Active loading of biomolecular cargo into engineered vesicles
AssigneeNorthwestern University
InventorsJustin Alexander Peruzzi, Taylor Franklin Gunnels, Neha Prashant Kamat, Joshua N. Leonard
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Abstract
The present disclosure provides, among other things, improved methods and compositions for loading an engineered vesicle (EngV) with a fusion polypeptide or biomolecular cargo, loaded EngVs or a plurality thereof (e.g., produced in accordance with the methods described herein), compositions comprising such loaded EngVs or a plurality thereof, and methods of treating a disease or disorder by administering compositions of the present disclosure.
Claims25
A method of loading an engineered vesicle (EngV) with a fusion polypeptide, comprising:
- contacting a cell with a nucleic acid encoding a fusion polypeptide comprising (i) a biomolecular cargo, and (ii) one or more lipid tags,
- wherein contacting occurs under conditions sufficient to permit (i) uptake of the nucleic acid by the cell, (ii) translation of the fusion polypeptide, and (iii) post-translational modification of the one or more lipid tags with one or more lipids by endogenous host machinery in the cell to thereby generate a lipid-modified fusion polypeptide, and wherein the lipid-modified fusion polypeptide is directed and loaded into or on the EngV.
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A fusion polypeptide comprising (i) a biomolecular cargo and (ii) one or more lipid tags, wherein the one or more lipid tags comprise a short amino acid sequence which a host cell can post-translationally modify with one or more lipids.
A method of loading an engineered vesicle (EV) with a biomolecular cargo, comprising:
- contacting a cell with a nucleic acid encoding the biomolecular cargo, wherein contacting occurs under conditions sufficient to permit (i) uptake of the nucleic acid by the cell, (ii) translation of the biomolecular cargo,
- wherein the biomolecular cargo comprises a transmembrane domain and the transmembrane domain is modified to improve direction and loading into or on the EngV relative to the biomolecular cargo with an unmodified transmembrane domain, thereby generating a transmembrane modified biomolecular cargo, and
- wherein the transmembrane modified biomolecular cargo is directed and loaded into or on the EngV.
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Dependencies are inferred from the claim text.