As functionalized chitosans hold great potential for the development of effective and broad-spectrum antibiotics, representative chitosan derivatives were tested for antimicrobial activity in neutral media: trimethyl chitosan (TMC), carboxy-methyl chitosan (CMC), and chitosan-thioglycolic acid (TGA; medium molecular weight: MMW-TGA; low molecular weight: LMW-TGA). Colony forming assays indicated that LMW-TGA displayed superior antimicrobial activity over the other derivatives tested: a 30 min incubation killed 100% Streptococcus sobrinus (Gram-positive bacteria) and reduced colony counts by 99.99% in Neisseria subflava (Gram-negative bacteria) and 99.97% in Candida albicans (fungi). To elucidate LMW-TGA effects at the cellular level, microscopic studies were performed. Use of fluorescein isothiocyanate (FITC)-labeled chitosan derivates in confocal microscopy showed that LMW-TGA attaches to microbial cell walls, while transmission electron microscopy indicated that this derivative severely affects cell wall integrity and intracellular ultrastructure in all species tested. We therefore propose LMW-TGA as a promising and effective broad-band antimicrobial compound.

A fluorescent labeled Wells-Dawson type POM ({P2W17O61Fluo}) was newly synthesized and characterized by a wide range of analytical methods. {P2W17O61Fluo} was functionalized with fluorescein amine through a stable amide bond, and its long time stability was verified by UV/vis spectroscopic techniques at physiologically relevant pH values. No significant impact on the cell viability or morphology of HeLa cells was observed for POM concentrations up to 100 μg mL(-1). Cellular uptake of fluorescent {P2W17O61Fluo} was monitored by confocal laser scanning microscopy. POM uptake occurs mainly after prolonged incubation times of 24 h resulting in different intracellular patterns, i.e. randomly distributed over the entire cytoplasm, or aggregated in larger clusters. This direct monitoring strategy for the interaction of POMs with cells opens up new pathways for elucidating their unknown mode of action on the way to POM-based drug development.

BackgroundPhotosensitizers are used in photodynamic therapy (PDT) to destruct tumor cells, however, their limited solubility and specificity hampers routine use, which may be overcome by encapsulation. Several promising novel nanoparticulate drug carriers including liposomes, polymeric nanoparticles, metallic nanoparticles and lipid nanocomposites have been developed. However, many of them contain components that would not meet safety standards of regulatory bodies and due to difficulties of the manufacturing processes, reproducibility and scale up procedures these drugs may eventually not reach the clinics. Recently, we have designed a novel lipid nanostructured carrier, namely Lipidots, consisting of nontoxic and FDA approved ingredients as promising vehicle for the approved photosensitizer m-tetrahydroxyphenylchlorin (mTHPC).ResultsIn this study we tested Lipidots of two different sizes (50 and 120 nm) and assessed their photodynamic potential in 3-dimensional multicellular cancer spheroids. Microscopically, the intracellular accumulation kinetics of mTHPC were retarded after encapsulation. However, after activation mTHPC entrapped into 50 nm particles destroyed cancer spheroids as efficiently as the free drug. Cell death and gene expression studies provide evidence that encapsulation may lead to different cell killing modes in PDT.ConclusionsSince ATP viability assays showed that the carriers were nontoxic and that encapsulation reduced dark toxicity of mTHPC we conclude that our 50 nm photosensitizer carriers may be beneficial for clinical PDT applications.Electronic supplementary materialThe online version of this article (doi:10.1186/s12951-016-0221-x) contains supplementary material, which is available to authorized users.

Novel tetraplatinated metalloporphyrin-based photosensitizers (PSs) show excellent phototoxic indexes up to 5800 against HeLa cells, which is, to the best of our knowledge, the highest value reported for any porphyrin...

BackgroundPhotodynamic therapy (PDT) as promising alternative to conventional cancer treatments works by irradiation of a photosensitizer (PS) with light, which creates reactive oxygen species and singlet oxygen (1O2), that damage the tumor. However, a routine use is hindered by the PS’s poor water solubility and extended cutaneous photosensitivity of patients after treatment. In our study we sought to overcome these limitations by encapsulation of the PS m-tetrahydroxyphenylchlorin (mTHPC) into a biocompatible nanoemulsion (Lipidots).ResultsIn CAL-33 tumor bearing nude mice we compared the Lipidots to the existing liposomal mTHPC nanoformulation Foslip and the approved mTHPC formulation Foscan. We established biodistribution profiles via fluorescence measurements in vivo and high performance liquid chromatography (HPLC) analysis. All formulations accumulated in the tumors and we could determine the optimum treatment time point for each substance (8 h for mTHPC, 24 h for Foslip and 72 h for the Lipidots). We used two different light doses (10  and 20 J/cm2) and evaluated immediate PDT effects 48 h after treatment and long term effects 14 days later. We also analyzed tumors by histological analysis and performing reverse transcription real-time PCR with RNA extracts. Concerning tumor destruction Foslip was superior to Lipidots and Foscan while with regard to tolerance and side effects Lipidots were giving the best results.ConclusionsWe could demonstrate in our study that nanoformulations are superior to the free PS mTHPC. The development of a potent nanoformulation is of major importance because the free PS is related to several issues such as poor bioavailability, solubility and increased photosensibility of patients. We could show in this study that Foslip is very potent in destroying the tumors itself. However, because the Lipidots' biocompatibility is outstanding and superior to the liposomes we plan to carry out further investigations and protocol optimization. Both nanoformulations show great potential to revolutionize PDT in the future.Electronic supplementary materialThe online version of this article (doi:10.1186/s12951-016-0223-8) contains supplementary material, which is available to authorized users.

We report the synthesis of novel tetraplatinated metalloporphyrin-based photosensitizers (PSs) for photodynamic therapy (PDT), their characterization, cellular uptake and localization, as well as the determination of their <i>in vitro</i> light-induced anticancer properties. The PSs show excellent phototoxic indexes up to 5800 against HeLa cells, which is, to the best of our knowledge, the highest value reported for any porphyrin so far. Furthermore, isotopic labelling of the porphyrin with a highly enriched ⁶⁷Zn isotope was performed in order to determine the distribution ratio of zinc to platinum by ICP-MS, allowing to differentiate between naturally occurring zinc and ⁶⁷Zn that was introduced into the cells by the PS. We conclude that the platinum units within the platinum-PS conjugates help to solubilize the PS and, at the same time, act as cell-penetrating vectors, enhancing the efficiency of the PS without causing a significant dark toxicity.<br>

We report the synthesis of novel tetraplatinated metalloporphyrin-based photosensitizers (PSs) for photodynamic therapy (PDT), their characterization, cellular uptake and localization, as well as the determination of their <i>in vitro</i> light-induced anticancer properties. The PSs show excellent phototoxic indexes up to 5800 against HeLa cells, which is, to the best of our knowledge, the highest value reported for any porphyrin so far. Furthermore, isotopic labelling of the porphyrin with a highly enriched ⁶⁷Zn isotope was performed in order to determine the distribution ratio of zinc to platinum by ICP-MS, allowing to differentiate between naturally occurring zinc and ⁶⁷Zn that was introduced into the cells by the PS. We conclude that the platinum units within the platinum-PS conjugates help to solubilize the PS and, at the same time, act as cell-penetrating vectors, enhancing the efficiency of the PS without causing a significant dark toxicity.<br>

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