Genome length determination in adeno-associated virus vectors with mass photometry.

Autor:	Hiemenz C; Coriolis Pharma Research GmbH, 82152 Martinsried, Germany., Baumeister N; Coriolis Pharma Research GmbH, 82152 Martinsried, Germany.; Department of Pharmacy, Pharmaceutical Technology and Biopharmaceutics, LMU Munich, 81377 Munich, Germany., Helbig C; Coriolis Pharma Research GmbH, 82152 Martinsried, Germany., Hawe A; Coriolis Pharma Research GmbH, 82152 Martinsried, Germany., Babutzka S; Department of Ophthalmology, University Hospital, LMU Munich, 80336 Munich, Germany., Michalakis S; Department of Ophthalmology, University Hospital, LMU Munich, 80336 Munich, Germany., Friess W; Department of Pharmacy, Pharmaceutical Technology and Biopharmaceutics, LMU Munich, 81377 Munich, Germany., Menzen T; Coriolis Pharma Research GmbH, 82152 Martinsried, Germany.
Jazyk:	angličtina
Zdroj:	Molecular therapy. Methods & clinical development [Mol Ther Methods Clin Dev] 2023 Nov 19; Vol. 31, pp. 101162. Date of Electronic Publication: 2023 Nov 19 (Print Publication: 2023).
DOI:	10.1016/j.omtm.2023.101162
Abstrakt:	Recombinant adeno-associated viruses (rAAVs) are attractive therapeutic viral vectors for gene delivery. To ensure the efficacy and safety of rAAV-based therapies, comprehensive characterization of the adeno-associated virus (AAV) capsids is essential. Mass photometry (MP) provides the advantage of short analysis times, low sample consumption, and high accuracy of molecular mass determination. Despite having just recently emerged, MP has already been used to characterize AAV genome content and quantify filled/empty capsid ratios. In this study, we explored three approaches for the application of MP to assess genome length in AAVs. In approach 1, genome length in intact AAVs was approximated with good precision (coefficient of variation [%CV] < 2.6%) and accuracy (±5%) by using a straightforward protein-based calibration. In approach 2, genome length was determined even more accurately (±1%, %CV < 2.9%) considering calibration with a set of additional AAVs of different genome length. In approach 3, genome length was assessed after genome release from the capsid by heating in 1% sodium dodecyl sulfate followed by surfactant removal with precision of %CV < 0.7% and accuracy of ±5%. In conclusion, the three developed MP-based approaches are fast, precise, and accurate methods for genome length determination in AAVs, differing in their calibration materials and efforts. Competing Interests: The authors declare no competing interests. (© 2023 The Authors.)
Databáze:	MEDLINE
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