Executive Summary

Scaling adeno-associated virus (AAV) vector production from preclinical to clinical-grade volumes introduces process changes that require a robust comparability exercise to satisfy MHRA review for an Investigational Medicinal Product Dossier (IMPD). A successful strategy depends on a predefined comparability protocol, a deep understanding of AAV key quality attributes (CQAs), and a suite of validated analytical assays capable of detecting minor process-related variations. This framework ensures that the scaled-up vector maintains the same identity, purity, potency, and biological characteristics as the original material used in IND-enabling toxicology studies.

Establishing a Baseline for AAV Process Comparability

A defensible comparability package for the MHRA begins long before scale-up. It starts with a well-characterized and reproducible manufacturing process at the pilot scale. Establishing a robust method for generating high-purity vectors, such as utilizing PEI-mediated transfection in HEK293 cells followed by iodixanol gradient centrifugation, creates a consistent analytical baseline (PMID: 20497038). Without this foundational data, demonstrating that a scaled-up process yields an equivalent product becomes analytically challenging and introduces regulatory risk.

The objective is not to prevent all process changes : it is to control and understand their impact. For AAV vectors, even minor shifts in upstream or downstream parameters can influence key quality attributes.

Key Quality Attributes in a Scaled-Up Environment

For any AAV program targeting UK and EU clinical trials, the comparability protocol must prospectively define the analytical methods and acceptance criteria for the vector’s CQAs.

• Identity & Purity: Verifying the AAV serotype, vector genome integrity, and capsid protein ratios (VP1:VP2:VP3) remains fundamental. Purity assessments must confirm the efficient removal of process-related impurities like host cell proteins and DNA.

• Quantity & Potency: Vector genome titer and the percentage of full capsids are primary metrics. A validated, cell-based potency assay is indispensable for demonstrating that the biological activity of the vector is unchanged after scale-up.

• Biological Characteristics: The in vivo performance of an AAV vector is directly linked to its analytical profile. A robust data package is required to provide regulators with confidence that product attributes remain consistent post-scale-up, ensuring predictable behavior in the intended patient population.