PNIPAM-stabilized cubosomes as fusogenic delivery nanovectors for anticancer applications.

In recent years, lipid cubic nanoparticles have emerged as promising nanocarriers for drug delivery, due to the several advantages they exhibit with respect to other lipid systems. Here, we report on lipid cubic nanoparticles stabilized by PNIPAM-based amphiphilic block copolymers, specifically, poly(N, N-dimethylacrylamide)-block-poly(N-isopropylacrylamide) (PDMA-b-PNIPAM), as a new class of drug delivery systems (DDS). In vitro studies on the internalization efficiency of the DDS towards two types of human cancer cells (colon HCT-116 and bladder T24 cells), carried out employing a set of sensitive techniques (confocal laser scanning microscopy (CLSM), flow cytometry, scanning electron microscopy (SEM), fluorescence spectroscopy), highlight a prominent role of PDMA-b-PNIPAM stabilizer in enhancing the uptake of cubosomes, compared to the standard Pluronic F127-based formulations. The drug delivery potential of cubosomes, tested by encapsulating a chemotherapeutic drug, camptothecin (CPT), and conducting cytotoxicity studies against 2D plated cells and 3D spheroids, confirm that PDMA-b-PNIPAM-stabilized cubosomes improve the efficacy of treatment with CPT. The origin of this effect lies in the higher lipophilicity of the stabilizer, as we confirm by studying the interaction between the cubosomes and biomimetic membranes of lipid vesicles with Small Angle X-Ray Scattering (SAXS) and CLSM experiments. These results corroborate our fundamental understanding of the interaction between cubosomes and cells, and on the role of polymer to formulate lipid cubic nanoparticles as DDS.

Rational design of block copolymer self-assemblies in photodynamic therapy.

Photodynamic therapy is a technique already used in ophthalmology or oncology. It is based on the local production of reactive oxygen species through an energy transfer from an excited photosensitizer to oxygen present in the biological tissue. This review first presents an update, mainly covering the last five years, regarding the block copolymers used as nanovectors for the delivery of the photosensitizer. In particular, we describe the chemical nature and structure of the block copolymers showing a very large range of existing systems, spanning from natural polymers such as proteins or polysaccharides to synthetic ones such as polyesters or polyacrylates. A second part focuses on important parameters for their design and the improvement of their efficiency. Finally, particular attention has been paid to the question of nanocarrier internalization and interaction with membranes (both biomimetic and cellular), and the importance of intracellular targeting has been addressed.

Role of Polymer Micelles in the Delivery of Photodynamic Therapy Agent to Liposomes and Cells.

The use of nanocarriers for hydrophobic photosensitizers, in the context of photodynamic therapy (PDT) to improve pharmacokinetics and bio-distribution, is well-established. However, the mechanisms at play in the internalization of nanocarriers are not well-elucidated, despite its importance in nanocarrier design. In this study, we focus on the mechanisms involved in copolymer poly(ethylene oxide)-block-poly(-caprolactone) PEO-PCL and poly(ethylene oxide)-block-poly styrene PEO-PS micelles - membrane interactions through complementary physico-chemical studies on biomimetic membranes, and biological experiments on two-dimensional (2D) and three-dimensional (3D) cell cultures. Förster Resonance Energy Transfer measurements on fluorescently-labelled lipid vesicles, and flow cytometry on two cancerous cell lines enabled the evaluation in the uptake of a photosensitizer, Pheophorbide a (Pheo), and copolymer chains towards model membranes, and cells, respectively. The effects of calibrated light illumination for PDT treatment on lipid vesicle membranes, i.e., leakage and formation of oxidized lipids, and cell viability, were assessed. No significant differences were observed between the ability of PEO-PCL and PEO-PS micelles in delivering Pheo to model membranes, but Pheo was found in higher concentrations in cells in the case of PEO-PCL. These higher Pheo concentrations did not correspond to better performances in PDT treatment. We demonstrated that there are subtle differences in PEO-PCL and PEO-PS micelles for the delivery of Pheo.

Amphiphilic polymers based on polyoxazoline as relevant nanovectors for photodynamic therapy.

An amphiphilic polymer (CmPOX) based on poly(2-methyl-2-oxazoline) linked to a hydrophobic part composed of an aliphatic chain ending with a photo-active coumarin group has been synthesized. It exhibits the ability of forming small polymeric self-assemblies, typically of ca. 10 nm in size, which were characterized by TEM, cryo-TEM and DLS. The nanocarriers were further formulated to yield photo-crosslinked systems by dimerization of coumarin units of coumarin-functionalized poly(methyl methacrylate) (CmPMMA) and CmPOX. The formed vectors were used to encapsulate Pheophorbide a, a known photosensitizer for photodynamic therapy. Cytotoxicity as well as phototoxicity experiments performed in vitro on human tumor cells revealed the great potential of these nanovectors for photodynamic therapy.

A journey from the endothelium to the tumor tissue: distinct behavior between PEO-PCL micelles and polymersomes nanocarriers.

Polymeric nanocarriers must overcome several biological barriers to reach the vicinity of solid tumors and deliver their encapsulated drug. This study assessed the in vitro and in vivo passage through the blood vessel wall to tumors of two well-characterized polymeric nanocarriers: poly(ethyleneglycol-b-ε-caprolactone) micelles and polymersomes charged with a fluorescent membrane dye (DiO: 3,3'-dioctadecyloxacarbo-cyanine perchlorate). The internalization and translocation from endothelial (human primary endothelial cells HUVEC) to cancer cells (human tumor cell line HCT-116) was studied in conventional 2D monolayers, 3D tumor spheroids, or in an endothelium model based on transwell assay. Micelles induced a faster DiO internalization compared to polymersomes but the latter crossed the endothelial monolayer more easily. Both translocation rates were enhanced by the addition of a pro-inflammatory factor or in the presence of tumor cells. These results were confirmed by early in vivo experiments. Overall, this study pointed out the room for the improvement of polymeric nanocarriers design to avoid drug losses when crossing the blood vessel walls.