Martin, Logan R.

Isothermal titration calorimetry (ITC) and circular dichroism (CD) were used to study the thermodynamics of RPC·G‐quadruplex DNA (G4) complex formation. The ruthenium polypyridyl complexes (RPCs) were [Ru(phen)3]2+ (12+), [Ru(phen)2(dppz)]2+ (22+), [Ru(phen)2(tatpp)]2+ (32+), and [Ru(phen)2(tatpp)(phen)2Ru]4+ (44+), and target DNAs were c‐MYC NHE‐III1 promoter sequence mutants forming 1‐2‐1 and 1‐6‐1 G‐quadruplexes. Formation of the 2:1 RPC·G4 complexes is characterized by entropy driven RPC binding to the top and bottom of G‐tetrad faces. 12+ appears to bind very weakly or not at all to G4 DNA. 22+ having a dipyridophenazine group to stack on the top and bottom of the G4 core, exhibits an average Ka = 6.7 × 104 m–1. 32+, with a larger G4 interactive tetraazatetrapyridopentacene group, binds with significantly higher affinity, Ka = 1.1 × 106 m–1. 22+ and 32+ appear to bind independently of G4 folding topology and RPC conformation. The thermograms for the titration of G4 DNA with rac‐44+ are characterized by two binding modes exhibiting higher and lower affinity (Ka,1 = 3.6 × 107 m–1 and Ka,2 = 3.2 × 105 m–1). The two binding modes are attributed to preferential binding of one of the 44+ enantiomers (e.g. ΛΛ) over the other isomers (e.g. ΔΔ or ΔΛ). Tighter binding of the preferred 44+ enantiomer, in comparison to 32+, is due to additional favorable entropy for locating a second [(phen)2Ru–]2+ moiety in a G4 groove. Weaker binding of the disfavored 44+ isomers must be due to a poorer fit of these isomers with the G4 faces.

AbstractInvited for the cover of this issue is the group of Edwin Lewis, Mississippi State University, in collaboration with Frederick MacDonnell, University of Texas at Arlington, USA. The cover image shows Ru(phen)2(tatpp)]2+ binding to DNA.

The ruthenium(II) polypyridyl complexes (RPCs), [(phen)2Ru(tatpp)]Cl2 (3Cl2) and [(phen)2Ru (tatpp)Ru(phen)2]Cl4 (4Cl4), containing the large planar and redox‐active tetraazatetrapyrido‐pentacene (tatpp) ligand, cleave DNA in the presence of reducing agents in cell‐free assays and show significant tumor regression in mouse tumor models with human non‐small cell lung carcinoma xenografts. ITC, CD, and ESI‐MS techniques were used to study the thermodynamics of RPC·DNA complex formation and the complex structure for binding three different RPCs to duplex DNA. The specific RPCs were [Ru(phen)3]2+ (12+), [Ru(phen)2(dppz)]2+ (22+), and [Ru(phen)2(tatpp)]2+ (32+). We examined the enatiomerically pure Δ‐RPC and Λ‐RPC isomers as well as the racemic mixture in terms of their binding to B‐DNA. B‐DNA binding of the three RPCs is characterized by a combination of groove binding (including electrostatic effects) and intercalation, with the thermodynamics being very similar for binding both the enantiomerically pure compounds and the racemic mixture. 12+ is the weakest DNA binder, exhibiting a Ka = 1.3 × 104 m–1 while 22+ binds with significantly higher affinity, Ka,1 = 1.4 × 106 m–1, and 32+ exhibits the tightest binding, Ka = 4.7 × 106 m–1. The trend is to increase the affinity as longer bridging ligands engage in additional π‐bonding with the DNA base pairs via an intercalative binding mode. A second binding mode, two orders of magnitude weaker was also seen for 22+. The ITC values for binding the racemic mixtures exhibit ΔGs ranging from –5.6 to –9.1 kcal mol–1 (12+·DNA and 32+·DNA respectively), while the ITC values for ΔH range from +4.9 to –5.0 kcal mol–1 (32+·DNA and mode 2 binding for 22+·DNA respectively). All of the primary complexes exhibit very negative values for –TΔS ranging from –7.3 to –14.0 kcal mol–1 (mode 1 binding for 22+·DNA and 32+·DNA respectively). To further understand the intercalation vs. groove binding contributions to the overall binding energy we compared the thermodynamics for formation of the 12+·DNA, 22+·DNA, and 32+·DNA complexes to the thermodynamics for formation of the 44+·DNA complex. The RPC binding affinities to duplex DNA follow the trend: 12+ < 44+ < 22+ < 32+. Differences in the affinity for binding 12+ vs. 22+ or 32+ are almost entirely due to the size of the intercalating moiety, e.g. phen which can only be partially intercalated in comparison to dppz and tatpp which can be completely intercalated. The lower affinity for the dinuclear ruthenium 44+ is due to the solvation penalty for the second Ru core complex that extends out from the major groove. Comparing these results, we have begun to develop the structure function relationships for the interaction of RPCs with B‐DNA.