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CPYPP Sale

目录号 : GC50065

An inhibitor of DOCK2, DOCK5, and DOCK180

CPYPP Chemical Structure

Cas No.:310460-39-0

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10 mg
¥1,350.00
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50 mg
¥4,410.00
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Sample solution is provided at 25 µL, 10mM.

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实验参考方法

Cell experiment [1]:

Cell lines

Spleen cells of BALB/c mice

Preparation Method

Spleen cells of BALB/c mice (1 × 107/ml) were incubated in RPMI1640 medium supplemented with 0.5% BSA or 10% FCS in the presence or absence of 100 µM CPYPP. 1 or 5 hr of incubation at 37°C.

Reaction Conditions

100µM for 1 hour or 5 hours

Applications

CPYPP was nontoxic when applied to spleen cells at 100 µM .

Animal experiment [2]:

Animal models

male C57BL/6J mice

Preparation Method

Mice were randomized into four groups (n = 6): the saline + vehicle group, LPS + vehicle group, saline + CPYPP group, and LPS + CPYPP group. ALI was induced by intraperitoneal (i.p.) injection of LPS (10 mg/kg) body weight; The mice in the LPS + vehicle and LPS + CPYPP groups received CPYPP (250 mg/kg) or an equivalent volume of vehicle via i.p. injection 10 min after LPS administration. The mice in the saline + vehicle and saline + CPYPP groups received CPYPP (250 mg/kg) or an equivalent volume of vehicle via i.p. injection 10 min after saline administration.

Dosage form

Intraperitoneal injection, 250 mg/kg

Applications

CPYPP-treated mice had decreased lung injury scores compared with vehicle-treated mice. Furthermore, CPYPP-treated mice exhibited significantly decreased MPO activity and W/D ratios, which indicated decreased pulmonary edema.

References:

[1]: Nishikimi?A, Uruno?T, Duan?X, Cao?Q, Okamura?Y, Saitoh?T, et al.?Blockade of inflammatory responses by a small-molecule inhibitor of the Rac activator DOCK2. Chem Biol?2012;19:488-97.
[2]: Xu, X., Su, Y., Wu, K., Pan, F. & Wang, A. DOCK2 contributes to endotoxemia-induced acute lung injury in mice by activating proinflammatory macrophages. Biochem. Pharmacol. 181, 114399 (2021).

产品描述

CPYPP bound to DOCK2 DHR-2 domain (DOCK2DHR-2) in a reversible manner and inhibited its catalytic activity[1].

CPYPP inhibited the guanine nucleotide exchange factor (GEF) activity of DOCK2DHR-2 for Rac1 in a dose-dependent manner, with a half-maximal inhibitory concentration (IC50) of 22.8 ± 2.4 μM. This inhibitory activity was independent on length of preincubation time, ranging from 2 min to 30 min. In addition, CPYPP was nontoxic when applied to spleen cells or thymoma cells (BW5147α-β-) at 100 μM for 3 hr or 3 days, respectively [1]. Pre-treatment of human neutrophils with CPYPP, a small molecule inhibitor of the DHR-2 domain of DOCK proteins and thus of the Rac GEF activity of human DOCK2 and DOCK5, significantly impaired neutrophil chemotaxis and ROS production [2].

CPYPP effectively decreased the secretion and gene expression of TNF-α, IL-1β and IL-6 in the lungs, suggested effects of DOCK2 on endotoxemia-induced inflammatory responses in mice. CPYPP remarkably inhibited the infiltration of total cells, macrophages and neutrophils into the bronchoalveolar lavage fluid [3]. A reseach found out that a joint usage of both CPYPP and C25-140 revealed a better consequence compared to monotherapy against hepatic I/R injury [4].

