Myelin nanovesicles and uses thereof
The invention concerns nanovesicles of nanostructured myelin and uses thereof in the treatment of demyelinating and neurodegenerative diseases of the central (CNS) and peripheral (PNS) nervous system. Under another aspect, processes for the preparation of myelin nanovesicles having particular characteristics that make them suitable for recovery of the myelin sheath, where it is compromised, as a drug delivery system for CNS or PNS, and for immunotolerance, are described.
1 . A nanovesicle having a diameter in the range from 30 to 200 nm and a zeta potential in the range from −10 to −50 mV, said nanovesicle comprising nanostructured myelin, wherein said nanostructured myelin comprises from 70% to 85% by weight of lipids and from 15% to 30% by weight of proteins, and wherein said myelin is extracted from brain tissue.
2 . The nanovesicle according to claim 1 , wherein said proteins comprise Myelin basic protein, Myelin proteolipid protein, Myelin-oligodendrocyte glycoprotein, 2′3′-cyclic-nucleotide 3′-phosphodiesterase, Calmodulin-1, and Myelin-associated glycoprotein, wherein said lipids and said proteins are present in the native structure and conformation and have not undergone any physico-chemical modifications selected from the group consisting of fusion, enzymatic cutting, bonding or addition of spacers or functional groups, cross-linking.
3 . The nanovesicle according to claim 1 , loaded with a substance or a molecule.
4 . The nanovesicle according to claim 3 , wherein said substance is a drug, a contrast agent, a fluorescent probe, a cytokine, growth factors, antioxidants, anti-inflammatoires, immunomodulators.
5 . A method for using a nanovesicle according to claim 1 , as a vehicle for vehiculating drugs and therapeutically active biomolecules to the central and peripheral nervous systems.
6 . A method for treating demyelinating diseases in a subject in need thereof, said method comprising the step of using the nanovesicle according to claim 1 .
7 . The method according to claim 6 , wherein said demyelinating diseases are multiple sclerosis, acute disseminated encephalomyelitis, adrenoleukodystrophy and adrenomyeloneuropathy, and Leber's hereditary optic neuropathy.
8 . A method for treating immunotolerance in a subject in need thereof, said method comprising the step of using the nanovesicle according claim 1 .
9 . A method for treating a neurodegenerative disease of the central and peripheral nervous system in a subject in need thereof, said method comprising the step of using the nanovesicle according to claim 1 .
10 . The method according to claim 9 , wherein said neurodegenerative diseases of the central and peripheral nervous system are Alzheimer's disease, Parkinson's disease, Amyotrophic Lateral Sclerosis.
11 . A process for the preparation of nanovesicles having the steps of:
a. preparing a myelin dispersion in an aqueous solvent at a concentration in the range from 0.1 mg/ml to 10 mg/ml, wherein said myelin is extracted from brain tissue;
b. homogenizing the myelin dispersion of step a. with an Ultra-Turrax at an intensity in the range from 8,000 to 22,000 rpm, for a time in the range from 5 to 15 minutes, to obtain a homogenized dispersion;
c. sonicating the homogenized dispersion of step b. with a sonicator at an intensity of 35-45 kHz for a time of 1-15 minutes, to obtain myelin nanovesicles having a diameter in the range from 30 to 200 nm, and a zeta potential in the range from −10 to −50 mV.
12 . The process according to claim 11 , wherein the myelin dispersion of step a. is prepared by injecting myelin, previously dissolved in an organic solvent miscible with water, in the aqueous medium, during step b.
13 . A process for the preparation of myelin nanovesicles having the steps of:
a. preparing a myelin dispersion in a polar solvent at a concentration in the range from 0.1 mg/ml to 10 mg/ml, wherein said myelin is extracted from brain tissue;
b. flowing the myelin dispersion of step a. through a microfluidic chip for micro-mixing, with a flow of between 0 and 5,000 μl/min, to obtain myelin nanovesicles having a diameter in the range from 30 to 200 nm, and a zeta potential in the range from −10 to −50 mV.
14 . The process according to claim 11 , having the additional step of freeze-drying the myelin nanovesicles obtained from step c., to obtain lyophilized myelin nanoparticles.
15 . A nanovesicle obtainable by the process according to claim 11 , wherein said nanovesicle has a diameter in the range from 30 to 200 nm, or in the range from 90 to 110 nm, and a zeta potential in the range from −10 to −50 mV, said nanovesicle comprising myelin, wherein said myelin comprises from 70 to 85% by weight of lipids and from 15 to 30% by weight of proteins.
16 . The nanovesicle according to claim 15 , wherein said proteins comprise Myelin basic protein, Myelin proteolipid protein, Myelin-oligodendrocyte glycoprotein, 2′3′-cyclic-nucleotide 3′-phosphodiesterase, Calmodulin-1 and Myelin-associated glycoprotein.
17 . The nanovesicle according to claim 16 , wherein said myelin is nanostructured.
18 . The nanovesicle according to claim 1 , wherein said nanovesicle has a diameter in the range from 90 to 110 nm.