Method for purifying an enveloped virus
Inventors
Sandig, Volker • Mühle, Michael • Krügener, Sven
Assignees
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Publication Number
US-11999975-B2
Publication Date
2024-06-04
Expiration Date
Abstract
The present invention relates to a method for purifying an enveloped virus. The present invention further relates to an enveloped virus or a plurality of enveloped viruses obtainable by said method.
Core Innovation
The invention relates to a method for purifying an enveloped virus by a bind-elute downstream process using mixed mode chromatography. The method binds an enveloped virus comprised in a preparation to a mixed mode chromatography carrier and then elutes the enveloped virus from the mixed mode chromatography carrier with an elution buffer.
The mixed mode chromatography carrier is a hydrophobic ion exchange chromatography carrier. The method is characterized in that the elution buffer comprises arginine, and the elution may be performed under conditions such that the elution buffer has a higher salt concentration than the equilibration buffer and washing buffer and/or a higher pH than the equilibration buffer and washing buffer.
The method may further include optional preparation processing before binding, such as cell lysis, virus clarification, and/or nuclease treatment. After elution, the mixed mode eluate may be subjected to one or more selected downstream processing steps including filtration, chromatography, and/or nuclease treatment.
Claims Coverage
The partial content includes one independent claim directed to a bind-elute purification method for an enveloped virus using a hydrophobic ion exchange mixed mode chromatography carrier with an arginine-containing elution buffer, with multiple dependent claims refining preparation handling, carrier conditioning, and downstream processing. The inventive features below reflect the main elements required by the independent claim and emphasized refinements described in the dependent claims.
Hydrophobic ion exchange mixed mode carrier for binding an enveloped virus
Binding an enveloped virus comprised in a preparation to a mixed mode chromatography carrier, wherein the mixed mode chromatography carrier is a hydrophobic ion exchange chromatography carrier.
Arginine-containing elution buffer
Eluting the enveloped virus from the mixed mode chromatography carrier with an elution buffer, wherein the elution buffer comprises arginine.
Carrier equilibration and washing while virus remains bound
Equilibrating the mixed mode chromatography carrier with an equilibration buffer and washing the mixed mode chromatography carrier with a washing buffer while the enveloped virus remains bound to the carrier.
Relative elution buffer salt concentration and/or pH versus equilibration and wash buffers
Eluting with an elution buffer having a higher salt concentration than the equilibration buffer and washing buffer and/or a higher pH than those buffers.
Optional upstream preparation processing including lysis, clarification, and/or nuclease treatment
Before binding, optionally processing the enveloped virus preparation by cell lysis, virus clarification, and/or nuclease treatment.
Downstream processing of the mixed mode eluate including filtration, chromatography, and/or nuclease treatment
Subjecting the eluate to one or more selected downstream processing steps selected from filtration, chromatography, and/or nuclease treatment.
Across the independent claim and its refinements, the main claim coverage centers on binding an enveloped virus to a hydrophobic ion exchange mixed mode chromatography carrier and eluting with an elution buffer comprising arginine. Dependent claims further specify optional upstream preparation processing, carrier equilibration and washing while virus remains bound, relative elution-condition constraints versus equilibration and wash buffers, and downstream processing steps applied to the mixed mode eluate.
Stated Advantages
Increase virus yield and purity while mitigating the need for high salt.
Host cell DNA reduction is reported for purified products.
Nuclease treatment effectiveness is described as maintained under high-salt/arginine conditions.
Robustness across runs is described.
Documented Applications
Purification of Modified Vaccinia Ankara (MVA) virus using the described mixed mode chromatography approach.
Purification of Newcastle disease virus (NDV) using the described mixed mode chromatography approach.
Downstream clarification/processing options for purified MVA and/or NDV products are described, including filtration, further chromatography, and/or nuclease treatment.
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