ZT-1a
目录号 : GC39826
ZT-1a 是一种有效的非 ATP 竞争性的选择性 SPAK 抑制剂。ZT-1a 抑制 SPAK 活性，在 ATP 浓度为 0.01、0.1 和 1 mM 时，IC50 分别为 44.3、35.0、46.7 μM。
Cas No.:212135-62-1
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
ZT-1a is a potent, non-ATP-competitive and selective SPAK inhibitor. ZT-1a inhibits SPAK activity with IC50s of 44.3, 35.0, 46.7 μM at ATP concentrations of 0.01, 0.1 and 1 mM, respectively[1].
[1]. Jinwei Zhang, et al. Modulation of Brain cation-Cl- Cotransport via the SPAK Kinase Inhibitor ZT-1a. Nat Commun. 2020 Jan 7;11(1):78.
| Cas No. | 212135-62-1 | SDF | |
| Canonical SMILES | O=C(NC1=CC(Cl)=C(C(C2=CC=C(Cl)C=C2)C#N)C=C1C)C3=CC(Cl)=CC=C3O | ||
| 分子式 | C22H15Cl3N2O2 | 分子量 | 445.73 |
| 溶解度 | DMSO: 125 mg/mL (280.44 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 | 2.2435 mL | 11.2176 mL | 22.4351 mL |
| 5 mM | 0.4487 mL | 2.2435 mL | 4.487 mL |
| 10 mM | 0.2244 mL | 1.1218 mL | 2.2435 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。
Modulation of brain cation-Cl- cotransport via the SPAK kinase inhibitor ZT-1a
Nat Commun 2020 Jan 7;11(1):78.PMID:31911626DOI:10.1038/s41467-019-13851-6.
The SLC12A cation-Cl- cotransporters (CCC), including NKCC1 and the KCCs, are important determinants of brain ionic homeostasis. SPAK kinase (STK39) is the CCC master regulator, which stimulates NKCC1 ionic influx and inhibits KCC-mediated efflux via phosphorylation at conserved, shared motifs. Upregulation of SPAK-dependent CCC phosphorylation has been implicated in several neurological diseases. Using a scaffold-hybrid strategy, we develop a novel potent and selective SPAK inhibitor, 5-chloro-N-(5-chloro-4-((4-chlorophenyl)(cyano)methyl)-2-methylphenyl)-2-hydroxybenzamide ("ZT-1a"). ZT-1a inhibits NKCC1 and stimulates KCCs by decreasing their SPAK-dependent phosphorylation. Intracerebroventricular delivery of ZT-1a decreases inflammation-induced CCC phosphorylation in the choroid plexus and reduces cerebrospinal fluid (CSF) hypersecretion in a model of post-hemorrhagic hydrocephalus. Systemically administered ZT-1a reduces ischemia-induced CCC phosphorylation, attenuates cerebral edema, protects against brain damage, and improves outcomes in a model of stroke. These results suggest ZT-1a or related compounds may be effective CCC modulators with therapeutic potential for brain disorders associated with impaired ionic homeostasis.
Efficacy of novel SPAK inhibitor ZT-1a derivatives (1c, 1d, 1g & 1h) on improving post-stroke neurological outcome and brain lesion in mice
Neurochem Int 2023 Jan;162:105441.PMID:PMC9839627DOI:10.1016/j.neuint.2022.105441.
SPAK inhibitor ZT-1a was previously shown to be neuroprotective in murine ischemic stroke models. In this study, we further examined the efficacy of four ZT-1a derivatives (ZT-1c, -1d, -1g and -1h) on reducing stroke-induced sensorimotor function impairment and brain lesions. Vehicle control (Veh) or ZT-1 derivatives were administered via osmotic pump to adult C57BL/6J mice during 3-21 h post-stroke. Neurological behavior of these mice was assessed at days 1, 3, 5, and 7 post-stroke and MRI T2WI and DTI analysis was subsequently conducted in ex vivo brains. Veh-treated stroke mice displayed sensorimotor function deficits compared to Sham mice. In contrast, mice receiving ZT-1a derivatives displayed significantly lower neurological deficits at days 3-7 post-stroke (p < 0.05), with ZT-1a, ZT-1c and ZT-1d showing greater impact than ZT-1h and ZT-1g. ZT-1a treatment was the most effective in reducing brain lesion volume on T2WI and in preserving NeuN + neurons (p < 0.01), followed by ZT-1d > -1c > -1g > -1h. The Veh-treated stroke mice displayed white matter tissue injury, reflected by reduced fractional anisotropy (FA) or axial diffusivity (AD) values in external capsule, internal capsule and hippocampus. In contrast, only ZT-1a-as well as ZT-1c-treated stroke mice exhibited significantly higher FA and AD values. These findings demonstrate that post-stroke administration of SPAK inhibitor ZT-1a and its derivatives (ZT-1c and ZT-1d) is effective in protecting gray and white matter tissues in ischemic brains, showing a potential for ischemic stroke therapy development.
NF-κB Signaling-Mediated Activation of WNK-SPAK-NKCC1 Cascade in Worsened Stroke Outcomes of Ang II-Hypertensive Mice
Stroke 2022 May;53(5):1720-1734.PMID:PMC9038703DOI:10.1161/STROKEAHA.121.038351.
