Zinc Phthalocyanine loaded polymeric micelles for photodynamic therapy

Photodynamic therapy (PDT) is a promising alternative to currently employed cancer treatments [1]. However, the high hydrophobicity of the photosensitizers (e.g. zinc phthalocyanine (ZnPC)) causes its aggregation in biological fluids. Moreover, unspecific accumulation in healthy tissues such as the skin, and low clearance rate of ZnPC, leads to prolong skin sensitization, forcing patients with a short life expectancy to remain inside and shielded from light. Consequently, clinical implementation is still limited. In this study, poly(caprolactone)n-poly(ethylene glycol) (pCL-PEG) micelles encapsulating ZnPC are being investigated to increase the solubility of ZnPC and its accumulation at the tumor site. pCLn-PEG (n=27) was synthesized by ring-opening polymerization following the protocol from Y. Liu et al. [2]. pLC-PEG micelles encapsulating ZnPC (ZnPC-M) at different drug loadings (0.2, 0.4 and 0.6 % w/w) were prepared by film hydration. Z-average hydrodynamic diameter and size distribution were measured by dynamic light scattering. In vitro stability and ZnPC transfer to human proteins and lipoproteins was compared between ZnPC-M with different drug loadings using asymmetric flow field flow fractionation after incubation of ZnPC-M with human plasma (1:2 v/v) for different timepoints. Phototoxicity of ZnPC-M was tested in vitro by exposing TFK1 (both in 2D and 3D) to a range of ZnPC concentrations. After incubation and washing, the cells were illuminated with a specific light intensity using a LED device and subsequently, further incubated overnight. The cell viability was measured with MTS assay and CellTiter-Glo® 3D Cell Viability Assay. Furthermore, association ZnPC with TFK1 cells was evaluated by fluorescent spectroscopy. The average hydrodynamic size of ZnPC-M increased with amount of ZnPC encapsulated, ranging from 56 ± 1 nm to 70 ± 1 nm. After incubation with human plasma, ZnPC was immediately and partly partitioned to lipoproteins but not to albumin. At 6 hours timepoint, less than 30% of the ZnPC content had been released from the ZnPC-M. The phototoxicity of ZnPC-M in TFK1 cells was inversely correlated with ZnPC loading for both 2D and 3D cells: 0.2% w/w ZnPC-M showed the lowest LC50. Quantification of the ZnPC associated with TFK1 cells before light exposure showed similar amounts of drug independently of the loading (0.2, 0.4 and 0.6 % w/w). These data suggests that ZnPC-M at 0.2% w/w have faster drug release, making the ZnPC available at the time of the illumination for the generation of ROS, and thus inducing higher phototoxicity. pCL-PEG micelles encapsulating ZnPC were developed to improve ZnPC solubility and stability. Increasing drug loading was shown to decrease phototoxicity of ZnPC-M in TFK1 cells, while not affecting plasma stability. Faster drug release seems to be the reasoning for the 0.2% w/w ZnPC-M outperform the higher loaded ZnPC-M. These results prompt further investigation in vivo to confirm the superiority of 0.2% w/w ZnPC-M in both pharmacokinetics and PDT efficacy.