Czechowska Agnieszka, Poplawski Tomasz, Drzewoski Jozef, Blasiak Janusz
Department of Molecular Genetics, University of Lodz, Ul. Banacha 12/16, 90-237 Lodz, Poland.
Chem Biol Interact. 2005 Apr 15;152(2-3):139-50. doi: 10.1016/j.cbi.2005.03.002.
Imatinib (STI571) is a 2-phenylaminopyrimidine derivative used mostly in the treatment of chronic myeloid leukaemia. It targets the BCR/ABL oncogenic tyrosine kinase, inhibiting its activity. Using the alkaline comet assay we showed that STI571 at concentrations ranging from 0.2 to 2 microM induced DNA damage in human leukemic K562 and BV173 cells expressing the BCR/ABL oncogene, whereas it had no effect in normal human lymphocytes and leukemic CCRF-CEM cells without the expression of BCR/ABL. Imatinib did not induce DNA strand breaks in the direct interaction with DNA as examined by the circular plasmid relaxation assay. Because the extent of DNA damage observed in the neutral and pH 12.1 versions of the comet assay was much lesser than in the alkaline version, we concluded that the drug induced DNA alkali-labile sites rather than strand breaks. K562 cells were unable to repair H(2)O(2)-induced DNA damage during a 120-min incubation, if they had been preincubated with STI571, whereas normal lymphocytes did so within 60 min. Pre-treatment of K562 cells with Vitamins A, C and E reduced the extent of DNA damage evoked by STI571. Similar results brought experiments with the nitrone spin traps POBN and PBN, suggesting that free radicals may be involved in the formation of DNA lesions induced by STI571 in K562 cells. These cells exposed to imatinib and treated with endonuclease III, formamidopyrimidine-DNA glycosylase and 3-methyladenine-DNA glycosylase II, the enzymes recognizing oxidized and alkylated bases, displayed greater extent of DNA damage than those not treated with these enzymes. Therefore, the mechanism of the anti-leukemic action of STI571 may involve not only the inhibition of BCR/ABL, but also DNA damage in the cells expressing this fusion protein. DNA damage induced by STI571 may follow from oxidative and alkylative base modifications.
伊马替尼（STI571）是一种2-苯基氨基嘧啶衍生物，主要用于治疗慢性粒细胞白血病。它作用于BCR/ABL致癌性酪氨酸激酶，抑制其活性。我们使用碱性彗星试验表明，浓度在0.2至2微摩尔范围内的STI571可诱导表达BCR/ABL癌基因的人白血病K562和BV173细胞发生DNA损伤，而对正常人类淋巴细胞和不表达BCR/ABL的白血病CCRF-CEM细胞没有影响。通过环状质粒松弛试验检测，伊马替尼在与DNA直接相互作用时不会诱导DNA链断裂。由于在彗星试验的中性和pH 12.1版本中观察到的DNA损伤程度远低于碱性版本，我们得出结论，该药物诱导的是DNA碱不稳定位点而非链断裂。如果K562细胞预先用STI571孵育，那么在120分钟的孵育过程中它们无法修复过氧化氢诱导的DNA损伤，而正常淋巴细胞在60分钟内就能修复。用维生素A、C和E预处理K562细胞可降低STI571引起的DNA损伤程度。使用硝酮自旋捕获剂POBN和PBN进行的实验也得到了类似结果，这表明自由基可能参与了STI571在K562细胞中诱导的DNA损伤形成过程。这些暴露于伊马替尼并经核酸内切酶III、甲酰胺嘧啶-DNA糖基化酶和3-甲基腺嘌呤-DNA糖基化酶II（识别氧化和烷基化碱基的酶）处理的细胞，与未用这些酶处理的细胞相比，显示出更大程度的DNA损伤。因此，STI571的抗白血病作用机制可能不仅涉及对BCR/ABL的抑制，还包括对表达这种融合蛋白的细胞的DNA损伤。STI571诱导的DNA损伤可能源于氧化和烷基化碱基修饰。
