Ligand discovery and gene delivery via retroviral surface display
Compositions of retroviruses and methods of using the same for gene delivery, wherein the retroviruses comprise a viral envelope protein comprising at least one mutation that diminishes its native function, a non-viral membrane-bound protein comprising a membrane-bound domain and an extracellular targeting domain.
1. A retrovirus comprising:
(a) a nucleic acid encoding a gene of interest; and
(b) a viral envelope that comprises at least two different proteins:
(i) a viral envelope protein comprising at least one mutation that diminishes the native viral tropism compared to the non-mutated viral envelope protein, wherein the viral envelope protein is a vesicular stomatitis virus (VSV-G) envelope protein that comprises an amino acid substitution at position K47 and/or amino acid substitution at position R354, said positions with reference to SEQ ID NO: 13 or a cocal virus G envelope that comprises an amino acid substitution at position K64 and/or position R371, said positions with reference to SEQ ID NO: 51; and
(ii) a non-viral membrane-bound protein comprising a transmembrane domain and an extracellular targeting domain that binds to a ligand expressed on the surface of a cell.
2. The retrovirus of claim 1 , wherein the retrovirus is a lentivirus.
3. The retrovirus of claim 1 , wherein:
(a) the at least one mutation of the VSV-G envelope protein is an amino acid substitution at position K47 to alanine (A) or glutamine (Q) and/or position R354 to alanine (A) or glutamine (Q), said positions with reference to SEQ ID NO: 13; or
(b) the at least one mutation of the cocal virus G envelope protein is an amino acid substitution at position K64 to alanine (A) or glutamine (Q) and/or position R371 to alanine (A) or glutamine (Q), said positions with reference to SEQ ID NO: 51.
4. The retrovirus of claim 1 , wherein the extracellular targeting domain comprises a protein, a peptide, or an antibody.
5. The retrovirus of claim 1 , wherein the extracellular targeting domain binds to a ligand expressed on the surface of a T cell or B cell.
6. The retrovirus of claim 1 , wherein the extracellular targeting domain comprises an interleukin-13 protein domain, a CD80 protein domain, a full-length antibody, an antibody fragment, a nanobody, a single chain antibody (scFv), an anti-CD19 antibody, an anti-TCR antibody, or an anti-CD3 antibody.
7. The retrovirus of claim 1 , wherein the ligand is a T cell receptor (TCR), a cytokine receptor, a cytokine, a T cell surface marker, CD3, CD19, or CD20.
8. A retrovirus comprising:
(a) a nucleic acid encoding a gene of interest; and
(b) a viral envelope that comprises at least two different proteins:
(i) a CD80 protein that comprises a CD80 extracellular domain and a transmembrane domain; and
(ii) a mutated viral envelope protein comprising at least one mutation that diminishes the native viral tropism of the mutated viral envelope protein compared to the non-mutated viral envelope protein, wherein the viral envelope protein is a VSV-G envelope protein that comprises an amino acid substitution at position K47 and/or an amino acid substitution at position R534, said positions with reference to SEQ ID NO: 13 or a cocal virus G envelope protein that comprises an amino acid substitution at position K64 and/or an amino acid substitution at position R371, said positions with reference to SEQ ID NO: 51.
9. The retrovirus of claim 8 , wherein the retrovirus is a lentivirus.
10. The retrovirus of claim 8 , wherein:
(a) the at least one mutation of the VSV-G envelope protein is an amino acid substitution at position K47 to alanine (A) or glutamine (Q) and/or position R354 to alanine (A) or glutamine (Q), said positions with reference to SEQ ID NO: 13; or
(b) the at least one mutation of the cocal virus G envelope protein is an amino acid substitution at position K64 to alanine (A) or glutamine (Q) and/or position R371 to alanine (A) or glutamine (Q), said positions with reference to SEQ ID NO: 51.
11. The retrovirus of claim 8 , further comprising a non-viral membrane-bound protein comprising an extracellular targeting domain that binds to a ligand expressed on the surface of a cell.
12. The retrovirus of claim 11 , wherein the non-viral membrane-bound protein comprises an extracellular targeting domain extracellular targeting domain that comprises an interleukin-13 protein domain, a full-length antibody, an antibody fragment, a nanobody, a single chain antibody (scFv), an anti-CD19 antibody, an anti-TCR antibody, or an anti-CD3 antibody.
13. The retrovirus of claim 11 , wherein the ligand is a T cell receptor (TCR), a cytokine receptor, a cytokine, a T cell surface marker, CD3, CD19, or CD20.
14. A retrovirus comprising:
(a) a nucleic acid encoding a gene of interest;
(b) a viral envelope that comprises at least two different proteins:
(i) a non-viral membrane-bound protein comprising a transmembrane domain and an extracellular targeting domain that comprises an anti-CD3 antibody; and
(ii) a mutated viral envelope protein comprising at least one mutation that diminishes the native viral tropism of the mutated viral envelope protein compared to the non-mutated viral envelope protein, wherein the viral envelope protein is a VSV-G envelope protein that comprises an amino acid substitution at position K47 and/or an amino acid substitution at position R354, said positions with reference to SEQ ID NO: 13 or a cocal virus G envelope protein that comprises an amino acid substitution at position K64 and/or an amino acid substitution at position R371, said positions with reference to SEQ ID NO: 51.
15. The retrovirus of claim 14 , wherein the retrovirus is a lentivirus.
16. The retrovirus of claim 14 , wherein:
(a) the at least one mutation of the VSV-G envelope protein is an amino acid substitution at position K47 to alanine (A) or glutamine (Q) and/or position R354 to alanine (A) or glutamine (Q), said positions with reference to SEQ ID NO: 13; or
(b) the at least one mutation of the cocal virus G envelope protein is an amino acid position at position K64 and/or position R371, said positions with reference to SEQ ID NO: 51.
17. The retrovirus of claim 14 , further comprising a non-viral membrane-bound protein comprising an extracellular targeting domain that binds to a ligand of a cell.
18. The retrovirus of claim 17 , wherein the non-viral membrane-bound protein comprises an extracellular targeting domain extracellular targeting domain that comprises an interleukin-13 protein domain, a CD80 protein domain, a full-length antibody, an antibody fragment, a nanobody, a single chain antibody (scFv), an anti-CD19 antibody, or an anti-TCR antibody.
19. The retrovirus of claim 17 , wherein the ligand is a T cell receptor (TCR), a cytokine receptor, a cytokine, a T cell surface marker, CD19, or CD20.