Observation of daunomycin and nogalamycin complexes with duplex DNA using electrospray ionisation mass spectrometry

Abstract

The noncovalent binding of the antitumour drugs daunomycin and nogalamycin to duplex DNA has been studied using electrospray ionisation mass spectrometry (ESI-MS). The conditions for the preparation of drug/duplex DNA complexes and for their detection by ESI-MS have been optimised. Ions corresponding to these complexes were most abundant relative to free DNA when prepared in the pH range 8-9, and using gentle ESI interface conditions. Self-complementary oligonucleotides, 5'-d(GGCTAGCC)-3' or 5'-d(CGGCGCCG)-3', annealed in the presence of a 5-fold molar excess of either nogalamycin or daunomycin gave ESI mass spectra in which the most intense ions corresponded to three molecules of drug bound to duplex DNA, with some evidence for four drug molecules bound. For binding to 5'-d(TGAGCTAGCTCA)(2)-3', complexes containing up to four nogalamycin and six daunomycin molecules were observed. These data are consistent with the neighbour exclusion principle whereby intercalation occurs between every other base pair such that up to four bound drugs would be expected for the 8 mers and up to six for the 12 mer. Competition experiments involving a single drug in an equimolar mixture of two oligonucleotides (5'-d(TGAGCTAGCTCA)(2)-3' with either 5'-d(CGGCGCCG)(2)-3' or 5'-d(GGCTAGCC)(2)-3') showed ions arising from complexes of drug/5'-d(CGGCGCCG)(2)-3' were more intense than complexes of drug/5'-d(GGCTAGCC)(2)-3', relative to those from the 12 mer in each mixture. While this suggests ESI-MS has the potential to detect differences in sequence selectivity, more detailed experiments involving a comparison of the relative ionisation efficiency of different oligonucleotides and a wider range of intercalators are required to establish this definitively. ESI mass spectra from experiments in which both drugs were reacted with the same oligonucleotide were more complex, such that a clear preference for one drug could not be established.

Evaluation of complexes of DNA duplexes and novel benzoxazoles or benzimidazoles by electrospray ionization mass spectrometry

“…The lack of a well-defined DNA binding stoichiometry by the antibiotic analogs is reminiscent of results reported by Gabelica et al for drug-DNA complexes containing known intercalators such as ethidium bromide, amsacrine, and ascididemin [16]; as well as Kapur et al for the more complex intercalators daunomycin and nogalamycin [36]. In the study by Gabelica, the DNA-binding modes observed in the gas-phase for ethidium bromide, amsacrine, and ascididemin exhibited the non-specific multiple-drug binding which parallels the binding behaviors observed in solution [16].…”

“…However, the relative peak intensities also paralleled ligand polarity, including ligand charge, and thus the observed binding pattern could simply be an artifact of the electrospray process. The peak distribution patterns reported by Kapur et al for daunomycin and nogalamycin were interpreted as following the neighbor exclusion principle predicted for intercalators, in which one drug molecule is inserted between every other base pair in the duplex DNA [36].…”

Comparison of the binding stoichiometries of positively charged DNA-binding drugs using positive and negative ion electrospray ionization mass spectrometry

“…Mixtures of dsDNA with [Ru(phen) 2 dpq]Cl 2 or [Ru(phen) 2 dpqC]Cl 2 were prepared as previously described [15]. It should be noted that in our laboratory we have established that no significant differences in the relative abundances of drug-dsDNA complexes are observed when the drugs are added either after or during annealing of the DNA with the non self-complementary 16 mers used in these experiments [6,14].…”

“…Electrospray ionization mass spec-trometry (ESI-MS) has substantial potential for this purpose. Several research groups have used ESI-MS to assess the relative affinities and stoichiometries of drugs that bind to DNA [5,6]. Although many noncovalent complexes have been studied by ESI-MS, questions remain as to the extent to which complexes detected in the gas phase reflect the solution equilibrium position existing prior to injection into the mass spectrometer [7][8][9].…”

“…This has been achieved using competition experiments to compare the affinities of intercalators for different DNA sequences [6], and to compare the affinities of different minor groove-binders for different DNA sequences [5,11,15].…”