Indirubin-3′-monoxime is a potent GSK-3β inhibitor, and weakly inhibits 5-Lipoxygenase, with IC₅₀s of 22 nM and 7.8-10 µM, respectively; Indirubin-3′-monoxime also shows inhibitory activities against CDK5/p25 and CDK1/cyclin B, with IC₅₀s of 100 and 180 nM.

Indirubin-3′-monoxime inhibits GSK-3β by competing with ATP, with K_i of 0.85 μM, and K_m of 110 μM. Indirubin-3′-monoxime also inhibits tau phosphorylation by GSK-3β, with an IC₅₀ value of around 100 nM. Indirubin-3′-monoxime completely inhibits the phosphorylation of the AT100 epitope^[1]. Indirubin-3′-monoxime inhibits vascular smooth muscle cell (VSMC) proliferation with IC₅₀ of ∼2 µM. Indirubin-3′-monoxime blunts migration of VSMC stimulated with the PDGF. Indirubin-3′-monoxime interferes with the migratory response in VSMC, and also suppresses the production of pro-migratory LT in monocytes. Moreover, Indirubin-3′-monoxime inhibits 5-lipoxygenase (5-LO) product synthesis in monocytes and neutrophils, with the same potency (IC₅₀=5.0±1.1 and 3.7±1.2 µM, respectively). Indirubin-3′-monoxime is an inhibitor of 5-LO, with IC₅₀ of 7.8-10 µM in cell-free assay^[3].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

Indirubin-3′-monoxime (0.1, 0.2 and 0.4 mg/kg, i.p) dose dependently reverses the cognitive impairment and combats the elevated oxidative stress markers in HFD fed mice. Indirubin-3′-monoxime also dose dependently lowers the serum glucose, TGs, TC and insulin levels, and improves the β-cell functioning in HFD fed mice. Moreover, Indirubin-3′-monoxime treatment significantly decreases HOMA-IR levels compared to HFD group. Indirubin-3′-monoxime (0.4 mg/kg) significantly attenuates the increased EL in the HFD group^[2].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

277.28

C₁₆H₁₁N₃O₂

160807-49-8

Room temperature in continental US; may vary elsewhere.

Powder	-20°C	3 years
	4°C	2 years
In solvent	-80°C	6 months
	-20°C	1 month

In Vitro: 

DMSO : ≥ 37 mg/mL (133.44 mM)

* “≥” means soluble, but saturation unknown.

*

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, 6 months; -20°C, 1 month。-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。

In Vivo:

请根据您的实验动物和给药方式选择适当的溶解方案。以下溶解方案都请先按照 In Vitro 方式配制澄清的储备液，再依次添加助溶剂：

——为保证实验结果的可靠性，澄清的储备液可以根据储存条件，适当保存；体内实验的工作液，建议您现用现配，当天使用； 以下溶剂前显示的百
分比是指该溶剂在您配制终溶液中的体积占比；如在配制过程中出现沉淀、析出现象，可以通过加热和/或超声的方式助溶

• 1.

  请依序添加每种溶剂： 10% DMSO    40% PEG300    5% Tween-80    45% saline

  Solubility: ≥ 2.5 mg/mL (9.02 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (9.02 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 25.0 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀；向上述体系中加入50 μL Tween-80，混合均匀；然后继续加入 450 μL生理盐水定容至 1 mL。

  将 0.9 g 氯化钠，完全溶解于 100 mL ddH₂O 中，得到澄清透明的生理盐水溶液

• [1]. Leclerc S, et al. Indirubins inhibit glycogen synthase kinase-3 beta and CDK5/p25, two protein kinases involved in abnormal tau phosphorylation in Alzheimer’s disease. A property common to most cyclin-dependent kinase inhibitors? J Biol Chem. 2001 Jan 5;276(1):251-60.