References:
[1]. Nishikimi?A, Uruno?T, Duan?X, Cao?Q, Okamura?Y, Saitoh?T, et al.?Blockade of inflammatory responses by a small-molecule inhibitor of the Rac activator DOCK2. Chem Biol?2012;19:488-97.
[2]. Moens, L., Gouwy, M., Bosch, B. et al. 2019. Human DOCK2 deficiency: report of a novel mutation and evidence for neutrophil dysfunction. J. Clin. Immunol. 39:298.
[3]. Xu, X., Su, Y., Wu, K., Pan, F. & Wang, A. DOCK2 contributes to endotoxemia-induced acute lung injury in mice by activating proinflammatory macrophages. Biochem. Pharmacol. 181, 114399 (2021).
[4]. Zuotian Huang, Junliang Pua, Yunhai Luo, et al. FAM49B, restrained by miR-22, relieved hepatic ischemia/reperfusion injury by inhibiting TRAF6/IKK signaling pathway in a Rac1-dependent manner. Molecular Immunology.143, 2022, 135-146.

CPYPP 以可逆方式与 DOCK2 DHR-2 结构域 (DOCK2DHR-2) 结合并抑制其催化活性[1]

CPYPP 以剂量依赖性方式抑制 DOCK2DHR-2 鸟嘌呤核苷酸交换因子 (GEF) 对 Rac1 的活性,半数最大抑制浓度 (IC50) 为 22.8 ± 2.4 μM .这种抑制活性与预孵育时间的长度无关,范围从 2 分钟到 30 分钟。此外,CPYPP 在以 100 μM 分别作用于脾细胞或胸腺瘤细胞 (BW5147α-β-) 3 小时或 3 天时是无毒的[1]。用 CPYPP 预处理人中性粒细胞,CPYPP 是一种小分子抑制剂,可抑制 DOCK 蛋白的 DHR-2 结构域,从而抑制人 DOCK2 和 DOCK5 的 Rac GEF 活性,显着损害中性粒细胞趋化性和 ROS 产生[2].

CPYPP 有效降低肺部 TNF-α、IL-1β 和 IL-6 的分泌和基因表达,表明 DOCK2 对小鼠内毒素血症诱导的炎症反应有影响。 CPYPP显着抑制总细胞、巨噬细胞和中性粒细胞向支气管肺泡灌洗液中的浸润[3]。一项研究发现,与单一疗法相比,CPYPP 和 C25-140 联合使用对肝 I/R 损伤具有更好的疗效[4]

Chemical Properties

Cas No. 310460-39-0 SDF
Canonical SMILES O=C(N(C3=CC=CC=C3)NC2=O)/C2=C\C=C\C1=C(Cl)C=CC=C1
分子式 C18H13ClN2O2 分子量 324.76
溶解度 DMSO: 50 mM 储存条件 Store at -20°C
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1 mg 5 mg 10 mg
1 mM 3.0792 mL 15.396 mL 30.792 mL
5 mM 0.6158 mL 3.0792 mL 6.1584 mL
10 mM 0.3079 mL 1.5396 mL 3.0792 mL
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Research Update

DOCK2 is involved in the host genetics and biology of severe COVID-19

Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge1-5. Here we conducted a genome-wide association study (GWAS) involving 2,393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3,289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target.

DOCK2 contributes to endotoxemia-induced acute lung injury in mice by activating proinflammatory macrophages

Dedicator of cytokinesis 2 (DOCK2), an atypical Rac activator, has important anti-inflammatory properties in blepharitis, enteric bacterial infection and colitis. However, the roles of DOCK2 in macrophage activation and acute lung injury (ALI) are still poorly elucidated. In vitro studies demonstrated that DOCK2 was essential for the nucleotide-sensing Toll-like receptor (TLR) 4-mediated inflammatory response in macrophages. We also confirmed that exposure of macrophages to LPS induced Rac activation through a TLR4-independent, DOCK2-dependent mechanism. Phosphorylation of I百B kinase (IKK) 汕 and nuclear translocation of transcription factor nuclear factor kappa B (NF-百B) were impaired in Ad-shDOCK2-expressing macrophages, resulting in a decreased inflammatory response. Similar results were obtained when EHop-016 (a Rac inhibitor) was used to treat uninfected macrophages. In summary, these data indicate that the DOCK2-Rac signaling pathway acts in parallel with TLR4 engagement to control IKK汕 activation for inflammatory cytokine release. Next, we investigated whether pharmacological inhibition of DOCK2 protects against endotoxemia-induced lung injury in mice. Treatment with 4-[3'-(2∪-chlorophenyl)-2'-propen-1'-ylidene]-1-phenyl-3,5-pyrazolidinedione (CPYPP), a small-molecule inhibitor of DOCK2, reduced the severity of lung injury, as indicated by decreases in the lung injury score and myeloperoxidase (MPO) activity. Moreover, CPYPP attenuated LPS-induced proinflammatory cytokine release in mice. Our studies suggest that inhibition of DOCK2 may suppress LPS-induced macrophage activation and that DOCK2 may be a novel target for treating endotoxemia-related ALI.