Background: Worsened stroke outcomes with hypertension comorbidity are insensitive to blood pressure-lowering therapies. In an experimental stroke model with comorbid hypertension, we investigated causal roles of ang II (angiotensin II)-mediated stimulation of the brain WNK (with no lysine [K] kinases)-SPAK (STE20/SPS1-related proline/alanine-rich kinase)-NKCC1 (Na-K-Cl cotransporter) complex in worsened outcomes. Methods: Saline- or ang II-infused C57BL/6J male mice underwent stroke induced by permanent occlusion of the distal branches of the middle cerebral artery. Mice were randomly assigned to receive either vehicle dimethyl sulfoxide/PBS (2 mL/kg body weight/day, IP), a novel SPAK inhibitor, 5-chloro-N-(5-chloro-4-((4-chlorophenyl)(cyano)methyl)-2-methylphenyl)-2-hydroxybenzamide (ZT-1a' 5 mg/kg per day, IP) or a NF-κB (nuclear factor-κB) inhibitor TAT-NBD (transactivator of transcription-NEMO-binding domain' 20 mg/kg per day, IP). Activation of brain NF-κB and WNK-SPAK-NKCC1 cascade as well as ischemic stroke outcomes were examined. Results: Stroke triggered a 2- to 5-fold increase of WNK (isoforms 1, 2, 4), SPAK/OSR1 (oxidative stress-responsive kinase 1), and NKCC1 protein in the ang II-infused hypertensive mouse brains at 24 hours after stroke, which was associated with increased nuclear translocation of phospho-NF-κB protein in the cortical neurons (a Pearson correlation r of 0.77, P<0.005). The upregulation of WNK-SPAK-NKCC1 cascade proteins resulted from increased NF-κB recruitment on Wnk1, Wnk2, Wnk4, Spak, and Nkcc1 gene promoters and was attenuated by NF-κB inhibitor TAT-NBD. Poststroke administration of SPAK inhibitor ZT-1a significantly reduced WNK-SPAK-NKCC1 complex activation, brain lesion size, and neurological function deficits in the ang II-hypertensive mice without affecting blood pressure and cerebral blood flow. Conclusions: The ang II-induced stimulation of NF-κB transcriptional activity upregulates brain WNK-SPAK-NKCC1 cascade and contributes to worsened ischemic stroke outcomes, illustrating the brain WNK-SPAK-NKCC1 complex as a therapeutic target for stroke with comorbid hypertension.
New drugs on the horizon for cerebral edema: what's in the clinical development pipeline?
Expert Opin Investig Drugs 2020 Oct;29(10):1099-1105.PMID:32815401DOI:10.1080/13543784.2020.1813715.
Introduction: Research has advanced our understanding of the molecular and cellular mechanisms of cerebral edema and has propelled the development of novel antiedema therapeutics. Current evidence supports aberrant neuro-glial ion transport as a central mechanism that underlies pathological fluid accumulation after central nervous system injury. Areas covered: Novel agents in clinical development show potential in altering the natural history and treatment of cerebral edema. Using the PubMed and Google Scholar databases, we review recent advances in our understanding of cerebral edema and describe agents under active investigation, their mechanism, and their application in recent and ongoing clinical trials. Expert opinion: Pharmacotherapies that target molecular mechanisms underlying the compensatory post-injury response of ion channels and transporters that lead to pathological alteration of osmotic gradients are the most promising therapeutic strategies. Repurposing of drugs such as glyburide that inhibit the aberrant upregulation of ion channels such as SUR1-TRPM4, and novel agents, such as ZT-1a, which reestablish physiological regulation of ion channels such as NKCC1/KCC, could be useful adjuvants to prevent and even reverse fluid accumulation in the brain parenchyma.
Role of SPAK-NKCC1 signaling cascade in the choroid plexus blood-CSF barrier damage after stroke
J Neuroinflammation 2022 Apr 12;19(1):91.PMID:35413993DOI:10.1186/s12974-022-02456-4.
Background: The mechanisms underlying dysfunction of choroid plexus (ChP) blood-cerebrospinal fluid (CSF) barrier and lymphocyte invasion in neuroinflammatory responses to stroke are not well understood. In this study, we investigated whether stroke damaged the blood-CSF barrier integrity due to dysregulation of major ChP ion transport system, Na+-K+-Cl- cotransporter 1 (NKCC1), and regulatory Ste20-related proline-alanine-rich kinase (SPAK). Methods: Sham or ischemic stroke was induced in C57Bl/6J mice. Changes on the SPAK-NKCC1 complex and tight junction proteins (TJs) in the ChP were quantified by immunofluorescence staining and immunoblotting. Immune cell infiltration in the ChP was assessed by flow cytometry and immunostaining. Cultured ChP epithelium cells (CPECs) and cortical neurons were used to evaluate H2O2-mediated oxidative stress in stimulating the SPAK-NKCC1 complex and cellular damage. In vivo or in vitro pharmacological blockade of the ChP SPAK-NKCC1 cascade with SPAK inhibitor ZT-1a or NKCC1 inhibitor bumetanide were examined. Results: Ischemic stroke stimulated activation of the CPECs apical membrane SPAK-NKCC1 complex, NF-κB, and MMP9, which was associated with loss of the blood-CSF barrier integrity and increased immune cell infiltration into the ChP. Oxidative stress directly activated the SPAK-NKCC1 pathway and resulted in apoptosis, neurodegeneration, and NKCC1-mediated ion influx. Pharmacological blockade of the SPAK-NKCC1 pathway protected the ChP barrier integrity, attenuated ChP immune cell infiltration or neuronal death. Conclusion: Stroke-induced pathological stimulation of the SPAK-NKCC1 cascade caused CPECs damage and disruption of TJs at the blood-CSF barrier. The ChP SPAK-NKCC1 complex emerged as a therapeutic target for attenuating ChP dysfunction and lymphocyte invasion after stroke.