Bocaparvovirus small noncoding rna and uses thereof
AssigneesUniversity of Iowa Research Foundation, University of Kansas
InventorsZiying Yan, John F. Engelhardt, Jianming Qiu, Zekun Wang
Life of this patent
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Abstract
Vectors having a nucleotide sequence having SEQ ID NO:1 or a nucleotide sequence having at least 85% identity to SEQ ID NO:1, or a portion thereof, that is capable of regulating bocaparvovirus replication, or vectors having the complement of the nucleotide sequence, and methods of using the vectors, are provided.
Claims30
A vector comprising a nucleotide sequence having SEQ ID NO:1 or a nucleotide sequence having at least 85% identity to SEQ ID NO:1, or a portion thereof, that is capable of regulating bocaparvovirus replication, wherein the vector does not encode bocavirus (BoV) NS1;
- a vector comprising a nucleic acid segment that has a sequence that is a complement to a nucleotide sequence having SEQ ID NO:1 or a nucleotide sequence having at least 85% identity to SEQ ID NO:1, or a portion thereof, that inhibits expression of SEQ ID NO:1;
- or a vector having a recombinant BoV genome comprising a LEH, a 3′ UTR and a REH, wherein the 3′ UTR includes a nucleotide sequence having at least 85% identity to SEQ ID NO:1, or a portion thereof that is capable of regulating BoV replication, wherein the REH includes a 46 nucleotide sequence comprising OriR having a TRS and a NS1 nicking or binding site, wherein the genome encodes bocavirus NP1, NS4 or NS2, and optionally VP1, VP2 and/or VP3, or any combination thereof, but not NS1.
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A method to prevent, inhibit or treat bocavirus infection in a mammal, comprising administering to a mammal a composition comprising an effective amount of a vector comprising a nucleic acid segment that has a sequence that is a complement to a nucleotide sequence having SEQ ID NO:1 or a nucleotide sequence having at least 85% identity to SEQ ID NO:1, or a portion thereof, that inhibits expression of SEQ ID NO:1.
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A helper virus-free method to produce recombinant adeno-associated virus (rAAV) or to prepare helper virus-free chimeric rAAV/BoV, comprising:
- contacting mammalian cells with i) one or more rAAV vectors, and one or more BoV vectors that express one or more of BoV NP1, BoV NS4 or NS2 and a nucleotide sequence having SEQ ID NO:1 or a nucleotide sequence having at least 85% identity to SEQ ID NO:1, or a portion thereof that is capable of regulating bocavirus replication, which cells, rAAV vector or BoV vector or other vector in the cells express AAV Rep and/or AAV Cap, in an amount effective to yield rAAV, wherein the cells and vectors do not express BoV VP1-3, or ii) one or more rAAV vectors, one or more BoV vectors that express one or more of BoV NP1, and BoV NS4 or NS2, and a vector that expresses at least adenovirus E2A, which cells, rAAV vector or BoV vector or other vector in the cells express AAV Rep and/or AAV Cap, in an amount effective to yield rAAV, wherein the cells and vectors do not express another viral capsid protein such as BoV VP1-3;
- and
- isolating adenovirus helper-free rAAV from the cells;
- or
- contacting mammalian cells with one or more rAAV vectors, and one or more BoV vectors that express one or more of BoV NP1, BoV NS4 or NS2, and a nucleotide sequence having SEQ ID NO:1 or a nucleotide sequence having at least 85% identity to SEQ ID NO:1, or a portion thereof that is capable of regulating bocavirus replication, in an amount effective to yield chimeric rAAV/BoV comprising a genome corresponding to at least one of the rAAV vectors, wherein the vectors and cells do not express BoV NS1 or AAV Cap and wherein at least one of the rAAV vectors, the BoV vectors or cells expresses AAV Rep and/or Bov VP1-3;
- and
- isolating chimeric AAV/BoV.
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An isolated mammalian cell for helper virus-free production of recombinant adeno-associated virus (rAAV), wherein the mammalian cell i) expresses a rAAV genome comprising a transgene, BoV NP1, BoV NS4 or NS2, a nucleotide sequence having SEQ ID NO:1 or a nucleotide sequence having at least 85% identity to SEQ ID NO:1, or a portion thereof that is capable of regulating bocavirus replication, and expresses AAV Rep and/or AAV Cap, or ii) expresses a rAAV genome comprising a transgene, one or more of BoV NP1, and BoV NS4 or NS2, adenovirus E2A, and AAV Rep and/or AAV Cap, in an amount effective to yield helper-free rAAV,
- wherein the mammalian cell does not express viral capsid protein other than AAV Cap.
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Dependencies are inferred from the claim text.