APR March 2024 - 19

« FORMULATION AND DEVELOPMENT
Use of AUC in AAV Analysis in a GMP Setting
Jurgen Hannig
Senior Principal Scientist, GMP Laboratory
PPD clinical research business of Thermo Fisher Scientific
Jie Ding
Senior Director, GMP Laboratory
PPD clinical research business of Thermo Fisher Scientific
The recent increase in the development of adenovirus associated
virus (AAV) based gene therapies has created a large demand for the
analysis of AAV-based drugs for the determination of percent empty
and percent full capsid as part of drug substance and drug product
quality control. The need for this analysis is triggered by fatalities
encountered in early adenovirus-based gene therapy trials due to
fatal immune responses.1
being considered experimental6
among high-resolution methods,
Subsequent research focused on AAV-based
therapies, which were known to provoke little or no immune response.
Nevertheless, the early therapy failures are still a concern such that
two recent guidances from the U.S. Food and Drug Administration
require the need for controlling the presence of empty capsids as
product related impurities to reduce the risk of (unnecessary) immune
response.2,3
and adaptive immune responses, may compete with full capsids for
receptor binding on target cells, which could necessitate an increase
in the required vector dose.4
Many analytical methodologies are available to determine the percent
empty and percent full capsid species in a sample. The most common
are: 1) anion exchange chromatography (AEX) exploring the difference
in the isoelectric points between empty and full capsids; 2) sizeexclusion
chromatography (SEC) with ultraviolet (UV), dRI (differential
refractive index) and multi-angle light scattering (MALS) detection
determining the empty to full capsid ratios based on the monomer
peak's average molecular weight and total concentration; 3) charge
density mass spectrometry (CDMS) measuring the charge and massto-charge
ratio of individual ions simultaneously, thereby allowing
direct determination of the mass of empty, partially filled and full
capsids, inclusive of their quantitation; and 4) sedimentation-velocity
analytical ultracentrifugation (SV-AUC) using the sedimentation
profiles of a sample subjected to centrifugal forces to quantify the
relative content of empty, partially filled and full capsids. For the
quantification of the content ratio, every method is subject to a tradeoff
between throughput and resolution.5
With CDMS methodology still
AAV empty capsids, in addition to stimulation of innate
AUC remains the only viable method capable of quantifying partially
filled capsids, despite requiring substantial sample amounts.7
One drawback of AUC is the current lack of availability of instrumentation
that could be used within a good manufacturing practice (GMP) setting
of a commercial quality control laboratory. Consistent and reproducible
routine AUC analysis has been made possible by the Optima AUC
(Beckman Coulter) analytical ultracentrifuge. Nevertheless, the fact that
the instrument operation software does not comply to GMP standards,
and that data analysis is performed with software that has not been
documented and validated for GMP use, complicates its application
in a GMP setting. The approach chosen to address that shortcoming is
described in this article.
Background
The PPD® Laboratory services GMP Lab employs Optima AUC
instruments controlled by the AUC Experimental Portal (on-board)
software. Of the two available AUC modes, sedimentation velocity
(SV-AUC) and sedimentation equilibrium (SE AUC), only the former
is currently used for analysis. The following workflow applies to the
data analysis:
* Setup, analysis run and data collection on the AUC
onboard computer.
* The acquired sedimentation velocity raw data are transferred
from the AUC onboard computer to a processing station
for analysis.
* SEDFIT software (freely distributed by the NIH) is used to
extract sedimentation coefficient data and create a continuous
sedimentation coefficient distribution plot, referred to as c(s).
www.americanpharmaceuticalreview.com |
| 19
»

