2',2'-Difluoro-2'-deoxyuridine
(Synonyms: 2'-脱氧-2',2'-二氟尿嘧啶核苷) 目录号 : GC46508
An active metabolite of gemcitabine
Cas No.:114248-23-6
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >95.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
2',2'-Difluoro-2'-deoxyuridine is an active metabolite of the anticancer nucleoside analog gemcitabine .1 It is formed by deamination of gemcitabine by cytidine deaminase in the liver. 2',2'-Difluoro-2'-deoxyuridine is cytotoxic to HepG2 and A549 cancer cells (IC50s = 3.13 and 3.92 µM, respectively). It enhances radiation-induced cell death of ECV304 and NCI H292 cells when used at concentrations ranging from 10 to 100 µM.2
1.Veltkamp, S.A., Pluim, D., van Eijndhoven, M.A.J., et al.New insights into the pharmacology and cytotoxicity of gemcitabine and 2',2'-difluorodeoxyuridineMol. Cancer Ther.7(8)2415-2425(2008) 2.Pauwels, B., Korst, A.E.C., Lambrechts, H.A.J., et al.The radiosensitising effect of difluorodeoxyuridine, a metabolite of gemcitabine, in vitroCancer Chemother. Pharmacol.58(2)219-228(2006)
| Cas No. | 114248-23-6 | SDF | |
| 别名 | 2'-脱氧-2',2'-二氟尿嘧啶核苷 | ||
| Canonical SMILES | O=C(N(C=C1)[C@H]2C(F)(F)[C@H](O)[C@@H](CO)O2)NC1=O | ||
| 分子式 | C9H10F2N2O5 | 分子量 | 264.2 |
| 溶解度 | DMF: 20 mg/ml,DMSO: 20 mg/ml,Ethanol: 25 mg/ml,PBS (pH 7.2): 10 mg/ml | 储存条件 | Store at -20°C |
| General tips | 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 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。 为了提高溶解度，请将管子加热至37℃，然后在超声波浴中震荡一段时间。 |
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| Shipping Condition | 评估样品解决方案：配备蓝冰进行发货。所有其他可用尺寸：配备RT，或根据请求配备蓝冰。 | ||
| 制备储备液 | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 3.785 mL | 18.9251 mL | 37.8501 mL |
| 5 mM | 0.757 mL | 3.785 mL | 7.57 mL |
| 10 mM | 0.3785 mL | 1.8925 mL | 3.785 mL |
| 第一步：请输入基本实验信息（考虑到实验过程中的损耗，建议多配一只动物的药量） | ||||||||||
| 给药剂量 | mg/kg |  |
动物平均体重 | g | |
每只动物给药体积 | ul | |
动物数量 | 只 |
| 第二步：请输入动物体内配方组成（配方适用于不溶于水的药物；不同批次药物配方比例不同，请联系GLPBIO为您提供正确的澄清溶液配方） | ||||||||||
| % DMSO % % Tween 80 % saline | ||||||||||
| 计算重置 | ||||||||||
计算结果:
工作液浓度： mg/ml；
DMSO母液配制方法： mg 药物溶于 μL DMSO溶液（母液浓度 mg/mL，
体内配方配制方法：取 μL DMSO母液，加入 μL PEG300，混匀澄清后加入μL Tween 80，混匀澄清后加入 μL saline，混匀澄清。
1. 首先保证母液是澄清的；
2.
一定要按照顺序依次将溶剂加入，进行下一步操作之前必须保证上一步操作得到的是澄清的溶液，可采用涡旋、超声或水浴加热等物理方法助溶。
3. 以上所有助溶剂都可在 GlpBio 网站选购。
Inhibition of thymidylate synthase by 2',2'-difluoro-2'-deoxycytidine (Gemcitabine) and its metabolite 2',2'-Difluoro-2'-deoxyuridine
Int J Biochem Cell Biol 2015 Mar;60:73-81.PMID:25562513DOI:10.1016/j.biocel.2014.12.010.
