Electron paramagnetic resonance (EPR) study of spin-labeled camptothecin derivatives: a different look of the ternary complex.
Camptothecin (CPT) derivatives are clinically effective poisons of DNA topoisomerase I (Top1) able to form a ternary complex with the Top1-DNA complex. The aim of this investigation was to examine the dynamic aspects of the ternary complex formation by means of site-directed spin labeling electron paramagnetic resonance (SDSL-EPR). Two semisynthetic CPT derivatives bearing the paramagnetic moiety were synthesized, and their biological activity was tested. A 22-mer DNA oligonucleotide sequence with high affinity cleavage site for Top1 was also synthesized. EPR experiments were carried out on modified CPT in the presence of DNA, of Top1, or of both. In the last case, a slow motion component in the EPR signal appeared, indicating the formation of the ternary complex. Deconvolution of the EPR spectrum allowed to obtain the relative drug amounts in the complex. It was also possible to demonstrate that the residence time of CPT "trapped" in the ternary complex is longer than hundreds of microseconds.
Department of Pharmaceutical Sciences, University of Padova, Via Marzolo 5, 30039 Padova, Italy.
Antonio
A
Ricci
Jessica
J
Marinello
Marco
M
Bortolus
Albert
A
Sánchez
Anna
A
Grandas
Enrique
E
Pedroso
Yves
Y
Pommier
Giovanni
G
Capranico
Anna Lisa
AL
Maniero
Giuseppe
G
Zagotto
D000972:Antineoplastic Agents, Phytogenic / Q000737:chemistry; D001483:Base Sequence; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry; D003497:Cyclic N-Oxides / Q000737:chemistry; D004247:DNA / Q000138:chemical synthesis / Q000737:chemistry; D004264:DNA Topoisomerases, Type I / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D017550:Spectroscopy, Fourier Transform Infrared; D059004:Topoisomerase I Inhibitors / Q000737:chemistry
D000972:Antineoplastic Agents, Phytogenic; D003497:Cyclic N-Oxides; D059004:Topoisomerase I Inhibitors; D004247:DNA; D004264:DNA Topoisomerases, Type I; C528909:TOP1 protein, human; C003959:TEMPO; D002166:Camptothecin
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