TAK-632 is a potent pan-RAF inhibitor with IC₅₀ of 1.4, 2.4 and 8.3 nM for CRAF, BRAF^V600E, BRAF^WT, respectively.

TAK-632 inhibits PDGFRβ, FGFR3, GSK3β, CDK2, P38α, PDGFRα, TIE2, and CDK1 with a range of IC₅₀ values from 120-790 nM. CHK1, IKKβ, and MEK1 are inhibited over an IC₅₀ range of 1400-1700 nM. With 1 h of preincubation time, TAK-632 inhibits BRAF and CRAF in an ATP competitive manner (at low ATP concentrations BRAF IC₅₀: 15 nM; CRAF: 8.1 nM). The respective biochemical activity of TAK-632 against BRAF and CRAF reduces to IC₅₀ values of 58 nM and 62 nM at high ATP concentrations.TAK-632 demonstrates strong inhibition of pMEK and pERK in HMVII cells with IC₅₀ values of 49 nM and 50 nM, respectively^[1]. TAK-632 shows strong antiproliferative effects both in A375 and SK-MEL-2 cells (GI₅₀ of 40-190 nM in A375 cells and GI₅₀ of 190-250 nM in SK-MEL-2 cells)^[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.

TAK-632 demonstrates dramatically improved solubility (740 μg/mL) in pH 6.8 phosphate buffer and exhibits significant oral absorption (at a dose of 25 mg/kg, AUC, 32.47 μg h/mL; F, 51.7%) in rats. In a dog PK study, 10 mg/kg administration of TAK-632 also shows superior oral bioavailability (F: 108%).Oral single administration of TAK-632 inhibits pERK in tumors at 8 h after its administration over a dose range of 1.9-24.1 mg/kg. In particular, 9.7-24.1 mg/kg dosing with TAK-632 strongly inhibits pERK levels to 11% of the control. TAK-632 exhibits dose-dependent antitumor efficacy without severe body weight reduction over a dose range of 3.9-24.1 mg/kg. Significant tumor regression is observed at 9.7 mg/kg and 24.1 mg/kg (T/C=−2.1% and −12.1%, respectively)^[1]. TAK-632 exhibits potent antitumor efficacy when orally administered at 60 mg/kg once daily (T/C=37%, P<0.001) or at 120 mg/kg once daily (T/C=29%, P<0.001) for 21 days without severe toxicity in NRAS-mutant melanoma using a SK-MEL-2 xenograft model^[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.

554.52

C₂₇H₁₈F₄N₄O₃S

1228591-30-7

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 : 100 mg/mL (180.34 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。

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 (4.51 mM); Suspended solution; Need ultrasonic

  此方案可获得 2.5 mg/mL (4.51 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 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

• 2.

  请依序添加每种溶剂： 10% DMSO    90% corn oil

  Solubility: ≥ 2.5 mg/mL (4.51 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (4.51 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 25.0 mg/mL 的澄清 DMSO 储备液加到 900 μL玉米油中，混合均匀。

• [1]. Okaniwa M, et al. Discovery of a selective kinase inhibitor (TAK-632) targeting pan-RAF inhibition: design, synthesis, and biological evaluation of C-7-substituted 1,3-benzothiazole derivatives. J Med Chem. 2013 Aug 22;56(16):6478-94.

  [2]. Nakamura A, et al. Antitumor activity of the selective pan-RAF inhibitor TAK-632 in BRAF inhibitor-resistant melanoma. Cancer Res. 2013 Oct 11.

Immunoprecipitated BRAF or CRAF is incubated with recombinant inactive MEK (K97R) at 30°C for 30 minutes in kinase reaction buffer containing ATP/Mg²⁺. RAS/RAF wild-type (A431, CsFb, and HeLa), KRAS-mutant (A549, HCT-116, and MIA PaCa-2), and NRAS-mutant melanoma (GAK, HMV-II, and SK-MEL-2) cells are treated with TAK-632 (0, 0.32, 1.6, 8, 40, 200, 1000 and 5000 nM) at the indicated concentrations for 2 hours. Cell lysates are analyzed by Western blot analysis^[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.

Cell viability is assessed (3 replicates) using the Sulforhodamine B assay or by the CellTiter-Glo luminescent cell viability assay. The concentrations of TAK-632 that produced 50% growth inhibition (GI₅₀) are calculated using PCP software. The combination index (CI) is calculated using CalcuSyn software. To investigate the antiproliferative activity of TAK-632, we performed proliferation assays in various cell lines harboring mutated BRAF, NRAS, or KRAS. HMV-II, SK-MEL-2, or A375 cells are cotreated with TAK-632 and TAK-733 at the indicated concentrations for 72 hours. Cell viability is measured. The CI value at EC₅₀ is calculated. A375 cells stably expressing NRAS^Q61K or ΔN-BRAF are cotreated with TAK-632 and TAK-733 at the indicated concentrations for 72 hours. Cell viability is measured. The CI value at EC₅₀ is calculated^[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.

Mice^[2]
The xenograft-implanted nude mice are used. Mice bearing SK-MEL-2 xenografts are treated once daily for 21 consecutive days with vehicle or TAK-632 at the indicated concentrations (10 mice per each treatment group). Day 0 indicates the beginning of treatment. Tumors are measured twice a week. Mice bearing SK-MEL-2 xenografts are treated once daily (QD) for 3 days with vehicle, TAK-632 at 60 mg/kg (60 mpk), or TAK-632 at 120 mg/kg (120 mpk). Tumor xenografts are obtained at indicated time points after the final treatment and analyzed by Western blot analysis. Individual blots with dividing lines are combined from a single electrophoresis gel. Bars represent densitometric analysis of phospho-ERK, normalized to vehicle-treated control (mean±SD).

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]. Okaniwa M, et al. Discovery of a selective kinase inhibitor (TAK-632) targeting pan-RAF inhibition: design, synthesis, and biological evaluation of C-7-substituted 1,3-benzothiazole derivatives. J Med Chem. 2013 Aug 22;56(16):6478-94.

  [2]. Nakamura A, et al. Antitumor activity of the selective pan-RAF inhibitor TAK-632 in BRAF inhibitor-resistant melanoma. Cancer Res. 2013 Oct 11.