title: Alkylation of 2'-deoxythymidylyl(3'----5')-2'-deoxythymidine with 2-hydroxyethylnitrosourea and stability of the resulting phosphotriester.
abstract: One of the main targets of RNA and DNA alkylation is the phosphate group. In contrast to RNA phosphotriesters, DNA phosphotriesters are relatively stable. Introduction of 2-hydroxyethyl phosphotriesters into DNA, however, has been reported to decrease its stability towards hydrolytic cleavage considerably. 2-Chloroethylnitrosoureas (CNUs) have been found to form predominantly 2-hydroxyethyl adducts of DNA. In the present study, we have determined the stability of 2'-deoxythymidylyl-(3'----5')-2'-deoxythymidine (dTpdT) after alkylation with various nitrosoureas, including N-methyl-N-nitrosourea (MNU) and 2-hydroxyethylnitrosourea (HENU) at alkaline and neutral pH (at 37 degrees C). The half-lives (t1/2) at pH 12.5 were, for example, 2.8 h for di(2'-deoxythymidine)methylphosphotriester [dTp(me)dT] but less than 1 min for di(2'-deoxythymidine) (2-hydroxyethyl)-phosphotriester [dTp(he)dT]. At pH 7.0, dTp(me)dT was stable for more than three days, whereas the t1/2 for dTp(he)dT was only 1.0 h at pH 7.0 (27 min at pH 9.0). The marked lability of dTp(he)dT, in comparison to other phosphotriester analogues, can be explained by intermediate formation of a dioxaphospholane ring resulting in triester bond breakage. Our data strongly support the hypothesis that a great proportion of DNA single-strand breaks induced in DNA by HENU or CNUs can be attributed to intermediate formation of 2-hydroxyethyl phosphotriesters.
mesh_terms: D000478:Alkylation; D055598:Chemical Phenomena; D002621:Chemistry; D004247:DNA / Q000378:metabolism; D015226:Dinucleoside Phosphates; D005038:Ethylnitrosourea / Q000031:analogs & derivatives* / Q000378:metabolism; D006207:Half-Life; D010710:Phosphates / Q000378:metabolism; D012313:RNA / Q000378:metabolism; D013942:Thymine Nucleotides / Q000378:metabolism*
chemical_list: D015226:Dinucleoside Phosphates; D010710:Phosphates; D013942:Thymine Nucleotides; C022146:1-(2-hydroxyethyl)-1-nitrosourea; C017758:thymidylyl-(3'-5')-thymidine; D012313:RNA; D004247:DNA; D005038:Ethylnitrosourea