Borges Crispina, Lemière Filip, Embrechts Jan, Van Dongen Walter, Esmans Eddy L
Department of Chemistry, Nucleoside Research and Mass Spectrometry Unit, University of Antwerp, Groenenborgerlaan 171, B-2020 Antwerp, Belgium.
Rapid Commun Mass Spectrom. 2004;18(19):2191-200. doi: 10.1002/rcm.1610.
Xenobiotic and endobiotic molecules can react with DNA leading to formation of so-called DNA adducts. This modified DNA can be repaired enzymatically, but, if not, these modifications are believed to be responsible for the initiation of carcinogenic processes. Hence, we studied the interaction of 2'-deoxynucleosides and 2'-deoxynucleotides with 3,4-estronequinone (3,4-E(1)Q), a metabolite of estrone (E(1)) and a supposed carcinogen. These estrone-nucleic acid adducts were analysed by capillary liquid chromatography (CapLC) coupled to electrospray ionization mass spectrometry (ESI-MS). Knowledge of their behaviour from in vitro studies is a prerequisite for detecting adducts in in vivo studies. Our initial attempts to synthesise nucleos(t)ide adducts of 3,4-E(1)Q in an aprotic solvent (dimethylformamide) yielded no adducts. However, under acidic aqueous conditions, adducts were obtained. With dGuo, a dGuo adduct was found in addition to a Gua adduct. Earlier publications on adduct formation in protic solvents failed to report formation of any adduct with dAdo. A N(3)-Ade adduct was reported upon reaction of 3,4-E(1)Q with Ade base and with DNA. With dAdo, we obtained two nucleoside adducts and six Ade adducts due to loss of 2'-deoxyribose. Thus, contrary to general belief that only 2,3-E(1)Q can form stable adducts, we showed formation of substantial amounts of intact DNA adducts with 3,4-E(1)Q in addition to deglycosylated adducts. Adducts were also obtained with dGMP and dAMP, but no phosphate alkylation was found. Adducts of dCyd, dCMP, dThd, and dTMP were not detected. Using chromatographic-MS data a structural relationship between the 2'-deoxynucleoside, 2'-deoxynucleotide and base adducts was found in the various reaction mixtures. The adducts of dGuo and dGMP reaction mixtures were alkylated at the same N(7)-position of the nucleobase, as indicated by the occurrence of a rapid deglycosylation reaction. In dAdo and dAMP reaction mixtures, 14 adducts were detected; their relationships from the LC and MS data reduced the number of structures to six adenine base alkylated adducts with respect to alkylation between N(1), N(3), N(7) and/or N(6) in the adenine and C(1), C(2) and/or C(6) in 3,4-E(1)Q. We could infer, in addition, whether they had an A ring attachment or a C(6) attachment on the estrone moiety.
外源性和内源性分子可与DNA发生反应,导致形成所谓的DNA加合物。这种修饰后的DNA可通过酶促作用进行修复,但如果未被修复,这些修饰被认为是致癌过程起始的原因。因此,我们研究了2'-脱氧核苷和2'-脱氧核苷酸与3,4-雌酮醌(3,4-E(1)Q)的相互作用,3,4-雌酮醌是雌酮(E(1))的一种代谢产物,也是一种假定的致癌物。这些雌酮-核酸加合物通过毛细管液相色谱(CapLC)与电喷雾电离质谱(ESI-MS)联用进行分析。了解它们在体外研究中的行为是在体内研究中检测加合物的前提条件。我们最初尝试在非质子溶剂(二甲基甲酰胺)中合成3,4-E(1)Q的核苷(酸)加合物,但未得到加合物。然而,在酸性水溶液条件下,获得了加合物。对于dGuo,除了鸟嘌呤加合物外还发现了一种dGuo加合物。早期关于在质子溶剂中形成加合物的出版物未能报道与dAdo形成任何加合物的情况。有报道称3,4-E(1)Q与腺嘌呤碱基及DNA反应会形成N(3)-Ade加合物。对于dAdo,由于2'-脱氧核糖的丢失,我们得到了两种核苷加合物和六种腺嘌呤加合物。因此,与普遍认为只有2,3-E(1)Q能形成稳定加合物的观点相反,我们表明除了去糖基化加合物外,3,4-E(1)Q还能形成大量完整的DNA加合物。用dGMP和dAMP也获得了加合物,但未发现磷酸烷基化现象。未检测到dCyd、dCMP、dThd和dTMP的加合物。利用色谱-质谱数据,在各种反应混合物中发现了2'-脱氧核苷、2'-脱氧核苷酸和碱基加合物之间的结构关系。dGuo和dGMP反应混合物的加合物在核苷酸碱基的相同N(7)位置发生了烷基化,快速去糖基化反应的出现表明了这一点。在dAdo和dAMP反应混合物中,检测到了14种加合物;根据液相色谱和质谱数据,它们之间的关系将结构数量减少到六种腺嘌呤碱基烷基化加合物,涉及腺嘌呤中N(1)、N(3)、N(7)和/或N(6)以及3,4-E(1)Q中C(1)、C(2)和/或C(6)之间的烷基化。此外,我们还可以推断它们在雌酮部分是具有A环连接还是C(6)连接。