Université De Nantes

Chu Nantes

Association Francaise Contre Les Myopathies

Institut National De La Sante Et De La Recherche Medicale

Institut National De La Sante Et De La Recherche Medicale (Inserm)

Inserm (Institut National De La Sante Et De La Recherche Medicale)

Association Francaise Contre Les Myopathies (Afm)

Nicole Brument

A method for obtaining purified recombinant Adeno-Associated Virus particles (rAAV) is provided. The method includes steps of: a) performing a depth filtration of a cell lysate or a culture supernatant obtained from cells producing rAAV particles, whereby a rAAV-containing clarified composition is provided; b) submitting the rAAV-containing clarified composition to a first step of anion-exchange chromatography on a chromatographic support wherein the rAAV-containing fraction is collected, whereby a first rAAV enriched composition is provided; c) submitting the first rAAV enriched composition at least once to a second step of anion-exchange chromatography on a chromatographic support wherein the rAAV-containing fraction is collected, whereby a second rAAV enriched composition is provided; d) submitting the second rAAV enriched composition to a step of tangential flow filtration to provide purified recombinant Adeno-Associated Virus particles (rAAV).

Recombinant adeno-associated virus particle purification with multiple-step anion exchange chromatography

The invention describes a method for obtaining purified recombinant Adeno-Associated Virus particles (rAAV), comprising performing a depth filtration of a starting material previously obtained from cells producing rAAV particles comprising a cell lysate and/or a culture supernatant, followed by a first step of anion-exchange chromatography, a second step of anion-exchange chromatography and a final step of tangential flow filtration, whereby purified recombinant Adeno-Associated Virus particles (rAAV) are provided.

The invention describes a method for obtaining purified recombinant Adeno-Associated Virus particles (rAAV), comprising the steps of: a) performing a depth filtration of a starting material previously obtained from cells producing rAAV particles, the said starting material being selected in a group comprising a cell lysate and a culture supernatant, whereby a rAAV-containing clarified composition is provided; b) submitting the rAAV-containing clarified composition to an affinity purification step, whereby a first rAAV enriched composition is provided; c) submitting the first rAAV enriched composition at least once to: c1) a step of anion-exchange chromatography on a chromatographic support wherein elution is performed by using a salt gradient, preferably a linear salt gradient, and wherein the rAAV-containing fraction is collected, whereby a second rAAV enriched composition is provided; or c2) a step of density gradient centrifugation, wherein the rAAV-containing fraction is collected, whereby a second rAAV enriched composition is provided; d) submitting the second rAAV enriched composition to a step of tangential flow filtration, whereby purified recombinant Adeno-Associated Virus particles (rAAV) are provided.

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Nantes, France

Nicole Brument