Bimetallic nanoparticles enhance photoactivity of conjugated photosensitizer.

Although photodynamic therapy (PDT) of cancer has been continuously improved, its efficiency is still limited by the high toxicity in the absence of irradiation, aggregation and deactivation by biomolecules of the most common photosensitizers (PS). The association of PS to nanoparticles (NPs) can be a promising tool to overcome these limitations and also to enhance PS tumoral selectivity. In addition, the association of PS to metallic NPs may provide the modulation of PS fluorescence and also the enhancement of PS photoactivity due to the electronic coupling with NPs plasmon effect. Adversely to the innumerous work on the coupling of PS to metallic NPs, the application of bimetallic NPs with this goal has not been explored yet. In this work we investigated the physicochemical properties and cytotoxicity of bimetallic gold-platinum NPs (AuPtNPs) conjugated to a chlorin molecule modified with a thiol group. Additionally, chlorin was coupled to AuNPs for comparative purposes since these have been the most commonly used NPs in PDT. The results showed that both platforms promoted the chlorin solubility in water which is crucial in biological applications. Despite the enhancement of photoactivity promoted by both NPs in comparison with chlorin in solution, chlorin-conjugated with AuPtNPs proved to be a more suitable platform for PDT application, since it showed a lower dark citotoxicity, as well as a higher generation of singlet oxygen and cell internalization compared with chlorin-conjugated AuNPs. It is important to highlight that this is the first work reporting on the enhancement of PS photoactivity by its conjugation to AuPtNPs.

A new Chlorin formulation promotes efficient photodynamic action in choriocapillaris of rabbit's eyes.

Age-related macular degeneration (AMD) as well as other choroidal diseases, demand novel therapeutic methods. Photodynamic therapy (PDT), which uses light and photosensitizer (PS) to cause specific vascular occlusion in the macula, is an interesting alternative. The only drug approved for the PDT treatment of AMD (Verteporfin) has a natural tendency to aggregate, demanding an expensive separation procedure during purification. We report a novel and affordable PS that is intrinsically protected against aggregation, the Monomeric Chlorin at High Concentration (MCHC-Chlorin), whose liposomal formulation was developed to provoke effective photodynamic action on the choroidal vasculature. Our report starts by stablishing the conditions to allow the efficient synthesis of MCHC-Chlorin in high yields (92%). We then tested the light stimulated occlusion of choriocapillary vessels in rabbit's eyes induced by the two MCHC-Chlorin isomers, which are directly obtained from the synthetic route. The PS formulation was infused in the rabbit's ear vein and eyes were immediately irradiated at 650 nm. Indirect ophthalmoscopy, fundus photography, fluorescein angiography and histopathological evaluations were used to evaluate levels of photo-thrombosis and collateral damage. Choriocapillary occlusion was achieved in all treated rabbits' eyes, while retina and sclera were completely preserved. There was no photochemical reaction in none of the eyes that received LASER without PS. Both MCHC-Chlorin isomers were separately tested and exhibited similar positive results with no systemic toxicity. Therefore, PDT occurred equally well in all treated eyes and none of the controls showed any effect in the ophthalmological exams. MCHC-Chlorin offers great potential and should be further studied as an alternative drug for choroidal diseases.

Photo-activated phase separation in giant vesicles made from different lipid mixtures.

Using giant unilamellar vesicles (GUVs) made from POPC, DPPC, cholesterol and a small amount of a porphyrin-based photosensitizer that we name PE-porph, we investigated the response of the lipid bilayer under visible light, focusing in the formation of domains during the lipid oxidation induced by singlet oxygen. This reactive species is generated by light excitation of PE-porf in the vicinity of the membrane, and thus promotes formation of hydroperoxides when unsaturated lipids and cholesterol are present. Using optical microscopy we determined the lipid compositions under which GUVs initially in the homogeneous phase displayed Lo-Ld phase separation following irradiation. Such an effect is attributed to the in situ formation of both hydroperoxized POPC and cholesterol. The boundary line separating homogeneous Lo phase and phase coexistence regions in the phase diagram is displaced vertically towards the higher cholesterol content in respect to ternary diagram of POPC:DPPC:cholesterol mixtures in the absence of oxidized species. Phase separated domains emerge from sub-micrometer initial sizes to evolve over hours into large Lo-Ld domains completely separated in the lipid membrane. This study provides not only a new tool to explore the kinetics of domain formation in mixtures of lipid membranes, but may also have implications in biological signaling of redox misbalance.

Synthesis of phthalocyanines-ALA conjugates: water-soluble compounds with low aggregation.

Syntheses of two water-soluble phthalocyanines (Pc) containing 5-aminolevulinic acid (ALA) linked to the core structure are described. These compounds were prepared by using original functionalizations, and they present remarkable structural and photophysical features, indicating that they could be applied to photodynamic therapy (PDT).

Antimicrobial photodynamic therapy combined to periodontal treatment: Experimental model.

BACKGROUND: Antimicrobial photodynamic therapy (aPDT) has been used as an adjuvant treatment for periodontitis. It combines a photosensitizer with a light source to induce reactive oxygen species and kill microbial cells. PpNetNI is a protoporphyrin derivative, and it has a chemical binding site at biofilm and great affinity to microbial cells. The aim of this study was to investigate the effects of aPDT as an adjuvant treatment for periodontitis. METHODS: Thirty healthy male rats Wistar (Rattus norvegicus) were used in this study (Approved by UNINOVE Ethical committee AN0029/2015). Periodontitis was induced by placing a cotton ligature around the first mandibular molar in a subgengival position. The contralateral mandibular first molar received neither a ligature nor any treatment, and was used as a control. After 7 days, the ligature was removed and all animals received scaling and root planing (SRP) and were divided according to the following treatments: SRP group (received SRP and irrigation with PpNetNI, 10µM) and aPDT group (PpNetNI 10µM followed by LED irradiation). aPDT was performed with a LED (630nm) with an output power of 400mW (fluence-rate 200mW/cm2; fluence 18J/cm2). Rats were euthanized at 24h, 48h and 7days postoperatively. The area of bone loss in vestibular region of the first molar was evaluated by Optical Coherence Tomography (OCT, THORLABS LTD., Ely, UK). Data were analyzed statistically (ANOVA and Tukey tests, p<0.05). RESULTS: The animals treated by aPDT showed bone gain of approximately 30% compared to the SRP group following 7days from the treatment. CONCLUSION: aPDT promoted bone recovery 7days after periodontal intervention.

Protoporphyrin IX nanoparticle carrier: preparation, optical properties, and singlet oxygen generation.

The present study is focused on developing a nanoparticle carrier for the photosensitizer protoporphyrin IX for use in photodynamic therapy. The entrapment of protoporphyrin IX (Pp IX) in silica spheres was achieved by modification of Pp IX molecules with an organosilane reagent. The immobilized drug preserved its optical properties and the capacity to generate singlet oxygen, which was detected by a direct method from its characteristic phosphorescence decay curve at near-infrared and by a chemical method using 1,3-diphenylisobenzofuran to trap singlet oxygen. The lifetime of singlet oxygen when a suspension of Pp IX-loaded particles in acetonitrile was excited at 532 nm was determined as 52 micros, which is in good agreement with the value determined for methylene blue in acetonitrile solution under the same conditions. The Pp IX-loaded silica particles have an efficiency of singlet oxygen generation (eta Delta) higher than the quantum yield of free porphyrins. This high efficiency of singlet oxygen generation was attributed to changes on the monomer-dimer equilibrium after photosentisizer immobilization.