CLP290
目录号 : GC35713
A prodrug form of CLP 257
Cas No.:1181083-81-7
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
CLP 290 is a carbamate prodrug form of the K+/Cl- cotransporter 2 (KCC2) activator CLP 257 .1 It increases the mechanical paw withdrawal threshold in a rat model of peripheral nerve injury and reverses morphine-induced hyperalgesia in rats when administered at a dose of 100 mg/kg.1,2 CLP 290 (35 mg/kg) also restores stepping ability, as well as increases body weight support and stride length, in a mouse model of paralysis induced by staggered spinal lesions.3
1.Gagnon, M., Bergeron, M.J., Lavertu, G., et al.Chloride extrusion enhancers as novel therapeutics for neurological diseasesNat. Med.19(11)1524-1528(2013) 2.Ferrini, F., Lorenzo, L.-E., Godin, A.G., et al.Enhancing KCC2 function counteracts morphine-induced hyperalgesiaSci. Rep.7(1)3870(2017) 3.Chen, B., Li, Y., Yu, B., et al.Reactivation of dormant relay pathways in injured spinal cord by KCC2 manipulationsCell174(3)521-535(2018)
| Cas No. | 1181083-81-7 | SDF | |
| Canonical SMILES | O=C(N1CCCC1)OC2=CC(F)=CC=C2/C=C(SC(N3CCCCN3)=N4)/C4=O | ||
| 分子式 | C19H21FN4O3S | 分子量 | 404.46 |
| 溶解度 | DMSO: 50 mg/mL (123.62 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 | 2.4724 mL | 12.3622 mL | 24.7243 mL |
| 5 mM | 0.4945 mL | 2.4724 mL | 4.9449 mL |
| 10 mM | 0.2472 mL | 1.2362 mL | 2.4724 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。
CLP290 promotes the sedative effects of midazolam in neonatal rats in a KCC2-dependent manner: A laboratory study in rats
PLoS One 2021 Mar 12;16(3):e0248113.PMID:33711029DOI:10.1371/journal.pone.0248113.
Immature neurons dominantly express the Na+-K+-2Cl- cotransporter isoform 1 (NKCC1) rather than the K+-Cl- cotransporter isoform 2 (KCC2). The intracellular chloride ion concentration ([Cl-]i) is higher in immature neurons than in mature neurons; therefore, γ-aminobutyric acid type A (GABAA) receptor activation in immature neurons does not cause chloride ion influx and subsequent hyperpolarization. In our previous work, we found that midazolam, benzodiazepine receptor agonist, causes less sedation in neonatal rats compared to adult rats and that NKCC1 blockade by bumetanide enhances the midazolam-induced sedation in neonatal, but not in adult, rats. These results suggest that GABA receptor activation requires the predominance of KCC2 over NKCC1 to exert sedative effects. In this study, we focused on CLP290, a novel KCC2-selective activator, and found that midazolam administration at 20 mg/kg after oral CLP290 intake significantly prolonged the righting reflex latency even in neonatal rats at postnatal day 7. By contrast, CLP290 alone did not exert sedative effects. Immunohistochemistry showed that midazolam combined with CLP290 decreased the number of phosphorylated cAMP response element-binding protein-positive cells in the cerebral cortex, suggesting that CLP290 reverted the inhibitory effect of midazolam. Moreover, the sedative effect of combined CLP290 and midazolam treatment was inhibited by the administration of the KCC2-selective inhibitor VU0463271, suggesting indirectly that the sedation-promoting effect of CLP290 was mediated by KCC2 activation. To our knowledge, this study is the first report showing the sedation-promoting effect of CLP290 in neonates and providing behavioral and histological evidence that CLP290 reverted the sedative effect of GABAergic drugs through the activation of KCC2. Our data suggest that the clinical application of CLP290 may provide a breakthrough in terms of midazolam-resistant sedation.
Enhancing KCC2 function counteracts morphine-induced hyperalgesia
Sci Rep 2017 Jun 20;7(1):3870.PMID:28634406DOI:10.1038/s41598-017-04209-3.
Morphine-induced hyperalgesia (MIH) is a severe adverse effect accompanying repeated morphine treatment, causing a paradoxical decrease in nociceptive threshold. Previous reports associated MIH with a decreased expression of the Cl- extruder KCC2 in the superficial dorsal horn (SDH) of the spinal cord, weakening spinal GABAA/glycine-mediated postsynaptic inhibition. Here, we tested whether the administration of small molecules enhancing KCC2, CLP257 and its pro-drug CLP290, may counteract MIH. MIH was typically expressed within 6-8 days of morphine treatment. Morphine-treated rats exhibited decreased withdrawal threshold to mechanical stimulation and increased vocalizing behavior to subcutaneous injections. Chloride extrusion was impaired in SDH neurons measured as a depolarizing shift in E GABA under Cl- load. Delivering CLP257 to spinal cord slices obtained from morphine-treated rats was sufficient to restore Cl- extrusion capacity in SDH neurons. In vivo co-treatment with morphine and oral CLP290 prevented membrane KCC2 downregulation in SDH neurons. Concurrently, co-treatment with CLP290 significantly mitigated MIH and acute administration of CLP257 in established MIH restored normal nociceptive behavior. Our data indicate that enhancing KCC2 activity is a viable therapeutic approach for counteracting MIH. Chloride extrusion enhancers may represent an effective co-adjuvant therapy to improve morphine analgesia by preventing and reversing MIH.