The Molecular Motor KIF21B Mediates Synaptic Plasticity and Fear Extinction by Terminating Rac1 Activation

Fear extinction is a component of cognitive flexibility that is relevant for important psychiatric diseases, but its molecular mechanism is still largely elusive. We established mice lacking the kinesin-4 motor KIF21B as a model for fear extinction defects. Postsynaptic NMDAR-dependent long-term depression (LTD) is specifically impaired in knockouts. NMDAR-mediated LTD-causing stimuli induce dynamic association of KIF21B with the Rac1GEF subunit engulfment and cell motility protein 1 (ELMO1), leading to ELMO1 translocation out of dendritic spines and its sequestration in endosomes. This process may essentially terminate transient activation of Rac1, shrink spines, facilitate AMPAR endocytosis, and reduce postsynaptic strength, thereby forming a mechanistic link to LTD expression. Antagonizing ELMO1/Dock Rac1GEF activity by the administration of 4-[3'-(2∪-chlorophenyl)-2'-propen-1'-ylidene]-1-phenyl-3,5-pyrazolidinedione (CPYPP) significantly reverses the knockout phenotype. Therefore, we propose that KIF21B-mediated Rac1 inactivation is a key molecular event in NMDAR-dependent LTD expression underlying cognitive flexibility in fear extinction.

Allosteric inhibition of the guanine nucleotide exchange factor DOCK5 by a small molecule

Rac small GTPases and their GEFs of the DOCK family are pivotal checkpoints in development, autoimmunity and bone homeostasis, and their abnormal regulation is associated to diverse pathologies. Small molecules that inhibit their activities are therefore needed to investigate their functions. Here, we characterized the mechanism of inhibition of human DOCK5 by C21, a small molecule that inhibits mouse Dock5 in cells and blocks bone degradation in mice models of osteoporosis. We showed that the catalytic DHR2 domain of DOCK5 has a high basal GEF activity in the absence of membranes which is not regulated by a simple feedback loop. C21 blocks this activity in a non-competitive manner and is specific for DOCK5. In contrast, another Dock inhibitor, CPYPP, inhibits both DOCK5 and an unrelated GEF, Trio. To gain insight into structural features of the inhibitory mechanism of C21, we used SAXS analysis of DOCK5DHR2 and crystallographic analysis of unbound Rac1-GDP. Together, these data suggest that C21 takes advantage of intramolecular dynamics of DOCK5 and Rac1 to remodel the complex into an unproductive conformation. Based on this allosteric mechanism, we propose that diversion of intramolecular dynamics is a potent mechanism for the inhibition of multidomain regulators of small GTPases.

Blockade of inflammatory responses by a small-molecule inhibitor of the Rac activator DOCK2

Tissue infiltration of activated lymphocytes is a hallmark of transplant rejection and organ-specific autoimmune diseases. Migration and activation of lymphocytes depend on DOCK2, an atypical Rac activator predominantly expressed in hematopoietic cells. Although DOCK2 does not contain Dbl homology domain typically found in guanine nucleotide exchange factors, DOCK2 mediates the GTP-GDP exchange reaction for Rac through its DHR-2 domain. Here, we have identified 4-[3'-(2∪-chlorophenyl)-2'-propen-1'-ylidene]-1-phenyl-3,5-pyrazolidinedione (CPYPP) as a small-molecule inhibitor of DOCK2. CPYPP bound to DOCK2 DHR-2 domain in a reversible manner and inhibited its catalytic activity in vitro. When lymphocytes were treated with CPYPP, both chemokine receptor- and antigen receptor-mediated Rac activation were blocked, resulting in marked reduction of chemotactic response and T cell activation. These results provide a rational of and a chemical scaffold for development of the DOCK2-targeting immunosuppressant.