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APR March 2024

Table of Contents for the Digital Edition of APR March 2024

Message from the Editor
Editorial Advisory Board
BIOPHARMACEUTICALS - Getting to GMP-Quality Biotherapeutics From Today’s Bench-Scale Continuous Manufacturing Systems: A Gap Analysis
MICROBIOLOGY - Critical Behavioral Attributes and the Application of COM-B Framework in Aseptic Processing
FORMULATION AND DEVELOPMENT - Use of AUC in AAV Analysis in a GMP Setting
FORMULATION AND DEVELOPMENT - Precision Medicine in Clinical Trials: A Statistical Perspective
An Interview with Dan Smithey, PhD President & CEO, Serán
FORMULATION AND DEVELOPMENT - Still Early Days for AI in Drug Discovery...Says Who?
QC Corner - Enhancing Material and Equipment Availability in Production Isolators
BIOPHARMACEUTICALS - Technologies for Aseptic Filling: The Choice is Clear
Vendor Viewpoint - Data Integrity and Rapid Micro Methods: Transforming to a Modern Microbiology Lab
MICROBIOLOGY - Bacterial Spore Formers in Disinfectant Efficacy Testing
Partner Perspective - Nanoparticle Technologies: Enablers for Ocular Drug Delivery
DRUG DELIVERY - Your Nails and You
DRUG DEVELOPMENT - Battling Exorbitance: High Costs in Sickle Cell Gene Therapies and the Imperative of Global Patient Registries for Equity
FORMULATION AND DEVELOPMENT - How Pharma Companies Are Solving Regulatory Challenges with AI-based Technology
Event Preview - CPHI North America
Event Preview - Excipient World 2024
P.I.N. Points
Advertiser's Index
APR March 2024 - CoverTip01
APR March 2024 - CoverTip02
APR March 2024 - Cover1
APR March 2024 - Cover2
APR March 2024 - 1
APR March 2024 - 2
APR March 2024 - 3
APR March 2024 - 4
APR March 2024 - 5
APR March 2024 - Message from the Editor
APR March 2024 - Editorial Advisory Board
APR March 2024 - BIOPHARMACEUTICALS - Getting to GMP-Quality Biotherapeutics From Today’s Bench-Scale Continuous Manufacturing Systems: A Gap Analysis
APR March 2024 - 9
APR March 2024 - 10
APR March 2024 - 11
APR March 2024 - 12
APR March 2024 - 13
APR March 2024 - MICROBIOLOGY - Critical Behavioral Attributes and the Application of COM-B Framework in Aseptic Processing
APR March 2024 - 15
APR March 2024 - 16
APR March 2024 - 17
APR March 2024 - 18
APR March 2024 - FORMULATION AND DEVELOPMENT - Use of AUC in AAV Analysis in a GMP Setting
APR March 2024 - 20
APR March 2024 - 21
APR March 2024 - 22
APR March 2024 - 23
APR March 2024 - FORMULATION AND DEVELOPMENT - Precision Medicine in Clinical Trials: A Statistical Perspective
APR March 2024 - 25
APR March 2024 - 26
APR March 2024 - 27
APR March 2024 - An Interview with Dan Smithey, PhD President & CEO, Serán
APR March 2024 - 29
APR March 2024 - 30
APR March 2024 - FORMULATION AND DEVELOPMENT - Still Early Days for AI in Drug Discovery...Says Who?
APR March 2024 - 32
APR March 2024 - 33
APR March 2024 - QC Corner - Enhancing Material and Equipment Availability in Production Isolators
APR March 2024 - 35
APR March 2024 - BIOPHARMACEUTICALS - Technologies for Aseptic Filling: The Choice is Clear
APR March 2024 - 37
APR March 2024 - 38
APR March 2024 - 39
APR March 2024 - Vendor Viewpoint - Data Integrity and Rapid Micro Methods: Transforming to a Modern Microbiology Lab
APR March 2024 - 41
APR March 2024 - MICROBIOLOGY - Bacterial Spore Formers in Disinfectant Efficacy Testing
APR March 2024 - 43
APR March 2024 - Partner Perspective - Nanoparticle Technologies: Enablers for Ocular Drug Delivery
APR March 2024 - 45
APR March 2024 - 46
APR March 2024 - 47
APR March 2024 - DRUG DELIVERY - Your Nails and You
APR March 2024 - 49
APR March 2024 - 50
APR March 2024 - 51
APR March 2024 - DRUG DEVELOPMENT - Battling Exorbitance: High Costs in Sickle Cell Gene Therapies and the Imperative of Global Patient Registries for Equity
APR March 2024 - 53
APR March 2024 - 54
APR March 2024 - 55
APR March 2024 - FORMULATION AND DEVELOPMENT - How Pharma Companies Are Solving Regulatory Challenges with AI-based Technology
APR March 2024 - 57
APR March 2024 - Event Preview - CPHI North America
APR March 2024 - 59
APR March 2024 - Event Preview - Excipient World 2024
APR March 2024 - 61
APR March 2024 - P.I.N. Points
APR March 2024 - 63
APR March 2024 - Advertiser's Index
APR March 2024 - Cover3
APR March 2024 - Cover4
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