RNA-based delivery systems with levels of control
AssigneeCalifornia Institute of Technology
InventorsXiaojing Gao, Lucy S. Chong, Michael B. Elowitz, Matthew S-M Kim
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Abstract
Disclosed herein include methods, compositions, and systems suitable for use in delivering a polynucleotide to a target cell of a subject in need thereof. In some embodiments, a viral vector comprises a polynucleotide encoding nucleoprotein (N), phosphoprotein (P), matrix protein (M), RNA-dependent RNA polymerase (L), and one or more transgenes. The viral vector can comprise one or more of a conditionally stable fusion protein, a protease fusion protein, a degron fusion protein, and/or a glycoprotein derived of another species than the viral vector polynucleotide to enable control of viral vector transduction and/or replication.
Claims20
A viral vector, comprising:
- a polynucleotide encoding a nucleoprotein (N), a phosphoprotein (P), a matrix protein (M), an RNA-dependent RNA polymerase (L), and one or more transgenes,
- wherein at least one of the N, P, M, or L is a degron fusion protein comprising a degron capable of binding a degron stabilizing molecule, and
- wherein the degron fusion protein changes from a destabilized state to a stabilized state when the degron binds to the degron stabilizing molecule;
- wherein at least one of the N, P, M, or L is a conditionally stable fusion protein comprising a stabilizing molecule binding domain capable of binding a stabilizing molecule, and wherein the conditionally stable fusion protein changes from a destabilized state to a stabilized state when the stabilizing molecule binding domain binds to the stabilizing molecule, and/or
- wherein two of the N, P, M, or L are expressed as a protease fusion protein comprising a first protein and a second protein of the two of the N, P, M, or L separated from a protease by a first cut site and a second cut site, respectively, wherein the protease is capable of cutting the first cut site and the second cut site when the protease is not bound by a protease inhibitor, and wherein the first protein and the second protein are in inactive states when the first cut site and the second cut site are not cut, respectively.
A method of controlling virus vector, comprising:
- causing a polynucleotide of a viral vector to be internalized into a cell of a subject, wherein the polynucleotide encodes a nucleoprotein (N), a phosphoprotein (P), a matrix protein (M), an RNA-dependent RNA polymerase (L), and one or more transgenes;
- and
- causing a stabilizing molecule to be present in or expressed by the cell, wherein at least one of the N, P, M, or L is a conditionally stable fusion protein comprising a stabilizing molecule binding domain, thereby the conditionally stable fusion protein changes from a destabilized state to a stabilized state when the stabilizing molecule binding domain binds the stabilizing molecule, and thereby a copy of the polynucleotide is generated from the polynucleotide;
- and/or
- causing a protease inhibitor to be absent in the cell, wherein two of the N, P, M, or L are expressed as a protease fusion protein comprising a first protein and a second protein of the two of the N, P, M, or L separated from a protease by a first cut site and a second cut site, respectively, thereby the protease cuts the first cut site and the second cut site to generate the first protein and the second protein are in active states, and thereby a copy of the polynucleotide is generated from the polynucleotide.
Dependencies are inferred from the claim text.