2',2'-Difluoro-2'-deoxycytidine (dFdC, gemcitabine) is a cytidine analogue active against several solid tumor types, such as ovarian, pancreatic and non-small cell lung cancer. The compound has a complex mechanism of action. Because of the structural similarity of one metabolite of dFdC, dFdUMP, with the natural substrate for thymidylate synthase (TS) dUMP, we investigated whether dFdC and its deamination product 2',2'-Difluoro-2'-deoxyuridine (dFdU) would inhibit TS. This study was performed using two solid tumor cell lines: the human ovarian carcinoma cell line A2780 and its dFdC-resistant variant AG6000. The specific TS inhibitor Raltitrexed (RTX) was included as a positive control. Using the in situ TS activity assay measuring the intracellular conversion of [5-(3)H]-2'-deoxyuridine or [5-(3)H]-2'-deoxycytidine to dTMP and tritiated water, it was observed that dFdC and dFdU inhibited TS. In A2780 cells after a 4h exposure to 1 μM dFdC tritium release was inhibited by 50% but did not increase after 24h, Inhibition was also observed following dFdU at 100 μM. No effect was observed in the dFdC-resistant cell line AG6000; in this cell line only RTX had an inhibitory effect on TS activity. In the A2780 cell line RTX inhibited TS in a time dependent manner. In addition, DNA specific compounds such as 2'-C-cyano-2'-deoxy-1-beta-D-arabino-pentafuranosylcytosine and aphidicoline were utilized to exclude DNA inhibition mediated down regulation of the thymidine kinase. Inhibition of the enzyme resulted in a relative increase of mis-incorporation of [5-(3)H]-2'-deoxyuridine into DNA. In an attempt to elucidate the mechanism of in situ TS inhibition the ternary complex formation and possible inhibition in cellular extracts of A2780 cells, before and after exposure to dFdC, were determined. With the applied methods no proof for formation of a stable complex was found. In simultaneously performed experiments with 5FU such a complex formation could be demonstrated. However, using purified TS it was demonstrated that dFdUMP and not dFdCMP competitively inhibited TS with a Ki of 130 μM, without ternary complex formation. In conclusion, in this paper we reveal a new target of dFdC: thymidylate synthase.
LC-MS/MS method for quantitation of gemcitabine and its metabolite 2',2'-Difluoro-2'-deoxyuridine in mouse plasma and brain tissue: Application to a preclinical pharmacokinetic study
J Pharm Biomed Anal 2021 May 10;198:114025.PMID:33744463DOI:10.1016/j.jpba.2021.114025.
A simple, sensitive, and relatively fast assay was developed and validated for the quantitation of gemcitabine (dFdC) and its major metabolite 2',2'-Difluoro-2'-deoxyuridine (dFdU) in mouse plasma and brain tissue. The assay used a small sample (25 μL plasma and 5 mg brain) for extraction by protein precipitation. After dilution of the supernatant extract, 1 μL was injected into HPLC system for reverse phase chromatographic separation with a total run time of 8 min. Chromatographic resolution of dFdC and dFdU was achieved on a Gemini C18 column (50 × 4.6 mm, 3 μm) utilizing gradient elution. Multiple reaction monitoring (MRM) with positive/negative ion switching was performed for detection of dFdC and its internal standard (dFdC-IS) in positive ion mode and dFdU and its IS (dFdU-IS) in negative ion mode. Two calibration curves ranging from 5-2000 ng/mL and 250-50,000 ng/mL were generated for dFdC and dFdU in mouse plasma, respectively. For measurement of dFdC and dFdU in mouse brain tissue, another two curves were used ranging from 0.02 to 40 ng/mg and 1-40 ng/mg, respectively. This assay demonstrated excellent precision and accuracy within day and between days for simultaneous measurement of dFdC and dFdU at all the concentration levels in both matrices. The other parameters such as selectivity, sensitivity, matrix effects, recovery, and storage stability were also assessed for both analytes in each matrix. Compared to the previously reported methods, the sample extraction in the current assay was simplified significantly, and the analysis time was greatly shortened. We successfully applied the validated method to the analysis of dFdC and dFdU in mouse plasma, brain, and brain tumor tissue in a preclinical pharmacokinetic study.
Determination of the antimetabolite Gemcitabine (2',2'-difluoro-2'-deoxycytidine) and of 2',2'-Difluoro-2'-deoxyuridine by 19F nuclear magnetic resonance spectroscopy
Anal Biochem 1993 Oct;214(1):25-30.PMID:8250231DOI:10.1006/abio.1993.1451.
