Isorhamnetin is a flavonoid compound extracted from the Chinese herb Hippophae rhamnoides L.. Isorhamnetin suppresses skin cancer through direct inhibition of MEK1 and PI3K.

Isorhamnetin is a plant flavonoid that occurs in fruits and medicinal herbs. Isorhamnetin binds directly to MEK1 in an ATP-noncompetitive manner and to PI3-K in an ATP-competitive manner. In vitro and ex vivo kinase assay data show that Isorhamnetin inhibits the kinase activity of MAP/ERK kinase (MEK) 1 and PI3-K and the inhibition is due to direct binding with Isorhamnetin^[1]. Isorhamnetin inhibits the Akt/mTOR and MEK/ERK signaling pathways, and promotes the activity of the mitochondrial apoptosis signaling pathway. The inhibitory effects of Isorhamnetin on breast cancer cells are determined using the CCK-8 method. Isorhamnetin inhibits the proliferation of numerous breast cancer cells (IC₅₀, ~10 µM), including MCF7, T47D, BT474, BT-549, MDA-MB-231 and MDA-MB-468, whereas less inhibitory activity is observed in the MCF10A normal breast epithelial cell line (IC₅₀, 38 µM)^[2].

Shanghai Jinpan Biotech Co Ltd has not independently confirmed the accuracy of these methods. They are for reference only.

Photographic data shows that Isorhamnetin treatment suppresses tumor development in mice. The average volume of tumors in untreated mice increases over time and reaches a volume of 623 mm³ at 4 weeks post-inoculation; however, at this time, in mice treated with 1 or 5 mg/kg Isorhamnetin, the average tumor volume is only 280 or 198 mm³, respectively. At the end of the study, Isorhamnetin treatment (1 or 5 mg/kg) reduces tumor weight compared with the untreated control group^[1].

Shanghai Jinpan Biotech Co Ltd has not independently confirmed the accuracy of these methods. They are for reference only.

316.26

C₁₆H₁₂O₇

480-19-3

异鼠李素

Room temperature in continental US; may vary elsewhere.

DMSO : 100 mg/mL (316.20 mM; Need ultrasonic)

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。

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: 2.08 mg/mL (6.58 mM); Suspended solution; Need ultrasonic

  此方案可获得 2.08 mg/mL (6.58 mM) 的均匀悬浊液，悬浊液可用于口服和腹腔注射。

  以 1 mL 工作液为例，取 100 μL 20.8 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]. Kim JE, et al. Isorhamnetin suppresses skin cancer through direct inhibition of MEK1 and PI3-K. Cancer Prev Res (Phila). 2011 Apr;4(4):582-91.

  [2]. Hu S, et al. Isorhamnetin inhibits cell proliferation and induces apoptosis in breast cancer via Akt and mitogen activated protein kinase kinase signaling pathways. Mol Med Rep. 2015 Nov;12(5):6745-51.

MCF7, T47D, BT474, BT-549, MDA-MB-231 and MDA-MB-468 breast cancer cell lines, as well as a MCF10A normal breast epithelial cell line (control) are seeded into 96-well plates at a density of 5×10³ cells/well in 100 µL DMEM and placed in cell incubator for 12 h at 37°C in an atmosphere containing 5% CO₂. The cells are then treated with various concentrations of Isorhamnetin (100, 33.3, 11.1, 3.7, 1.2, 0.4 and 0 µM) for 48 h, and cell proliferation rates are determined by adding 10 µL CCK-8 solution prior to incubation at 37°C for 2 h. The absorbance is measured at a wavelength of 450 nm using a SpectraMax 190 Microplate Reader. For each assay, four parallel wells are included, and the half maximal inhibitory concentration (IC₅₀) is measured using the inhibition curve and presented as the mean of three independent experiments^[2].

Shanghai Jinpan Biotech Co Ltd has not independently confirmed the accuracy of these methods. They are for reference only.

Mice^[1]
Female athymic nude mice are injected subcutaneously in the flank with A431 cells (1×10⁶ cells in 50 μL of medium and 50 μL of Matrigel). Cells are allowed to form tumors, and once the tumors reach a size of 40 mm³, the mice are randomly assigned into groups (6 mice/group) and treated with (1 or 5 mg/kg body weight) or without Isorhamnetin in 40% DMSO/PBS buffer, administered intraperitoneally every other day for 28 days. Tumor size is measured every week with calipers, and the tumor volume is calculated. Mice are sacrificed after 28 days of treatment when the control tumors reach approximately 600 mm³. The tumors are harvested, photographed, and weighed. Tumor tissues are used for western blot analysis and immunohistochemical analysis.

Shanghai Jinpan Biotech Co Ltd has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Kim JE, et al. Isorhamnetin suppresses skin cancer through direct inhibition of MEK1 and PI3-K. Cancer Prev Res (Phila). 2011 Apr;4(4):582-91.

  [2]. Hu S, et al. Isorhamnetin inhibits cell proliferation and induces apoptosis in breast cancer via Akt and mitogen activated protein kinase kinase signaling pathways. Mol Med Rep. 2015 Nov;12(5):6745-51.