Belinostat (PXD101; PX105684) is a potent HDAC inhibitor with an IC₅₀ of 27 nM in HeLa cell extracts.

Belinostat (PXD101) induces a concentration-dependent (0.2-5 μM) increase in acetylation of histone H4 in tumor cell lines. Belinostat is cytotoxic in vitro in a number of tumor cell lines with IC₅₀s in the range 0.2-3.4 μM as determined by a clonogenic assay and induces apoptosis. Belinostat inhibits the growth of a number of human tumor cell lines in vitro with IC₅₀s determined by a clonogenic assay in the range 0.2-3.4 μM^[1]. Belinostat (PXD101) is a potent histone deacetylase (HDAC) inhibitor, potently inhibits the enzymatic activity of purified recombinant HDAC6 (IC₅₀ of 82 nM)^[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.

Treatment of nude mice bearing human ovarian and colon tumor xenografts with Belinostat (10-40 mg/kg/day i.p.) daily for 7 days causes a significant dose-dependent growth delay with no obvious signs of toxicity to the mice. Growth delay is also observed for xenografts of cisplatin-resistant ovarian tumor cells. A marked increase in acetylation of H4 is detected in blood and tumor of mice 3 h after treatment with Belinostat (PXD101). The inhibition of growth of human tumor xenografts in mice, with no apparent toxicity^[1]. Belinostat (PXD101) displays single-agent antitumor activity on human A2780 ovarian cancer s.c. xenografts which is enhanced via combination therapy with Carboplatin^[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.

318.35

C₁₅H₁₄N₂O₄S

866323-14-0

贝利司他

Room temperature in continental US; may vary elsewhere.

DMSO : 100 mg/mL (314.12 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: ≥ 2.5 mg/mL (7.85 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (7.85 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

• 2.

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

  Solubility: ≥ 2.5 mg/mL (7.85 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (7.85 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 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
• 3.

  请依序添加每种溶剂： 10% DMSO    90% corn oil

  Solubility: ≥ 2.5 mg/mL (7.85 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (7.85 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]. Plumb JA, et al. Pharmacodynamic response and inhibition of growth of human tumor xenografts by the novel histonedeacetylase inhibitor PXD101. Mol Cancer Ther. 2003 Aug;2(8):721-8.

  [2]. Qian X, et al. Activity of PXD101, a histone deacetylase inhibitor, in preclinical ovarian cancer studies. Mol Cancer Ther. 2006 Aug;5(8):2086-95.

  [3]. Chia S, et al. Phenotype-driven precision oncology as a guide for clinical decisions one patient at a time. Nat Commun. 2017 Sep 5;8(1):435.

For activity assays, the reaction is carried out in a total volume of 150 μL of buffer [60 mM Tris (pH 7.4) containing 30% glycerol] containing 2 μL of cell extract and, where used, 2 μL of Belinostat. The reaction is started by the addition of 2 μL of [³H]labeled substrate (acetylated histone H4 peptide corresponding to the 20 NH₂-terminal residues). Samples are incubated at 37°C for 45 min, and the reaction stopped by the addition of HCl and acetic acid (0.72 and 0.12 M final concentrations, respectively). Released [³H]acetate is extracted into 750 μL of ethyl acetate, and samples are centrifuged at 12,000× g for 5 min. The upper phase (600 μL) is transferred to 3 mL of scintillation fluid and counted^[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.

The human ovarian cell line A2780 and Cisplatin (A2780/cp70) and Doxorubicin (2780AD) resistant derivatives are grown in RPMI 1640 supplemented with glutamine (2 mM) and FCS (10%). The human colon (HCT116 and HT29), melanoma (HS852), prostate (PC3), lung (CALU-3), and breast (MCF7) cell lines are grown in RPMI 1640 and the rest in DMEM supplemented as above. The human non-small cell lung cancer cell line WIL is grown in DMEM supplemented as above. Drug sensitivity is determined by a clonogenic assay. Briefly, cells are plated in 5 mL of medium at a density of 8×10⁴ cells/25 cm² flask and allowed to attach and grow for 48 h. Cells are exposed to Belinostat (five concentrations from 0.016 to 10 μM) for 24 h. The medium is removed, and 1 mL of trypsin/EDTA is added to each flask. Once the cells have detached, 1 mL of medium is added, the cells are resuspended, and those from the control untreated flask are counted. Cells are diluted and plated into 6-cm Petri dishes (three per flask) at a density of 500-2000 cells/dish depending on the cell line. Cells from the drug-treated flasks are diluted and plated as for the control flasks. Dishes are incubated for 10-15 days at 37°C. Cells are washed with PBS, fixed in methanol, and stained with crystal violet, and colonies that contained ≥50 cells counted. Sensitivity is expressed as the IC₅₀ (mean±SE of three experiments) defined as the concentration of drug required to reduce the number of colonies to 50% of that of the control untreated cells^[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]
For the human tumor xenograft studies, monolayer cultures are harvested with trypsin/EDTA (0.25%/1 mM in PBS) and resuspended in PBS. About 10⁷ cells are injected s.c. into the right flank of athymic nude mice (CD1 nu/nu mice). After 10-15 days when the mean tumor diameter is ≥0.5 cm, animals are randomized into groups of six for experiments. Belinostat is dissolved in DMSO and then diluted in water to give a final concentration of DMSO of 10% and is administered i.p. at the times specified. This formulation gives sufficient solubility for doses of ≤ 40 mg/kg. Mice are weighed daily, and tumor volumes are estimated by caliper measurements assuming spherical geometry (volume=d³×π/6).

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]. Plumb JA, et al. Pharmacodynamic response and inhibition of growth of human tumor xenografts by the novel histonedeacetylase inhibitor PXD101. Mol Cancer Ther. 2003 Aug;2(8):721-8.

  [2]. Qian X, et al. Activity of PXD101, a histone deacetylase inhibitor, in preclinical ovarian cancer studies. Mol Cancer Ther. 2006 Aug;5(8):2086-95.

  [3]. Chia S, et al. Phenotype-driven precision oncology as a guide for clinical decisions one patient at a time. Nat Commun. 2017 Sep 5;8(1):435.