The human telomeric sequence d[T(2)AG(3)](4) has been demonstrated to form different types of G-quadruplex structures, depending upon the incubation conditions. For example, in sodium (Na(+)), a basket-type G-quadruplex structure is formed. In this investigation, using circular dichroism (CD), biosensor-surface plasmon resonance (SPR), and a polymerase stop assay, we have examined how the addition of different G-quadruplex-binding ligands affects the conformation of the telomeric G-quadruplex found in solution. The results show that while telomestatin binds preferentially to the basket-type G-quadruplex structure with a 2:1 stoichiometry, 5,10,15,20-[tetra-(N-methyl-3-pyridyl)]-26-28-diselena sapphyrin chloride (Se2SAP) binds to a different form with a 1:1 stoichiometry in potassium (K(+)). CD studies suggest that Se2SAP binds to a hybrid G-quadruplex that has strong parallel and antiparallel characteristics, suggestive of a structure containing both propeller and lateral, or edgewise, loops. Telomestatin is unique in that it can induce the formation of the basket-type G-quadruplex from a random coil human telomeric oligonucleotide, even in the absence of added monovalent cations such as K(+) or Na(+). In contrast, in the presence of K(+), Se2SAP was found to convert the preformed basket G-quadruplex to the hybrid structure. The significance of these results is that the presence of different ligands can determine the type of telomeric G-quadruplex structures formed in solution. Thus, the biochemical and biological consequences of binding of ligands to G-quadruplex structures found in telomeres and promoter regions of certain important oncogenes go beyond mere stabilization of these structures.

Cationic porphyrins are known to bind to and stabilize different types of G-quadruplexes. Recent studies have shown the biological relevance of the intramolecular parallel G-quadruplex as a transcriptional silencer in the c-MYC promoter. TMPyP4 also binds to this G-quadruplex and most likely converts it to a mixed parallel/antiparallel G-quadruplex with two external lateral loops and one internal propeller loop, suppressing c-MYC transcriptional activation. To achieve therapeutic selectivity by targeting G-quadruplexes, it is necessary to synthesize drugs that can differentiate among the different types of G-quadruplexes. We have designed and synthesized a core-modified expanded porphyrin analogue, 5,10,15,20-[tetra(N-methyl-3-pyridyl)]-26,28-diselenasapphyrin chloride (Se2SAP). Se2SAP converts the parallel c-MYC G-quadruplex into a mixed parallel/antiparallel G-quadruplex with one external lateral loop and two internal propeller loops, resulting in strong and selective binding to this G-quadruplex. A Taq polymerase stop assay was used to evaluate the binding of TMPyP4 and Se2SAP to G-quadruplex DNA. Compared to TMPyP4, Se2SAP shows a greater selectivity for and a 40-fold increase in stabilization of the single lateral-loop hybrid. Surface plasmon resonance and competition experiments with duplex DNA and other G-quadruplexes further confirmed the selectivity of Se2SAP for the c-MYC G-quadruplex. Significantly, Se2SAP was found to be less photoactive and noncytotoxic in comparison to TMPyP4. From this study, we have identified an expanded porphyrin that selectively binds with the c-MYC G-quadruplex in the presence of duplex DNA and other G-quadruplexes.

The synthesis, spectral and structural characterization of meso-aryl sapphyrins and rubyrins containing heteroatoms such as S, O, Se in addition to pyrrole nitrogens are reported. The synthesis of the desired expanded porphyrins has been achieved using a single precursor, the modified tripyrranes containing heteroatoms, through an unprecedented oxidative coupling reaction in moderately good yields. The product distribution and the isolated yields were found to be dependent on the nature of the acid catalyst and its concentration. Use of 0.1 equiv of acid exclusively gave 26π rubyrins while a higher concentration of acid gave a mixture of 18π porphyrin, 22π sapphyrin, and 26π rubyrin. Two additional products, 22π oxasmaragdyrin and 18π oxacorrole, were isolated in the reaction of oxatripyrrane. All of the sapphyrins and rubyrins exhibit well-defined intense Soret and Q-bands in the visible region, and the intensity and the position of the absorption maxima were dependent on the number and the nature of the heteroatoms present in the cavity. The solid-state structures of sapphyrins 8 and 9 show small deviations from planarity with formation of supramolecular ladders stabilized by weak C−H···S, C−H···Se, and C−H···N hydrogen bonds. 1H NMR studies reveal retainment of supramolecular arrays in solution. The TFA adduct of 8 shows unusual binding in which both the hydroxyl oxygen and the carbonyl oxygen participate, which is reminiscent of metal carboxylate binding and in total contrast to that observed for β-substituted sapphyrins. 1H NMR studies on rubyrins indicate rapid rotation of heterocyclic rings at room temperature, and protonation leads to a decrease in rate of rotation at room temperature. 1H NMR spectra of 10 and 17 in its free base form recorded at −50 °C reveal that the heterocyclic rings are inverted and protonation leads to dramatic ring flipping. However, 11 shows normal structure in the solution. The single-crystal X-ray structures of 10, 11, and 17 show that the heterocyclic rings, thiophene in 10, selenophene in 11, and furan and thiophene in 17, are inverted in the solid state.