The analysis of the new antimetabolite 2',2'-difluoro-2'-deoxycytidine (Gemcitabine, Eli Lilly Corp.) and of its metabolic deamination product, 2',2'-Difluoro-2'-deoxyuridine, in urine and plasma by means of 19F NMR is described. Both compounds show AB-type NMR spectra which are characterized as a function of pH at a magnetic field strength of 9.4 T. In the physiological pH range the spectra of both compounds are distinct, despite the fact that both compounds have almost identical average 19F shifts. A linear relation between NMR intensity and concentration in urine has been established. The NMR method does not require sample pretreatment and allows for rapid, nondestructive analysis of Gemcitabine concentrations over 0.01 mM. Separation of the two compounds in vivo by 19F NMR spectroscopy is difficult to achieve.
Preanalytical Stability of Gemcitabine and its Metabolite 2', 2'-Difluoro-2'-Deoxyuridine in Whole Blood-Assessed by Liquid Chromatography Tandem Mass Spectrometry
J Pharm Sci 2015 Dec;104(12):4427-4432.PMID:26372902DOI:10.1002/jps.24638.
Gemcitabine (2',2'-difluoro-2'-deoxycytidine, dFdC) and metabolite (2',2'-Difluoro-2'-deoxyuridine, dFdU) quantification is warranted for individualized treatment strategies. Analyte stability is crucial for the validity of such quantification. We therefore studied the impact of the time interval from blood sampling to separation of plasma on gemcitabine stability. Blood from gemcitabine-treated patients was drawn into tetrahydrouridine (THU)-spiked heparin and ethylenediaminetetraacetic acid tubes and kept on ice until separation. Plasma was separated sequentially up to 24 h after sampling and dFdC and dFdU were quantified by liquid chromatography tandem mass spectrometry (LC-MS/MS). The change in plasma concentrations over time was compared with the highest imprecision for concentrations above the lower limit of quantification of the LC-MS/MS method. Analyte concentrations decreased slightly over time, but for samples stored for 4 h on ice, the decline was smaller than the expected analytical imprecision. After 24 h, the maximum decline was 14.0%, which exceeded the expected analytical imprecision. dFdC and dFdU stabilities were acceptable for at least 4 h when THU-spiked whole blood samples were kept on ice. This is within the scope of routine sampling procedures. Further, variations in separation time intervals within this time frame are negligible when interpreting drug concentrations.
Modulation of gemcitabine (2',2'-difluoro-2'-deoxycytidine) pharmacokinetics, metabolism, and bioavailability in mice by 3,4,5,6-tetrahydrouridine
Clin Cancer Res 2008 Jun 1;14(11):3529-35.PMID:18519786DOI:10.1158/1078-0432.CCR-07-4885.
Purpose: In vivo, 2',2'-difluoro-2'-deoxycytidine (dFdC) is rapidly inactivated by gut and liver cytidine deaminase (CD) to 2',2'-Difluoro-2'-deoxyuridine (dFdU). Consequently, dFdC has poor oral bioavailability and is administered i.v., with associated costs and limitations in administration schedules. 3,4,5,6-Tetrahydrouridine (THU) is a potent CD inhibitor with a 20% oral bioavailability. We investigated the ability of THU to decrease elimination and first-pass effect by CD, thereby enabling oral dosing of dFdC. Experimental design: A liquid chromatography-tandem mass spectrometry assay was developed for plasma dFdC and dFdU. Mice were dosed with 100 mg/kg dFdC i.v. or orally with or without 100 mg/kg THU i.v. or orally. At specified times between 5 and 1,440 min, mice (n = 3) were euthanized. dFdC, dFdU, and THU concentrations were quantitated in plasma and urine. Results: THU i.v. and orally produced concentrations >4 microg/mL for 3 and 2 h, respectively, whereas concentrations of >1 microg/mL have been associated with near-complete inhibition of CD in vitro. THU i.v. decreased plasma dFdU concentrations but had no effect on dFdC plasma area under the plasma concentration versus time curve after i.v. dFdC dosing. Both THU i.v. and orally substantially increased oral bioavailability of dFdC. Absorption of dFdC orally was 59%, but only 10% passed liver and gut CD and eventually reached the systemic circulation. Coadministration of THU orally increased dFdC oral bioavailability from 10% to 40%. Conclusions: Coadministration of THU enables oral dosing of dFdC and warrants clinical testing. Oral dFdC treatment would be easier and cheaper, potentially prolong dFdC exposure, and enable exploration of administration schedules considered impractical by the i.v. route.