  [2]. Sharma S, et al. Neuroprotective role of Indirubin-3′-monoxime, a GSKβ inhibitor in high fat diet induced cognitive impairment in mice. Biochem Biophys Res Commun. 2014 Oct 3;452(4):1009-15.

  [3]. Blazevic T, et al. Indirubin-3′-monoxime exerts a dual mode of inhibition towards leukotriene-mediated vascular smooth muscle cell migration. Cardiovasc Res. 2014 Mar 1;101(3):522-32.

  [4]. Cao Z, et al. Indirubin Derivatives as Dual Inhibitors Targeting Cyclin-Dependent Kinase and Histone Deacetylase for Treating Cancer [published online ahead of print, 2021 Oct 8]. J Med Chem. 2021;10.1021/acs.jmedchem.1c01311.

GSK-3β is expressed in and purified from insect Sf9 cells. It is assayed, following a 1/100 dilution in 1 mg/mL BSA, 10 mM DTT, with 5 μL of 40 μM GS-1 peptide as a substrate, in buffer A, in the presence of 15 μM[γ-³²P]ATP (3000 Ci/mmol; 1 mCi/mL) in a final volume of 30 μL. After 30-min incubation at 30°C, 25-μL aliquots of supernatant are spotted onto 2.5×3-cm pieces of Whatman P81 phosphocellulose paper, and, 20 s later, the filters are washed five times (for at least 5 min each time) in a solution of 10 mL of phosphoric acid/liter of water. The wet filters are counted in the presence of 1 mL of ACS scintillation fluid^[1].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

Cytotoxicity of Indirubin-3′-monoxime in monocytes is analysed by MTT assay in a 96-well format using a multi-well scanning spectrophotometer. Neutrophils (5×10⁶ cells/mL) or monocytes (2×10⁶ cells/mL) are incubated for 30 min with Indirubin-3′-monoxime, and the viability of the cells is analysed by MTT assay. Compared with vehicle (0.3% DMSO), no significant acute cytotoxicity is observed (neutrophils: 103.9±4.4%; monocytes: 129.4±5.4%; n=3, each)^[3].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

Male mice (5-6 weeks old) are randomLy assigned into five groups (n=10). Group 1: receive normal pellet diet (NPD); Group 2: receive a HFD; Group 3-5 receive HFD for 8 weeks followed by Indirubin-3′-monoxime treatment (0.1, 0.2 and 0.4 mg/kg i.p, respectively) once daily for 1 week. Indirubin-3′-monoxime is dissolved in (2.5% v/v) DMSO in saline. The mice in NPD and HFD groups receive an equivalent volume of vehicle (2.5% v/v DMSO in saline). Doses of Indirubin-3′-monoxime are selected. Mice are kept under standard husbandry conditions (22±1°C and 60% humidity) and maintained on a 12/12-h light/dark schedule with free access to food and water for 8 weeks. Body weight is recorded weekly throughout the experimental period^[2].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Leclerc S, et al. Indirubins inhibit glycogen synthase kinase-3 beta and CDK5/p25, two protein kinases involved in abnormal tau phosphorylation in Alzheimer’s disease. A property common to most cyclin-dependent kinase inhibitors? J Biol Chem. 2001 Jan 5;276(1):251-60.

  [2]. Sharma S, et al. Neuroprotective role of Indirubin-3′-monoxime, a GSKβ inhibitor in high fat diet induced cognitive impairment in mice. Biochem Biophys Res Commun. 2014 Oct 3;452(4):1009-15.

  [3]. Blazevic T, et al. Indirubin-3′-monoxime exerts a dual mode of inhibition towards leukotriene-mediated vascular smooth muscle cell migration. Cardiovasc Res. 2014 Mar 1;101(3):522-32.

  [4]. Cao Z, et al. Indirubin Derivatives as Dual Inhibitors Targeting Cyclin-Dependent Kinase and Histone Deacetylase for Treating Cancer [published online ahead of print, 2021 Oct 8]. J Med Chem. 2021;10.1021/acs.jmedchem.1c01311.