MAgnetic VEsicle Rotation-Induced Cell-Killing – MAVERICK

Magnetic polymersomes developed around 2005 are a spectacular example of versatile nanovectors capable of releasing active substances at a tumor site under the action of a high frequency (HF) magnetic field that provokes local (i.e. nanoscale) heating of the membrane, enhancing its permeability. In a static magnetic field, these nano-objects undergo substantial elongation. If one fixes that shape by chemical crosslinking under magnetic field, these anisometric entities will give access to new delivery modes, such as the application of a magnetic torque on the cellular membranes (outer plasma membrane and intracellular organelle membranes) by applying a low frequency (LF) magnetic field. The MAVERICK project aims at studying the ellipsoid-like or more complex deformations of magnetic polymersomes (WP1), the biocompatibility and internalization pathways as a function of polymersome morphologies (WP2), and the magneto-induced toxicity on cells induced by LF or HF magnetic fields (WP3). More precisely WP1 comprises at first preparation of several types of magnetic polymersomes (MagPol) starting from amphiphilic di-block copolymers whose hydrophilic block is poly(oxyethylene) (POE) and the hydrophobic block either poly(butadiene) (PBut) or poly(trimethylene carbonate) (PTMC), in which magnetic iron oxide nanoparticles (IONPs) were embedded. Deformation of these soft magnetic shells under a static magnetic field will be studied theoretically by numerical simulation and by several experimental methods: electron microscopy, dynamic depolarized scattering (DDLS) and small angle neutron scattering (SANS). The novelty compared to previous studies will consist in a "quench" of the expected elongated shapes of MagPol under field through chemical crosslinking of the hydrophobic blocks of copolymers. Different crosslinking reactions will be tried out, first by radical initiation by an oxidizing agent for PBut, and also by photo-crosslinking induced par UV irradiation UV for PBut and PTMC (with the aid of copolymerization with a monomer bearing pendant unsaturated groups). One last type of polymersomes will comprise a thermosensitive component, either a semi-crystalline PTMC or poly(caprolactone) block, or a fraction phospholipids in the gel state such as dipalmytoyl phosphocholine (DPPC) that exhibits melting at 41°C. Once these different MagPol samples will be characterized on a physicochemical aspect, their biocompatibility will be assayed within the frame of WP2. Several cell lines will be used, fibroblasts, epithelial and immune system cells (monocytes and macrophages), in order to check the biocompatibility of the different MagPol types. For each chemical composition, comparison will be made between the isotropic polymersomes (IsoMagPol) and the same after crosslinking under magnetic field (AnisoMagPol). Then, the efficacy and kinetics of cell internalization of different MagPol batches will be studied by diverse methods: magnetophoresis and flow cytometry for quantification, specific fluorescent labelling for confocal microscopy, and finally by electron microscopy and tomography, in order to precise the subcellular localization of MagPol within lysosomes or on the contrary inside cytosol. Finally, WP3 will consist in the building of magnetic field inductors oscillating at low frequency (from 1 à 500 Hz) to apply magnetic torques on the cell walls through oscillation of the AnisoMagPol, either with permanent magnets or electromagnets. This low frequency excitation mode will be compared to the action of a radiofrequency magnetic field (from 100 to 750 kHz) inducting thermal effect onto the cell walls, mediated by the MagPol. In this case, the release of a fluorescent probe or model drug induced magnetically will be evidenced.