Key result
Native digestion extracts host cell proteins from AAV serotypes more effectively than denaturing conditions.
Why the study?
Highly abundant biotherapeutics hamper detection of low-level host cell protein contaminants that can impact product quality, efficacy, and toxicity.
Native digestion improves the detection of low-level host cell protein contaminants in adeno-associated virus vectors used for gene therapy.
Native digestion may aid low-level HCP detection in AAV vectors; hypothesis-generating for gene therapy manufacturing and requires clinical validation.
Host cell proteins (HCPs) are contaminants of biotherapeutics produced from engineered living systems; they can influence the product's quality, efficacy, and toxicity. Liquid chromatography coupled to mass spectrometry can detect HCPs thereby mitigating their risks. However, highly abundant biotherapeutics hamper the detection of low-level HCPs. Sample preparation termed native digestion has proven effective to preferentially digest and draw out HCPs from intact antibodies. Here, we adapted native digestion to adeno-associated viruses (AAV), which is a vector gaining popularity for gene therapy. We leveraged quantitative proteomics using capillary-flow liquid chromatography-mass spectrometry (LC-MS) and demonstrated that native digestion was more effective than applying denaturing conditions to extract the HCPs associated with different AAV serotypes.
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Lodge et al. (2024) studied this question. Native digestion vs. Denaturing conditions was evaluated on Extraction of host cell proteins associated with different AAV serotypes. Native digestion was more effective than applying denaturing conditions to extract host cell proteins associated with different adeno-associated virus serotypes.
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