Search

Photodynamic therapy (PDT) is a clinical, minimally invasive method for destroying cancer cells in the presence of a photosensitizer, oxygen, and a light source. The main obstacle for the PDT treatment of deep tumors is a strong reduction of the excitation light intensity as a result of its refraction, reflection, and absorption by biological tissues. Internal light sources based on bioluminescence resonance energy transfer can be a solution of this problem. Here we show that luciferase NanoLuc being expressed as a fusion protein with phototoxic flavoprotein miniSOG in cancer cells in the presence of furimazine (highly specific NanoLuc substrate) induces a photodynamic effect of miniSOG comparable with its LED-excited (Light Emitting Diode) phototoxicity. Luminescence systems based on furimazine and hybrid protein NanoLuc-miniSOG targeted to mitochondria or cellular membranes possess the similar energy transfer efficiencies and similar BRET-induced cytotoxic effects on cancer cells, though the mechanisms of BRET-induced cell death are different. As the main components of the proposed system for BRET-mediated PDT are genetically encoded (luciferase and phototoxic protein), this system can potentially be delivered to any site in the organism and thus may be considered as a promising approach for simultaneous delivery of light source and photosensitizer in deep-lying tumors and metastasis anywhere in the body.

Despite an enhanced permeability and retention effect typical of many solid tumors, drug penetration is not always sufficient. Possible strategies for the drug delivery improvement are a modification of the tumor cell-to-cell junctions and usage of cell membrane permeabilization proteins. In this review we discuss epithelial cell junctions as targets for a combined anticancer therapy and propose new possible sources of such agents. We suggest considering viral and bacterial pathogens disrupting epithelial layers as plentiful sources of new therapeutic agents for increasing tumor permeability for other effector agents. We also observe the application of pore forming proteins and peptides of different origin for cytoplasmic delivery of anti-cancer agents and consider the main obstacles of their use in vivo.

Cytotoxic effects of a new targeted phototoxin DARPin-miniSOG and mechanism of its action were investigated in vitro. It was determined that DARPin-miniSOG causes light-induced death of HER2/neu-positive cancer cells (IC50 0.8 μM). Treatment of the cells with DARPin-miniSOG in the presence of ascorbic acid eliminated the light-induced cytotoxic action of the protein. This observation suggests the involvement of oxidative stress in the mechanism of the phototoxin action. DNA fragmentation analysis, caspase-3 activity assay and PI-staining of HER2/neu-positive cancer cells treated with DARPin-miniSOG indicated that phototoxin induces necrotic cell death under blue light illumination. Co-localization analysis showed that DARPin-miniSOG accumulates mostly in endosomes and lysosomes.