Method of increasing the function of an AAV vector

摘要

A method of correcting singletons in a selected AAV sequence in order to increasing the packaging yield, transduction efficiency, and/or gene transfer efficiency of the selected AAV is provided. This method involves altering one or more singletons in the parental AAV capsid to conform the singleton to the amino acid in the corresponding position(s) of the aligned functional AAV capsid sequences.

主权项

1. A method of increasing the packaging yield, transduction efficiency, and/or gene transfer efficiency of a parental AAV, said method comprising the steps of:
(a) comparing a parental AAV capsid sequence with a library of functional AAV capsid sequences in an alignment, wherein said library comprises at least three functional AAV capsids which are at least 85% identical over the full-length of the aligned vp1 sequences and which represent at least three different clades, wherein the library comprises at least two AAV sequences from each clade; (b) identifying at least one singleton in the parental AAV capsid, said singleton being an amino acid residue in a position in the parental AAV capsid which differs from the amino acid in the corresponding position of the aligned functional AAV capsid sequences in the library all of which have the same amino acid residue in that position; (c) altering the singleton in the parental AAV capsid to the amino acid located in the corresponding position in the functional AAV capsid sequences in the library to provide a modified AAV capsid sequence, and (d) assessing the modified AAV for increased packaging yield, transduction efficiency, and/or gene transfer efficiency as compared to the parental AAV.