Photodynamic Inactivation of the Sarcoplasmic Reticulum Ca2+-ATPase by a Ruthenium Polypyridyl Complex Featuring π-Expansive Ligands

Sarcoplasmic reticulum Ca2+-ATPase (SERCA) is a key protein involved in calcium homeostasis and the pivotal role of Ca2+ in a plethora of biological activities makes SERCA a powerful biological target. The intertwining between Ca2+ signalling and reactive oxygen species (ROS) has drawn much attention to investigate the photodynamic effect that photosensitizers (PSs), prodrugs that are light-activated to produce ROS in photodynamic therapy (PDT), pursue on this important target. However, only limited classes of PSs were considered so far, with most of them being porphyrin-based scaffolds, without considering, to the best of our knowledge, the important family of ruthenium(II) polypyridyl complexes (RPCs), which still attracts increasing interest in PDT thanks to its unique and versatile chemical-physical repertoire. With the goal of exploring the potential arising from considering SERCA a possible target for highly effective RPC-based PSs, herein we studied the photodynamic effect on this protein of [Ru(dppn)2(bpy-morph)](PF6)2 (RuPS) (dppn = benzo[i]dipyrido[3,2-a:2′,3′-c]phenazine, bpy-morph = 2,2′-bipyridine-4,4′-diylbis(morpholinomethanone), where two π-expansive dppn units guarantees a high 1O2 sensitization (ϕΔ = 0.54 ± 0.06 in CH3CN). To this aim, the solid supported membrane (SSM) biosensing method was employed, equipped with a LED light (λem = 434 nm, 30 W) to perform irradiation experiments. Light activation of RuPS (10 μM) induced a prompt decrease (30%) of Ca2+ translocation by SERCA, leading to the almost complete loss of residual current after 15 min irradiation (more than 90% decrease). The inhibition of SERCA was then evaluated in vivo in C2C12 myotubes, chosen as a model in skeletal muscle biology. Upon establishing cellular internalization via confocal microscopy, light-activated RuPS was found to enhance the cytosolic calcium concentration up to 1.5-fold-higher than thapsigargin, used as positive control. Moreover, the differentiated phenotype of myotubes was reverted by light-activated RuPS, an effect which was likely related to the impaired activity of this key protein.