Structural Basis of Topotecan−DNA Recognition Probed by Flow Linear Dichroism, Circular Dichroism, and Raman Spectroscopy

Abstract

Camptothecin (CPT) and its clinically important antitumor derivative topotecan (Tpt) were traditionally described as unique antitumor compounds exhibiting no affinity toward DNA alone or DNA topoisomerase I (top1) alone but interacting with both the enzyme and the DNA within the so-called ternary cleavable complexes. We present here the first experimental data on the molecular structure and geometry of Tpt-DNA complexes in solution. Tpt interacts with DNA within the DNA minor groove and demonstrates the preferential binding to GC-rich DNA. The flow linear dichroism (FLD) spectra show that the Tpt binds DNA only in lactone form and its chromophore forms the angle nearly 55°with the DNA long axis. Induced circular dichroism (CD) data independently confirm conclusions about Tpt preferable orientation drawn from the FLD experiments. The Raman spectroscopy data confirm the FLD and CD results and further demonstrate direct interactions of Tpt lactone ring with dG. The capability of Tpt to bind DNA in the minor groove of GC-rich DNA regions must be taken into account when considering molecular structure of ternary cleavable complexes of CPTs, DNA, and top1 in solution.

Structure and properties of camptothecin derivatives, their protonated forms, and model interaction with the topoisomerase I–DNA complex

“…Same enantiomeric forms are obtained for the studied‐CPTs with exception of (1), where R(−),S(+) diastereoisomer is stabilized (Figure 2). The obtained [α] D values (Figure 3 and Table II), correlated also well with the reported data for the optical properties of CPTs 46, 47…”

Binding of topotecan to chromatin: Insights into cooperative binding and comparison with DNA

“…Although, binding of TPT to DNA alters DNA helicity and base stacking, in agreement with other reports suggesting direct binding of TPT to DNA [11,28], in chromatin ellipticity at positive extreme (275 nm) remains almost unchanged indicating that the interaction of histones with DNA in chromatin structure covers some sites of TPT binding. A negative extreme at 220 nm, which corresponds to the protein moiety of chromatin becomes more positive upon TPT binding implying that the secondary structure of histones that is mainly in the form of ␣-helix is considerably reduced.…”

“…Although, direct interaction of TPT with DNA has been the subject of several reports [11,12,28], no work has been published on the effect of this drug on DNA-histone complex in chromatin structure. Therefore the goal of this study was to define binding affinity of TPT to chromatin compared to DNA to elucidate the mechanism of TPT action at the chromatin level and illustrate the possible role of histone proteins in this binding process.…”

Multiple Binding Modes of the Camptothecin Family to DNA Oligomers

“…These experiments strongly suggest that the G1 base, located at the edge of the duplex, is the predominant site of TPT binding, consistently with spectrophotometric studies. [14] However, our studies suggest that the interaction is one of stacking against a base pair rather than an interaction with the minor groove. [11] An issue that should be addressed in drug-DNA interactions is the possibility of spin diffusion as a source of indirect cross-peaks.…”