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.
Claims25
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 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;
- and
- an envelope comprising a glycoprotein not encoded by the polynucleotide,
- wherein the glycoprotein is of the species of any of the N, P, M, and L encoded by the polynucleotide, and/or wherein the glycoprotein, or a portion thereof, is derived of another species, and
- wherein a glycoprotein binding domain of a first bridge protein is capable of binding the glycoprotein.
A method of controlling virus vector, comprising:
- causing a polynucleotide of a viral vector to be internalized into a first 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 wherein the viral vector comprises an envelope comprising a glycoprotein not encoded by the polynucleotide;
- and
- causing a first glycoprotein to be expressed by the first cell, wherein the first glycoprotein is of the species of any of the N, P, M, and L encoded by the polynucleotide, thereby a first virus particle, comprising a first replicated copy of the polynucleotide and the first glycoprotein, buds from the first cell and is internalized into a second cell;
- causing a second glycoprotein to be expressed by the first cell and a first bridge protein to be introduced into the subject, wherein the second glycoprotein, or a portion thereof, is derived of another species, thereby a second virus particle, comprising a second replicated copy of the polynucleotide and the second glycoprotein, buds from the first cell and is internalized into a second cell after a glycoprotein binding domain of the first bridge protein binds the second glycoprotein;
- causing a stabilizing molecule to be present in or expressed by the second 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 third copy of the polynucleotide is generated from the second copy of the polynucleotide;
- and/or
- causing a protease inhibitor to be absent in the second 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 fourth copy of the polynucleotide is generated from the second copy of the polynucleotide.
Dependencies are inferred from the claim text.