Tetrahydrouridine dihydrate (THU dihydrate) is potent inhibitor of cytidine deaminase (CDA), which competitively blocks the enzyme’s active site more effectively than intrinsic cytidine.

cytidine deaminase (CDA)^[1]

Tetrahydrouridine (THU) is a specific inhibitor of cytidine deaminase (CDA) which can suppress deamination in the catabolism of cytotoxic deoxycytidine analogues like ara-C and Gemcitabine. To test how Tetrahydrouridine affects the Gemcitabine-mediated anti-neoplastic effect on pancreatic and lung carcinoma cells, a combination therapy is performed. As expected, high CDA expression in BxPC-3 and H441 results in improved Gemcitabine sensitivity after a 100 µM Tetrahydrouridine treatment. The sensitivity of BxPC-3 and H441 cell lines increases by as much as approximately 2.1 and 4.4 fold respectively. On the other hand, MIAPaCa-2 and H1299 cells unexpectedly become more sensitive to Gemcitabine with low CDA expression. MIAPaCa-2 and H1299 cells show a change in IC₅₀ of 2.2 and 2.3 fold respectively. However, Panc-1 and H322 cells do not show significant changes in drug sensitivity. These data suggested that Tetrahydrouridine can sensitize some pancreatic and lung carcinoma cells to Gemcitabine-induced cell death regardless of CDA expression levels. Tetrahydrouridine inhibits S-phase without apoptosis^[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.

Administration of 167 mg/kg Tetrahydrouridine (THU) followed by 1.0 mg/kg DAC results in death in one male and eight females. Animals surviving to scheduled termination are generally asymptomatic with no treatment related effects observed in body weights, food consumption, clinical chemistry and urinalysis for a treatment up to 1.0 mg/kg DAC in combination with 167 mg/kg Tetrahydrouridine in animals^[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.

284.26

C₉H₂₀N₂O₈

四氢尿苷

Room temperature in continental US; may vary elsewhere.

Solution, -20°C, 2 years

DMSO : ≥ 100 mg/mL (351.79 mM)

H₂O : 50 mg/mL (175.90 mM; Need ultrasonic)

* “≥” means soluble, but saturation unknown.

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 (8.79 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (8.79 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 (8.79 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (8.79 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 (8.79 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (8.79 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]. Funamizu N, et al. Tetrahydrouridine inhibits cell proliferation through cell cycle regulation regardless of cytidine deaminase expression levels. PLoS One. 2012;7(5):e37424.

  [2]. Terse P, et al. Subchronic oral toxicity study of decitabine in combination with tetrahydrouridine in CD-1 mice. Int J Toxicol. 2014 Mar-Apr;33(2):75-85.

Cell growth for pancreatic and lung carcinoma cell lines is carried out using the colorimetric methylene blue assay in 96-well plates at a density of 5,000 cells/well. Cells are either exposed or not exposed to Tetrahydrouridine (100 µM), counting the first 12 hrs as Day 0. Mean values are calculated from three different wells in triplicates for four days^[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^[2]
CD-1 mice (male 30-38 g and female 24-31g) from are individually housed in polycarbonate cages suspended on stainless steel racks with SaniChip certified hardwood bedding. Mice are assigned to four dose groups and a vehicle control group. Animals are gavaged with DAC or its vehicle 1 hour ± 5 minutes after administration of THU or its vehicle at a dose volume of 10 mL/kg. The DAC doses are selected based on the range finding study in which the mice tolerated six oral doses (2x/week) of 0.1, 0.2 and 0.4 mg/kg DAC in combination with a fixed dose of 167 mg/kg THU. A fixed Tetrahydrouridine dose (500 mg/m²) and the optimal timing between Tetrahydrouridine and DAC administration (60 min) are selected. Conversion of milligrams per body surface area dose in mice into milligrams per kilogram body weight dose estimation is based on Michaelis constant (k_m) values for mice obtained from US Food and Drug Administration published guidelines. In brief, the mouse dose in milligrams per body surface area (500 mg/m²) is divided by the k_m of 3 to convert the dose to milligrams per kilogram body weight (167 mg/kg).

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]. Funamizu N, et al. Tetrahydrouridine inhibits cell proliferation through cell cycle regulation regardless of cytidine deaminase expression levels. PLoS One. 2012;7(5):e37424.

  [2]. Terse P, et al. Subchronic oral toxicity study of decitabine in combination with tetrahydrouridine in CD-1 mice. Int J Toxicol. 2014 Mar-Apr;33(2):75-85.