Ensartinib dihydrochloride (X-396 dihydrochloride) is a potent and dual ALK/MET inhibitor with IC₅₀s of <0.4 nM and 0.74 nM, respectively.

IC50: <0.4 nM (ALK), 0.74 nM (MET)^[1]

Ensartinib (X-396) dihydrochloride is a potent and dual ALK/MET inhibitor with IC₅₀s of <0.4 nM and 0.74 nM, respectively. Ensartinib dihydrochloride is potent in H3122 lung cancer cells harboring EML4-ALK E13;A20 (IC₅₀: 15 nM). Ensartinib dihydrochloride is also potent in H2228 lung cancer cells harboring EML4-ALK E6a/b; A20 (IC₅₀: 45 nM). Furthermore, X-376 is potent in SUDHL-1 lymphoma cells harboring NPM-ALK (IC₅₀: 9 nM)^[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.

Ensartinib (X-396) dihydrochloride shows substantial bioavailability and moderate half-lives in vivo. Nude mice harboring H3122 xenografts are treated with Ensartinib dihydrochloride at 25 mg/kg bid. Ensartinib dihydrochloride significantly delays the growth of tumors compared to vehicle alone. In the xenograft experiments, Ensartinib dihydrochloride appears well-tolerated in vivo. Mouse weight is unaffected by Ensartinib dihydrochloride treatment. Drug-treated mice appear healthy and do not display any signs of compound related toxicity. To further assess potential side effects of Ensartinib dihydrochloride, additional systemic toxicity and toxico-kinetic studies are performed in Sprague Dawley (SD) rats. Following 10 days of repeated oral administration of Ensartinib dihydrochloride at 20, 40, 80 mg/kg in SD rats, all animals survive to study termination. The no significant toxicity (NST) levels are determined to be 80 mg/kg for Ensartinib dihydrochloride. At NST levels, Ensartinib dihydrochloride achieves an AUC of 66 μM×hr and a C_max of 7.19 μM^[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.

634.36

C₂₆H₂₉Cl₄FN₆O₃

2137030-98-7

Room temperature in continental US; may vary elsewhere.

4°C, sealed storage, away from moisture

*In solvent : -80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture)

DMSO : 35.71 mg/mL (56.29 mM; Need ultrasonic)

H₂O : 33.33 mg/mL (52.54 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 (sealed storage, away from moisture)。-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： PBS

  Solubility: 14.29 mg/mL (22.53 mM); Clear solution; Need ultrasonic
• 2.

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

  Solubility: ≥ 2.08 mg/mL (3.28 mM); Clear solution

  此方案可获得 ≥ 2.08 mg/mL (3.28 mM，饱和度未知) 的澄清溶液。

  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

• 3.

  请依序添加每种溶剂： 10% DMSO    90% (20% SBE-β-CD in saline)

  Solubility: ≥ 2.08 mg/mL (3.28 mM); Clear solution

  此方案可获得 ≥ 2.08 mg/mL (3.28 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 20.8 mg/mL 的澄清 DMSO 储备液加到 900 μL 20% 的 SBE-β-CD 生理盐水水溶液中，混合均匀。

  将 2 g 磺丁基醚 β-环糊精加入 5 mL 生理盐水中，再用生理盐水定容至 10 mL，完全溶解，澄清透明
• 4.

  请依序添加每种溶剂： 10% DMSO    90% corn oil

  Solubility: ≥ 2.08 mg/mL (3.28 mM); Clear solution

  此方案可获得 ≥ 2.08 mg/mL (3.28 mM，饱和度未知) 的澄清溶液，此方案不适用于实验周期在半个月以上的实验。

  以 1 mL 工作液为例，取 100 μL 20.8 mg/mL 的澄清 DMSO 储备液加到 900 μL玉米油中，混合均匀。

• [1]. Lovly CM, et al. Insights into ALK-driven cancers revealed through development of novel ALK tyrosine kinaseinhibitors. Cancer Res. 2011 Jul 15;71(14):4920-31.

For viability experiments, cells are seeded in 96-well plates at 25%-33% confluency and exposed to drugs. The human lung adenocarcinoma cell lines H3122 and H2228 are treated with Ensartinib (10, 30, 100, 300 and 1000 nM). SUDHL-1 lymphoma cells are treated with Ensartinib (5, 10, 30, 100 and 300 nM). SY5Y neuroblastoma cells are treated with Ensartinib (30, 100, 300 and 1000 nM). At 72 hours post Ensartinib addition, Cell Titer Blue Reagent is added and fluorescence is measured on a Spectramax spectrophotometer. All experimental points are set up in hextuplicate replicates and are performed at least two independent times. IC₅₀s are calculated using GraphPad Prism version 5 for Windows. The curves are fit using a nonlinear regression model with a log (inhibitor) vs. response formula^[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.

Mice^[1]
Nude mice (nu/nu) are injected with H3122 cells. Once tumors reach an average volume of 450 mm³, a total of 27 athymic mice harboring H3122 tumors are randomized and dosed via oral gavage with 25 mg/kg Ensartinib or the control vehicle. Two, five, and fifteen hours after the single treatment (3 tumors/timepoint/group), mice are sacrificed and serum is collected for assessment of drug concentration using an LC-MS based bioanalytical method^[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.

• [1]. Lovly CM, et al. Insights into ALK-driven cancers revealed through development of novel ALK tyrosine kinaseinhibitors. Cancer Res. 2011 Jul 15;71(14):4920-31.