title: N-(2-chloroethyl)-N-nitrosoureas covalently bound to nonionic and monocationic lexitropsin dipeptides. Synthesis, DNA affinity binding characteristics, and reactions with 32P-end-labeled DNA.
abstract: The synthesis and characterization of a series of compounds that contain an N-alkyl-N-nitrosourea functionality linked to DNA minor groove binding bi- and tripeptides (lexitropsins or information-reading peptides) based on methylpyrrole-2-carboxamide subunits are described. The lexitropsins (lex) synthesized have either a 3-(dimethylamino)propyl or propyl substituent on the carboxyl terminus. The preferred DNA affinity binding sequences of these compounds were footprinted in 32P-end-labeled restriction fragments with methidiumpropyl-EDTA.Fe(II), and in common with other structural analogues, e.g., distamycin and netropsin, these nitrosoureas recognize A-T-rich runs. The affinity binding of the compound with the dimethylamino terminus, which is ionized at near-neutral pH, appeared stronger than that observed for the neutral dipeptide. The sequence specificity for DNA alkylation by (2-chloroethyl)nitrosourea-lex dipeptides (Cl-ENU-lex), with neutral and charged carboxyl termini, using 32P-end-labeled restriction fragments, was determined by the conversion of the adducted sites into single-strand breaks by sequential heating at neutral pH and exposure to base. The DNA cleavage sites were visualized by polyacrylamide gel electrophoresis and autoradiography. The alkylation of DNA by Cl-ENU-lex was compared to that by N-(2-chloroethyl)-N'-cyclohexyl-N-nitrosourea (CCNU), which has no DNA affinity binding properties. While all the Cl-ENU compounds generate DNA breaks as a consequence of the formation of N7-alkyl-guanine, the Cl-ENU-lex compounds induced, in a time- and dose-dependent fashion, intense DNA cleavage bands at adenine, cytosine, and thymine residues associated with affinity binding sites. These non-G cleavages induced by Cl-ENU-lex were inhibited by the coaddition of distamycin at concentrations that did not affect G alkylation break sites. CCNU, even at much higher concentrations, does not generate any similar detectable lesions at non-G sites. Therefore, linking the Cl-ENU moiety to minor groove binders is a viable strategy to qualitatively and quantitatively control the delivery and release of the ultimate DNA alkylating agent in a sequence-dependent fashion.
authors.affiliation: Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha 68198-6805.
authors.forename: K M
authors.initials: KM
authors.lastname: Church
authors.forename: R L
authors.initials: RL
authors.lastname: Wurdeman
authors.forename: Y
authors.initials: Y
authors.lastname: Zhang
authors.forename: F X
authors.initials: FX
authors.lastname: Chen
authors.forename: B
authors.initials: B
authors.lastname: Gold
mesh_terms: D000478:Alkylation; D001483:Base Sequence; D001665:Binding Sites; D002412:Cations; D004247:DNA; D004262:DNA Restriction Enzymes; D004214:Distamycins; D005038:Ethylnitrosourea / Q000138:chemical synthesis / Q000031:analogs & derivatives; D006146:Guanidines; D006147:Guanine; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009429:Netropsin / Q000031:analogs & derivatives
chemical_list: D002412:Cations; D004214:Distamycins; D006146:Guanidines; C053654:lexitropsin; C017139:1-(2-chloroethyl)-1-nitrosourea; D006147:Guanine; D009429:Netropsin; D004247:DNA; D004262:DNA Restriction Enzymes; D005038:Ethylnitrosourea