Traumatic Brain Injury Temporal Proteome Guides KCC2-Targeted Therapy
J Neurotrauma 2019 Nov 15;36(22):3092-3102.PMID:31122143DOI:10.1089/neu.2019.6415.
Advancing therapeutics for traumatic brain injury (TBI) remains a challenge, necessitating testable targets with interventions appropriately timed to intercede on evolving secondary insults. Neuroproteomics provides a global molecular approach to deduce the complex post-translational processes that underlie secondary events after TBI. Yet method advancement has outpaced approaches to interrogate neuroproteomic complexity, in particular when addressing the well-recognized temporal evolution of TBI pathobiology. Presented is a detailed account of the temporal neuroproteomic response to mild-moderate rat controlled cortical impact within perilesioned somatosensory neocortex across the first two weeks after injury. Further, this investigation assessed use of artificial neural network and functional enrichment analyses to discretize the temporal response across some 2047 significantly impacted proteins. Results were efficiently narrowed onto ion transporters with phenotypic relevance to abnormal GABAergic transmission and a delayed decline amenable to intervention under managed care conditions. The prototypical target potassium/chloride co-transporter 2 (KCC2 or SLC12A5) was investigated further with the KCC2-selective modulator CLP290. Guided by post-translational processing revealed one-day after insult to precede KCC2 protein loss a day after, CLP290 was highly effective at restoring up to 70% of lost KCC2 localization, which was significantly correlated with recovery of sham-level function in assessed somatosensory behavioral tasks. The timing of administration was important, with no significant improvement observed if given earlier, one-hour after insult, or later when KCC2 protein decline begins. Results portend importance for a detailed post-translational characterization when devising TBI treatments, and support the therapeutic promise of KCC2-targeted CLP290 intervention for positive functional recovery after brain injury.
Targeting ischemia-induced KCC2 hypofunction rescues refractory neonatal seizures and mitigates epileptogenesis in a mouse model
Sci Signal 2021 Nov 9;14(708):eabg2648.PMID:34752143DOI:10.1126/scisignal.abg2648.
Neonatal seizures pose a clinical challenge in their early detection, acute management, and long-term comorbidities. They are often caused by hypoxic-ischemic encephalopathy and are frequently refractory to the first-line antiseizure medication phenobarbital. One proposed mechanism for phenobarbital inefficacy during neonatal seizures is the reduced abundance and function of the neuron-specific K+/Cl− cotransporter 2 (KCC2), which maintains chloride homeostasis and promotes GABAergic inhibition upon its phosphorylation during postnatal development. Here, we investigated whether this mechanism is causal and whether it can be rescued by KCC2 functional enhancement. In a CD-1 mouse model of refractory ischemic neonatal seizures, treatment with the KCC2 functional enhancer CLP290 rescued phenobarbital efficacy, increased KCC2 abundance, and prevented the development of epileptogenesis, as quantified by video electroencephalogram monitoring. These effects were prevented by knock-in expression of nonphosphorylatable mutants of KCC2 (S940A or T906A and T1007A), indicating that KCC2 phosphorylation regulates both neonatal seizure susceptibility and CLP290-mediated KCC2 functional enhancement. Our findings therefore validate KCC2 as a clinically relevant target for refractory neonatal seizures and provide insights for future drug development.
Therapeutic effects of KCC2 chloride transporter activation on detrusor overactivity in mice with spinal cord injury
Am J Physiol Renal Physiol 2023 Apr 1;324(4):F353-F361.PMID:36656987DOI:10.1152/ajprenal.00271.2022.
This study aimed to clarify whether downregulation of K+-Cl- cotransporter 2 (KCC2) in the sacral parasympathetic nucleus (SPN) of the lumbosacral spinal cord, from which the efferent pathway innervating the bladder originates, causes cellular hyperexcitability and triggers detrusor overactivity (DO) in spinal cord injury (SCI). SCI was produced by Th8-9 spinal cord transection in female C57BL/6 mice. At 4 wk after SCI, CLP290, a KCC2 activator, was administered, and cystometry was performed. Thereafter, neuronal activity with c-fos staining and KCC2 expression in cholinergic preganglionic parasympathetic neurons in the SPN was examined using immunohistochemistry. Firing properties of neurons in the SPN region were evaluated by extracellular recordings in the spinal cord slice preparations. DO evident as nonvoiding contractions was significantly reduced by CLP290 treatment in SCI mice. The number of c-fos-positive cells and coexpression of c-fos in choline acetyltransferase-positive cells were decreased in the SPN region of the SCI CLP290-treated group versus the SCI vehicle-treated group. KCC2 immunoreactivity was present on the cell membrane of SPN neurons and normalized fluorescence intensity of KCC2 in choline acetyltransferase-positive SPN neurons was decreased in the SCI vehicle-treated group versus the spinal intact vehicle-treated group but recovered in the SCI CLP290-treated group. Extracellular recordings showed that CLP290 suppressed the high-frequency firing activity of SPN neurons in SCI mice. These results indicated that SCI-induced DO is associated with downregulation of KCC2 in preganglionic parasympathetic neurons and that activation of KCC2 transporters can reduce DO, increase KCC2 expression in preganglionic parasympathetic neurons, and decrease neuronal firing of SPN neurons in SCI mice.NEW & NOTEWORTHY This study is the first report to suggest that activation of the Cl- transporter K+-Cl- cotransporter 2 may be a therapeutic modality for the treatment of spinal cord injury-induced detrusor overactivity by targeting bladder efferent pathways.