mediated dUTP nick?end labeling staining/fluoro?jade B staining. Expression levels of both CX3CR1 and CX3CL1 reached their peak 10 days after the injury, followed by a dramatic downward trend at 14 days. The enhanced expression of CX3CR1 was detected in astrocytes and microglia of the injured spinal cord. AZD8797 improved locomotive recovery after 10 days of SCI and was as effective as methylprednisolone. The effect of AZD8797 was mediated by suppressing apoptosis, necrosis and inflammatory responses, as assessed by WB/ELISA and morphological examinations. The current study has demonstrated that AZD8797 can effectively block overwhelming inflammation, apoptosis and necrosis after SCI and facilitate early recovery of locomotive function.
FKN/CX3CR1 axis facilitates migraine-Like behaviour by activating thalamic-cortical network microglia in status epilepticus model rats
Background: The incidence of migraines is higher among individuals with epilepsy than in healthy individuals, and these two diseases are thought to shared pathophysiological mechanisms. Excitation/inhibition imbalance plays an essential role in the comorbidity of epilepsy and migraine. Microglial activation is crucial for abnormal neuronal signal transmission. However, it remains unclear whether and how microglia are activated and their role in comorbidities after being activated. This study aimed to explore the characteristics and mechanism of microglial activation after seizures and their effect on migraine.
Methods: Model rats of status epilepticus (SE) induced by intraperitoneal injection of lithium chloride (LiCl)-pilocarpine and migraine induced by repeated dural injections of inflammatory soup (IS) were generated, and molecular and histopathologic evidence of the microglial activation targets of fractalkine (FKN) signalling were examined. HT22-BV2 transwell coculture assays were used to explore the interaction between neurons and microglia. LPS (a microglial agonist) and FKN stimulation of BV2 microglial cells were used to evaluate changes in BDNF levels after microglial activation.
Results: Microglia were specifically hyperplastic and activated in the temporal lobe cortex, thalamus, and spinal trigeminal nucleus caudalis (sp5c), accompanied by the upregulation of FKN and CX3CR1 four days after seizures. Moreover, SE-induced increases in nociceptive behaviour and FKN/CX3CR1 axis expression in migraine model rats. AZD8797 (a CX3CR1 inhibitor) prevented the worsening of hyperalgesia and microglial activation in migraine model rats after seizures, while FKN infusion in migraine model rats exacerbated hyperalgesia and microglial activation associated with BDNF-Trkb signalling. Furthermore, in neuron-microglia cocultures, microglial activation and FKN/CX3CR1/BDNF/iba1 expression were increased compared with those in microglial cultures alone. Activating microglia with LPS and FKN increased BDNF synthesis in BV2 microglia.
Conclusions: Our results indicated that epilepsy facilitated migraine through FKN/CX3CR1 axis-mediated microglial activation in the cortex/thalamus/sp5c, which was accompanied by BDNF release. Blocking the FKN/CX3CR1 axis and microglial activation are potential therapeutic strategies for preventing and treating migraine in patients with epilepsy.
Antagonizing the CX3CR1 Receptor Markedly Reduces Development of Cardiac Hypertrophy After Transverse Aortic Constriction in Mice
Left-ventricular hypertrophy, characterized by cardiomyocyte hypertrophy, interstitial cell proliferation, and immune cell infiltration, is a high risk factor for heart failure and death. Chemokines interacting with G protein-coupled chemokine receptors probably play a role in left-ventricular hypertrophy development by promoting recruitment of activated leukocytes and modulating left-ventricular remodeling. Using the minimally invasive model of transverse aortic constriction in mice, we demonstrated that a variety of chemokine and chemokine receptor messenger Ribonucleic Acid are overexpressed in the early and late phase of hypertrophy progression. Among the chemokine receptors, Cx3cr1 and Ccr2 were most strongly overexpressed and were significantly upregulated at 3, 7, and 14 days after transverse aortic constriction. Ligands of CX3CR1 (Cx3cl1) and CCR2 (Ccl2, Ccl7, Ccl12) were significantly overexpressed in the left ventricle at the early stages after mechanical pressure overload. Pharmacological inhibition of CX3CR1 signaling using the antagonist AZD8797 led to a significant reduction of hypertrophy, whereas inhibition of CCR2 with the RS504393 antagonist did not show any effect. Furthermore, AZD8797 treatment reduced the expression of the hypertrophic marker genes Nppa and Nppb as well as the profibrotic genes Tgfb1 and Col1a1 at 14 days after transverse aortic constriction. These findings strongly suggest the involvement of the CX3CR1/CX3CL1 pathway in the pathogenesis of left-ventricular hypertrophy.