Background
Daunorubicin is an inhibitor of DNA topoisomerase II [1].
Daunorubicin is an anthracycline antibiotic. It is also used as an effective chemotherapeutic agent against tumors especially acute myeloid leukaemia and acute lymphocytic leukaemia. Daunorubicin can affect the metabolism and synthesis of DNA and RNA. In the in vitro assay, daunorubicin inhibits the incorporation of thymidine and uridine into L1210 cells. It also inhibits the incorporation of labeled precursors into the isolated DNA and RNA from incubated cells. When treated with leukemic cells isolated from acute lymphocytic leukemia patients, daunorubicin significantly inhibits the biosynthesis of the DNA and RNA macromolecules [2, 3].
多柔比星是DNA拓扑异构酶II的抑制剂[1]。
多柔比星是蒽环类抗生素，也是治疗肿瘤，特别是急性髓细胞白血病和急性淋巴细胞白血病的有效化疗药物。多柔比星可以影响DNA和RNA的代谢和合成。在体外实验中，多柔比星抑制了thymidine和uridine的进入L1210细胞，同时抑制了孵育细胞后分离的DNA和RNA中标记前体的进入。当用多柔比星处理急性淋巴细胞白血病患者分离的白血病细胞时，多柔比星明显抑制了DNA和RNA大分子的生物合成[2，3]。
参考文献:
[1] Hande K R. Etoposide: four decades of development of a topoisomerase II inhibitor. European Journal of Cancer, 1998, 34(10): 1514-1521.
[2] Momparler R L, Karon M, Siegel S E, et al. Effect of adriamycin on DNA, RNA, and protein synthesis in cell-free systems and intact cells. Cancer Research, 1976, 36(8): 2891-2895.
[3] Meriwether W D, Bachur N R. Inhibition of DNA and RNA metabolism by daunorubicin and adriamycin in L1210 mouse leukemia. Cancer research, 1972, 32(6): 1137-1142.
Protocol
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Cell experiment:
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The chemosensitivity to Daunorubicin is assessed using the MTT assay. In brief, the 96 well plates are set up with cells at the initial density of 2×105 cells/mL and are incubated at 37°C for 72 h in an atmosphere of 5% CO2 in the absence and presence of nine different concentrations of Daunorubicin (Dnr) or Dox ranging from 1.90 to 0.007 μM in triplicate. After incubation, 10 μL of MTT solution (5 mg/mL tetrazolium salt) is added to each well and the plates are incubated for a further 4 h at 37°C. The formazan salt crystals are dissolved by adding 100 μL 10% SDS in 10 mM HCl solution and incubating over night at 37°C. The absorbance is measured at 540 nm with a reference at 650 nm by a 96-well enzyme-linked immunosorbent assay (ELISA) plate reader. Chemosensitivity is expressed as the IC50, which is the concentration of drug causing 50% cell survival compare to control cells grown without drug. Calculations are carried out using Microsoft Excel[2].
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Animal experiment:
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Rat[4]Eight-week-old male Sprague-Dawley rats are used. The animals are quarantined and acclimatized for the additional 2 weeks prior to the initiation of the experiments. On day 0, each animal receives a single intravenous injection of Daunorubicin at a dose of 3 mg/kg (i.v.). Daunorubicin is administered in three equal injections at 48 h intervals for a period of one week to achieve an accumulative dose of 9 mg/kg, which is well documented to produce cardiotoxicity and nephrotoxicity. Age-matched rats are injected with corresponding volumes of 0.9% NaCl and used as a control (group Control;n=5). Twenty-two DNR-treated rats are randomly divided into two groups and received oral administration of Telmisartan (10 mg/kg/day; group Daunorubicin+Telmisartan; n=10) or vehicle (group Daunorubicin; n=12). The dose of Telmisartan is chosen on the basis of a previous report. Administration of Telmisartan is started on the same day as Daunorubicin administration and continued for 5 additional weeks after cessation of Daunorubicin administration (6 weeks total period). This duration of study is chosen on the basis of previous reports. On day 41, rats are placed individually in metabolic cages for 24-h urine collections for the measurement of protein concentrations and body weight (BW) is measured. After the end of the study period (6 weeks), rats are sacrificed and kidney tissue is harvested for semi-quantitative immunoblotting and immunohistochemical studies.
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参考文献:
[1]. Lehmann M, et al. Activity of topoisomerase inhibitors daunorubicin, idarubicin, and aclarubicin in the Drosophila Somatic Mutation and Recombination Test. Environ Mol Mutagen. 2004;43(4):250-7.
[2]. Dano K, et al. Inhibition of DNA and RNA synthesis by daunorubicin in sensitive and resistant Ehrlich ascites tumor cells in vitro. Cancer Res. 1972 Jun;32(6):1307-14.
[3]. Svensson SP, et al. Melanin inhibits cytotoxic effects of Doxorubicin and Daunorubicin in MOLT 4 cells. Pigment Cell Res. 2003 Aug;16(4):351-4
[4]. Gervasoni JE Jr, et al. An effective in vitro antitumor response against human pancreatic carcinoma with paclitaxel and Daunorubicin by induction of both necrosis and apoptosis. Anticancer Res. 2004 Sep-Oct;24(5A):2617-26
[5]. Arozal W, et al. Telmisartan prevents the progression of renal injury in daunorubicin rats with the alteration of angiotensin II and endothelin-1 receptor expression associated with its PPAR-γ agonist actions. Toxicology. 2011 Jan 11;279(1-3):91-9.
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