• A dinuclear ruthenium(II) complex excited by near-infrared light through two-photon absorption induces phototoxicity deep within hypoxic regions of melanoma cancer spheroids

      Raza, Ahtasham; Archer, Stuart A; Fairbanks, Simon D; Smitten, Kirsty L.; Botchway, Stanley W.; Thomas, Jim A; MacNeil, Sheila; Haycock, John W.; The University of Derby (American Chemical Society (ACS), 2020-02-17)
      The dinuclear photo-oxidizing RuII complex [{Ru(TAP2)}2(tpphz)]4+ (TAP = 1,4,5,8- tetraazaphenanthrene, tpphz = tetrapyrido[3,2-a:2′,3′-c:3′′,2′′- h:2′′′,3′′′-j]phenazine), 14+ is readily taken up by live cells localizing in mitochondria and nuclei. In this study, the two-photon absorption cross-section of 14+ is quantified and its use as a two-photon absorbing phototherapeutic is reported. It was confirmed that the complex is readily photo-excited using near infrared, NIR, light through two-photon absorption, TPA. In 2-D cell cultures, irradiation with NIR light at low power results in precisely focused photo-toxicity effects in which human melanoma cells were killed after 5 minutes of light exposure. Similar experiments were then carried out in human cancer spheroids that provide a realistic tumor model for the development of therapeutics and phototherapeutics. Using the characteristic emission of the complex as a probe, its uptake into 280 µm spheroids was investigated and confirmed that the spheroid takes up the complex. Notably TPA excitation results in more intense luminescence being observed throughout the depth of the spheroids, although emission intensity still drops off toward the necrotic core. As 14+ can directly photo-oxidize DNA without the mediation of singlet oxygen or other reactive oxygen species, photo-toxicity within the deeper, hypoxic layers of the spheroids was also investigated. To quantify the penetration of these phototoxic effects, 14+ was photo-excited through TPA at a power of 60 mW, which was progressively focused in 10 µm steps throughout the entire z-axis of individual spheroids. These experiments revealed that, in irradiated spheroids treated with 14+, acute and rapid photo-induced cell death was observed throughout their depth, including the hypoxic region.
    • A dinuclear ruthenium(II) phototherapeutic that targets duplex and quadruplex DNA

      Archer, Stuart; Raza, Ahtasham; Dröge, Fabian; Robertson, Craig C.; Auty, Alexander J; Chekulaev, Dimitri M; Weinstein, Julia; Keane, Theo; Meijer, Anthony J. H. M.; Haycock, John; et al. (Royal Society of Chemistry, 2019-02-18)
      With the aim of developing a sensitizer for photodynamic therapy, a previously reported luminescent dinuclear complex that functions as a DNA probe in live cells was modified to produce a new isostructural derivative containing RuII(TAP)2 fragments (TAP = 1,4,5,8- tetraazaphenanthrene). The structure of the new complex has been confirmed by a variety of techniques including single crystal X-ray analysis. Unlike its parent, the new complex displays RuL-based 3MLCT emission in both MeCN and water. Results from electrochemical studies and emission quenching experiments involving guanosine monophosphate are consistent with an excited state located on a TAP moiety. This hypothesis is further supported by detailed DFT calculations, which take into account solvent effects on excited state dynamics. Cell-free steady-state and time-resolved optical studies on the interaction of the new complex with duplex and quadruplex DNA show that the complex binds with high affinity to both structures and indicate that its photoexcited state is also quenched by DNA, a process that is accompanied by the generation of the guanine radical cation sites as photo-oxidization products. Like the parent complex, this new compound is taken up by live cells where it primarily localizes within the nucleus and displays low cytotoxicity in the absence of light. However, in complete contrast to [{RuII(phen)2}2(tpphz)]4+, the new complex is therapeutically activated by light to become highly phototoxic toward malignant human melanoma cell line showing that it is a promising lead for the treatment of this recalcitrant cancer.