Development and application of metallo-phthalocyanines as potent G-quadruplex DNA binders and photosensitizers.

Metallo-phthalocyanines (MPc) are common photosensitizers with ideal photophysical and photochemical properties. Also, these molecules have shown to interact with non-canonical nucleic acid structures, such as G-quadruplexes, and modulate oncogenic expression in cancer cells. Herein, we report the synthesis and characterisation of two metallo-phthalocyanines containing either zinc (ZnPc) or nickel (NiPc) in the central aromatic core and four alkyl ammonium lateral chains. The interaction of both molecules with G-quadruplex DNA was assessed by UV-Vis, fluorescence and FRET melting experiments. Both molecules bind strongly to G-quadruplexes and stabilise these structures, being NiPc the most notable G-quadruplex stabiliser. In addition, the photosensitizing ability of both metal complexes was explored by the evaluation of the singlet oxygen generation and their photoactivation in cells. Only ZnPc showed a high singlet oxygen generation either by direct observation or by indirect evaluation using a DPBF dye. The cellular evaluation showed mainly cytoplasmic localization of ZnPc and a decrease of the IC50 values of the cell viability of ZnPc upon light activation of two orders of magnitude. Two metallo-phthalocyanines containing zinc and nickel within the aromatic core have been investigated as G-quadruplex stabilizers and photosensitizers. NiPc shows a high G4 binding but negligible photosensitizing ability while ZnPc exhibits a moderate binding to G-quadruplex together with a high potency to generate singlet oxygen and photocytotoxicity. The interaction with G4s and capacity to be photosensitized is associated with the geometry adopted by the central metal core of the phthalocyanine scaffold.

Impact of the zinc complexation of polytopic polyaza ligands on the interaction with double and single stranded DNA/RNA and antimicrobial activity.

Metal complexes have gained a huge interest in the biomedical research in the last decade because of the access to unexplored chemical space with regards to organic molecules and to present additional functionalities to act simultaneously as diagnostic and therapeutic agents. Herein, we evaluated the interaction of two polytopic polyaza ligands and their zinc complexes with DNA and RNA by UV thermal denaturation, fluorescence and circular dichroism spectroscopic assays. The zinc coordination was investigated by X-ray diffraction and afforded the structure of the binuclear zinc complex of PYPOD. Thermal denaturation of DNA and RNA and fluorimetry analysis revealed preferential binding of the zinc-PHENPOD complexes towards GC-containing DNA in contrast to the free ligands. On the other hand, PYPOD metal complexes, compared to the free ligand, stabilized AT-based DNA (B-form) better than AU-RNA (A-form). With regards to single stranded RNA, the binuclear complex of PHENPOD and the free ligand can efficiently identify polyadenylic acid (poly A) among other RNA sequences by circular dichroism spectroscopy. The antimicrobial activity in S. aureus and E. coli bacteria showed the highest activity for the free ligands and their trinuclear zinc complexes. This work can provide valuable insights into the impact of the nuclearity of polytopic polyaza ligands in the binding to DNA/RNA and the antimicrobial effect.

Modulating the G-Quadruplex and Duplex DNA Binding by Controlling the Charge of Fluorescent Molecules.

Two fluorescent and non-toxic spirobifluorene molecules bearing either positive (Spiro-NMe3) or negative (Spiro-SO3) charged moieties attached to the same aromatic structure have been investigated as binders for DNA. The novel Spiro-NMe3 containing four alkylammonium substituents interacts with G-quadruplex (G4) DNA structures and shows preference for G4s over duplex by means of FRET melting and fluorescence experiments. The interaction is governed by the charged substituents of the ligands as deduced from the lower binding of the sulfonate analogue (Spiro-SO3). On the contrary, Spiro-SO3 exhibits higher binding affinity to duplex DNA structure than to G4. Both molecules show a moderate quenching of the fluorescence upon DNA binding. The confocal microscopy evaluation shows the internalization of both molecules in HeLa cells and their lysosomal accumulation.

Alkaloids as Photosensitisers for the Inactivation of Bacteria.

Antimicrobial photodynamic therapy has emerged as a powerful approach to tackle microbial infections. Photodynamic therapy utilises a photosensitiser, light, and oxygen to generate singlet oxygen and/or reactive oxygen species in an irradiated tissue spot, which subsequently react with nearby biomolecules and destroy the cellular environment. Due to the possibility to irradiate in a very precise location, it can be used to eradicate bacteria, fungus, and parasites upon light activation of the photosensitiser. In this regard, natural products are low-cost molecules capable of being obtained in large quantities, and some of them can be used as photosensitisers. Alkaloids are the largest family among natural products and include molecules with a basic nature and aromatic rings. For this study, we collected the naturally occurring alkaloids used to treat microorganism infections using a photodynamic inactivation approach. We gathered their main photophysical properties (excitation/emission wavelengths, quantum yields, and oxygen quantum yield) which characterise the ability to efficiently photosensitise. In addition, we described the antibacterial activity of alkaloids upon irradiation and the mechanisms involved in the microorganism killing. This review will serve as a reference source to obtain the main information on alkaloids used in antimicrobial photodynamic therapy.

Triazolopyridyl ketones as a novel class of antileishmanial agents. DNA binding and BSA interaction.

A new series of triazolopyridyl pyridyl ketones has been synthetized by regioselective lithiation of the corresponding [1,2,3]triazolo[1,5-a]pyridine at 7 position followed by reaction with different electrophiles. The in vitro antileishmanial activity of these compounds was evaluated against Leishmaniainfantum, Leishmaniabraziliensis, Leishmaniaguyanensis and Leishmaniaamazonensis. Compounds 6 and 7 were found to be the most active leishmanicidal agents. Both of them showed activities at micromolar concentration against cultured promastigotes of Leishmania spp. (IC50=99.8-26.8 µM), without cytotoxicity on J774 macrophage cells. These two compounds were also tested in vivo in a murine model of acute infection by L. infantum. The triazolopyridine derivative 6 was effective against both spleen and liver parasites forms, while 7 was inactive against liver parasites. Mechanistic aspects of the antileishmanial activity were investigated by means of DNA binding studies (UV-titration and viscosimetry). Results have revealed that these active ligands are able to interact strongly with DNA [Kb=1.14 × 10(5)M(-1) (6) and 3.26 × 10(5)M(-1) (7)]. Moreover, a DNA groove binding has been proposed for both 6 and 7. To provide more insight on the mode of action of compounds 6 and 7 under biological conditions, their interaction with bovine serum albumin (BSA) was monitored by fluorescence titrations and UV-visible spectroscopy. The quenching constants and binding parameters were determined. Triazolopyridine ketones 6 and 7 have exhibited significant affinity towards BSA [Kb=2.5 × 10(4)M(-1) (6) and 1.9 × 10(4)M(-1) (7)]. Finally, to identify the binding location of compounds 6 and 7 on the BSA, competitive binding experiments were carried out, using warfarin, a characteristic marker for site I, and ibuprofen as one for site II. Results derived from these studies have indicated that both compounds interact at BSA site I and, to a lesser extent, at site II.