Recombinant adeno-associated virus compositions and methods for producing same

A therapeutic composition comprising recombinant adeno-associated virus (rAAV) particles, wherein the rAAV particles comprise an adeno-associated virus 9 (AAV9) capsid protein and an expression cassette encoding a gene product of interest, wherein the therapeutic composition comprises more than 1E+13 viral genomes per milliliter (vg/mL) rAAV particles,

• wherein the therapeutic composition comprises less than 15% empty rAAV particles and at least 80% full rAAV particles,
• wherein the gene product of interest is human glucocerebrosidase (GCase), and the expression cassette comprises the nucleotide sequence of SEQ ID NO:
• 2,
• wherein the therapeutic composition is produced by infecting Sf9 insect cells with a population of Baculovirus vectors and lysing the insect cells to produce a cellular lysate comprising the rAAV particles, wherein the cellular lysate is subjected to a method comprising the steps of:
• (i) contacting an affinity chromatography column with the cellular lysate, wherein the affinity column comprises a binding agent specific for a capsid protein of the rAAV particles under conditions under which the rAAV particles bind to the affinity chromatography column;
• (ii) eluting the bound rAAV particles from the column thereby producing a first eluate;
• (iii) performing anion-exchange chromatography on the first eluate to produce a second eluate, wherein the second eluate comprises fewer empty rAAV particles than the first eluate;
• and
• (iv) concentrating the second eluate by performing tangential flow filtration using a flow buffer comprising Tris, MgCl2, NaCl, and Poloxamer 188, thereby producing the therapeutic composition comprising rAAV particles, and
• wherein the therapeutic composition comprises less than or equal to 1.3 ng residual host cell DNA from the insect cells per 1E+14 vg/ml of the rAAV particles of the therapeutic composition.