Adeno-associated virus capsid variants and methods of use thereof
The present disclosure provides recombinant adeno-associated virus virions with variant capsid protein, where the recombinant AAV (rAAV) virions exhibit one or more of increased ability to cross neuronal cell barriers, increased infectivity of a neural stem cell, increased infectivity of a neuronal cell, and reduced susceptibility to antibody neutralization, compared to a control AAV, and where the rAAV virions comprise a heterologous nucleic acid. The present disclosure provides methods of delivering a gene product to a neural stem cell or a neuronal cell in an individual. The present disclosure also provides methods of modifying a target nucleic acid present in a neural stem cell or neuronal cell.
1. A recombinant adeno-associated virus (rAAV) virion comprising:
a) a variant AAV capsid protein, wherein the variant AAV capsid protein comprises an amino acid sequence having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:1, and wherein the variant capsid protein confers
increased infectivity of a neural stem cell, compared to the infectivity of the neural stem cell by a control AAV virion comprising a wild-type AAV capsid protein;
and
b) a heterologous nucleic acid comprising a nucleotide sequence encoding a heterologous gene product.
2. The rAAV virion of claim 1 , wherein the rAAV virion exhibits at least 5-fold increased infectivity of a neural stem cell compared to the infectivity of the neural stem cell by the control AAV virion comprising a wild-type AAV capsid protein.
3. The rAAV virion of claim 1 , wherein the variant AAV capsid protein exhibits increased resistance to human AAV neutralizing antibodies compared to the resistance exhibited by a control AAV virion comprising a wild-type AAV capsid protein.
4. The rAAV virion of claim 1 , wherein the gene product is an interfering RNA or an aptamer.
5. The rAAV virion of claim 1 , wherein the gene product is a polypeptide.
6. The rAAV virion of claim 5 , wherein the polypeptide is a neuroprotective polypeptide, an anti-angiogenic polypeptide, a polypeptide that induces differentiation of a neural stem cell, or a polypeptide that enhances function of a neural stem cell.
7. The rAAV virion of claim 5 , wherein the polypeptide is cerebrolysin, laminin-IKVAV, cripto, pituitary adenylate cyclase-activating polypeptide, nerve growth factor, brain derived neurotrophic factor, glial derived neurotrophic factor, fibroblast growth factor 2, neurturin, ciliary neurotrophic factor, epidermal growth factor, X-linked inhibitor of apoptosis, aromatic L-amino acid decarboxylase, glutamic acid decarboxylase, tripeptidyl peptidase, aspartoacyclase, or Sonic hedgehog.
8. A pharmaceutical composition comprising:
a) a recombinant adeno-associated virus virion of claim 1 ; and
b) a pharmaceutically acceptable excipient.
9. A method of delivering a gene product to a neural stem cell in an individual, the method comprising administering to the individual a recombinant adeno-associated virus (rAAV) virion according to claim 1 .
10. The rAAV virion of claim 1 , wherein the amino acid sequence has at least 98% amino acid sequence identity to the amino acid sequence of SEQ ID NO:1.
11. The rAAV virion of claim 1 , wherein the amino acid sequence has at least 99% amino acid sequence identity to the amino acid sequence of SEQ ID NO:1.
12. The rAAV virion of claim 1 , wherein the amino acid sequence has the amino acid sequence of SEQ ID NO:1.
13. An isolated nucleic acid comprising a nucleotide sequence that encodes a variant adeno-associated virus (AAV) capsid protein comprising an amino acid sequence having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:1, and wherein the variant capsid protein confers increased infectivity of a neural stem cell compared to the infectivity of the neural stem cell by a control AAV virion comprising a wild-type AAV capsid.
14. An isolated, genetically modified host cell comprising the nucleic acid of claim 13 .
15. The isolated nucleic acid of claim 13 , wherein the amino acid sequence has at least 98% amino acid sequence identity to the amino acid sequence of SEQ ID NO:1.
16. The isolated nucleic acid of claim 13 , wherein the amino acid sequence has at least 99% amino acid sequence identity to the amino acid sequence of SEQ ID NO:1.
17. The isolated nucleic acid of claim 13 , wherein the amino acid sequence has the amino acid sequence of SEQ ID NO:1.
18. A variant adeno-associated virus (AAV) capsid protein, wherein the variant AAV capsid protein comprises an amino acid sequence having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:1, and wherein the variant capsid protein confers increased infectivity of a neural stem cell compared to the infectivity of the neural stem cell by a control AAV virion comprising a wild-type AAV capsid.