Subject(s)
Anti-HIV Agents/pharmacology , Drug Design , RNA/chemistry , Terphenyl Compounds/pharmacology , rev Gene Products, Human Immunodeficiency Virus/antagonists & inhibitors , Anti-HIV Agents/chemical synthesis , Anti-HIV Agents/chemistry , Binding Sites , Dose-Response Relationship, Drug , HIV-1/drug effects , Humans , Molecular Structure , Structure-Activity Relationship , Terphenyl Compounds/chemical synthesis , Terphenyl Compounds/chemistry , Virus Replication/drug effects , rev Gene Products, Human Immunodeficiency Virus/metabolismABSTRACT
The bisnaphthalimide cytotoxic agent elinafide exhibits a mixed DNA binding mode including groove-association and intercalation. We have compared the interaction of elinafide and two bisnaphthalimide analogues with various natural and modified DNA sequences using solution NMR and UV-melting methods and surface plasmon resonance (SPR) experiments at different pH conditions. The combined data obtained with these techniques established a high-affinity binding mode comprising intercalation and strong electrostatic contacts with guanine bases in the major groove, and a weaker interaction with A·T pairs likely involving groove association. However, the SPR binding constants and the NMR and UV-melting binding parameters responded differently to variations in DNA bases and ligand intercalating moieties. The rates and equilibrium constants determined by SPR clearly responded to changes in pH and DNA groove composition, but were rather insensitive to alterations in drug rings and DNA bases affecting the intercalation process. Conversely, the intermolecular stacking interactions detected by NMR and the ligand-induced thermal stabilizations measured by UV depended on both sets of factors and were controlled by the sequence-dependent properties of the DNA helices, indicating that these data were modulated by naphthalimide stacking in addition to groove association. A two-step binding process where a groove-bound state is required prior to intercalation is proposed as an explanation for these observations. These findings may be useful for studying other classes of DNA- and RNA-binding drugs, which frequently combine groove-binding and stacking moieties.
Subject(s)
Amides/chemistry , Antineoplastic Agents/chemistry , DNA/chemistry , Isoquinolines/chemistry , Naphthalimides/chemistry , Hydrogen-Ion Concentration , Magnetic Resonance Spectroscopy , Nucleic Acid Denaturation , Solutions , Surface Plasmon ResonanceABSTRACT
The sequence-specific structural and dynamic properties of double-helical DNA play important roles in many biological processes involving DNA recognition. Using a combination of NMR spectroscopy, surface plasmon resonance, and UV thermal denaturation experiments, we have investigated how sequences not making direct contact with the drug modulate the interaction between the cytotoxic agent elinafide and its preferred bisintercalation sites on double-helical DNA. Our combined data are consistent with two superposed interactions, one process involving ligand binding to the DNA duplex with nanomolar dissociation constants and another process of ring intercalation characterized by faster dissociation rates and substantially higher dissociation constants in some cases. The sequence of the base pairs flanking the bisnaphthalimide binding tetranucleotides influence both events through indirect readout effects, but these effects appear to be particularly relevant for the second (intercalation) process. The most unfavorable sequences contain specifically oriented A-tracts that oppose DNA intercalation of the naphthalimide rings, as reflected by strikingly different thermal stability and thermodynamic binding profiles. The complexes of elinafide with these sequences are characterized by poor DNA-naphthalimide and DNA-DNA stacking interactions and by enhanced dynamics of the ligand's intercalated rings and of the base pairs forming the tetranucleotide binding site.
