Ni2+ and Cu2+ complexes of a phenanthroline-based ligand bind to G-quadruplexes at non-overlapping sites.

Transition metal complexes allow fine tuning of DNA binding affinity and selectivity. Here we report on the nucleic acid recognition properties of a phenanthroline-based ligand coordinated to Ni(2+) or Cu(2+). The resulting complexes clearly bind to telomeric G-quadruplexes at different sites according to the nature of the bound metal ion.

Effect of G-quadruplex polymorphism on the recognition of telomeric DNA by a metal complex.

The physiological role(s) played by G-quadruplexes renders these 'non-canonical' DNA secondary structures interesting new targets for therapeutic intervention. In particular, the search for ligands for selective recognition and stabilization of G-quadruplex arrangements has led to a number of novel targeted agents. An interesting approach is represented by the use of metal-complexes, their binding to DNA being modulated by ligand and metal ion nature, and by complex stoichiometry. In this work we characterized thermodynamically and stereochemically the interactions of a Ni(II) bis-phenanthroline derivative with telomeric G-quadruplex sequences using calorimetric, chiroptical and NMR techniques. We employed three strictly related sequences based on the human telomeric repeat, namely Tel22, Tel26 and wtTel26, which assume distinct conformations in potassium containing solutions. We were able to monitor specific enthalpy/entropy changes according to the structural features of the target telomeric sequence and to dissect the binding process into distinct events. Interestingly, temperature effects turned out to be prominent both in terms of binding stoichiometry and ΔH/ΔS contributions, while the final G-quadruplex-metal complex architecture tended to merge for the examined sequences. These results underline the critical choice of experimental conditions and DNA sequence for practical use of thermodynamic data in the rational development of effective G-quadruplex binders.

Metal ion-mediated assembly of effective phenanthroline-based G-quadruplex ligands.

Transition metal ions can drive the assembly of small planar ligands to produce structures able to efficiently recognize G-quadruplex DNA arrangements.

Perylene side chains modulate G-quadruplex conformation in biologically relevant DNA sequences.

The stabilisation of different G-quadruplex intra- and intermolecular structures by a number of perylene derivatives characterised by side chains ending with linear or cyclic amines was investigated by electrophoretic (EMSA) and spectroscopic (CD) techniques. The G-rich sequences included the biologically relevant human telomeric TTAGGG runs and the NHE region of the c-myc oncogene. The test compounds could be subdivided into two families: derivatives carrying a cyclic amine in the side chains, which show a reduced binding to the G-quadruplex form, and linear amine congeners, exhibiting enhanced affinity. The latter efficiently induce pairing of multiple DNA chains, while the former are not able to overcome the original folding of the nucleic acid sequence which is preserved in the complex. Remarkably, addition of the perylenes to G-rich sequences paired in a double helical form results in G-quadruplex induction by weak binders only. This is likely related to the ability of strong G-quadruplex binders, but not of weak G-quadruplex binders, to efficiently intercalate into the double-stranded arrangement, which becomes stabilised and is not prone to undergo denaturation and subsequent G-quadruplex folding essentially for kinetic reasons. Hence, two apparently conflicting requirements emerge from this work. In fact, linear alkylamino terminals in the perylene side chains are capable of strong and selective G-quadruplex recognition, but only cyclic amine end groups favour duplex-quadruplex transitions that are likely crucial to produce biological and pharmacological effects in living systems.

Antitumor AZA-anthrapyrazoles: biophysical and biochemical studies on 8- and 9-aza regioisomers.

Aza-bioisosteres of anthrapyrazoles (Aza-APs) bearing the C-N substitution at position 9 are powerful anticancer agents now in clinical trials. In contrast, their 8-substituted regioisomers are practically devoid of chemotherapeutic effects. To understand the molecular basis for a dramatically different response by otherwise very similar compounds, we performed a detailed investigation on the physico-chemical properties of several aza-APs belonging to the two families, on their DNA-binding affinity and specificity as well as on their capacity to impair the activity of the two isoforms of human Topoisomerase II (top2alpha and top2beta). Our results indicate that molecular size and shape, electronic distribution, redox properties, lipophilicity and protonation equilibria are essentially the same when comparing 9- with 8-substituted congeners. Although no major difference could be picked up when comparing the DNA binding properties of corresponding members of the 8- and 9-aza families, interestingly the affinity and specificity for the nucleic acid is modulated by the nature of the side-arms linked to the aza-AP scaffold, suggesting structural motifs that may determine DNA sequence recognition by the studied drug. Topoisomerase II poisoning activity was much higher for 9-aza derivatives than 8-aza analogues as shown by a cleavage assay with purified recombinant top2 isoforms. The difference appears to account for the divergent anticancer potential exhibited by different aza-AP regioisomers and suggests a specific molecular recognition of the cleavage complex by the studied drugs.

Characterization of anthracenediones and their photoaffinity analogs.

In an attempt to overcome the cardiotoxicity and cross-resistance problems caused by the anticancer drugs anthracyclines and anthracenediones during chemotherapy, we have developed a series of aza-anthracenedione compounds by modifying the chromophore and the side arms of anthracyclines and anthracenediones. One of these aza-anthracenediones, 6,9-bis[(2-aminoethyl)amino]benzo[g]isoquinoline-5,10-dione (BBR 2778), which is currently under phase II clinical trials, showed remarkable antitumor activity and appeared to lack a cardiotoxic effect in preclinical studies. However, it was still cross-resistant against multidrug resistance (MDR) cells expressing P-glycoprotein (P-gp). In contrast, another aza-anthracenedione, 6,9-bis[[2-(dimethylamino)ethyl]amino]benzo[g]isoquinoline-5,10-dione, which has side arm structures different from those of BBR 2778, was highly active against MDR cells. In this study, BBR 2778, BBR 2378, and an anthracenedione compound, 1,4-bis[(2-aminoethyl)amino]-5,8-dimethyl-9,10-anthracenedione, were used to assess the relationship between the chemical structures of these drugs and their interactions with DNA and P-gp. In addition, the biological and pharmacological influences of photoaffinity labeling were also studied for BBR 2778 and DEH. As the results indicate, the photolabeled analogs of BBR 2778 and DEH were less DNA-reactive and less cytotoxic. The more lipophilic compound, BBR 2378, and the photolabeled analogs of BBR 2778 and DEH inhibited P-gp labeling by azidopine better than did the more hydrophilic parental compounds. These studies suggested that the DNA binding affinity of BBR 2778 and DEH could be important in determining their cytotoxicity, and that the chemical structure of the side arms and the lipophilicity of these drugs are critical in determining their cross-resistance.