Franklin Biolabs Achieves CLIA Certification for AAV NAb Testing
Learn more
Programmatic Asset
EXECUTIVE SUMMARY
Downstream Filtration and Concentration Techniques for AAV Vectors
CELL & GENE | RNA | BIOLOGICS
Effective downstream processing is a determining factor in the viability of an AAV vector program. The primary objectives are to achieve high purity by removing process-related impurities (host cell proteins, DNA, empty capsids) and to concentrate the vector to a formulation suitable for clinical administration. Franklin Biolabs employs a data-driven approach to develop and optimize purification strategies, including chromatography and filtration techniques, ensuring the final product meets stringent quality attributes required for Investigational Medicinal Product Dossier (IMPD) and global regulatory submissions. This process is foundational to achieving the 18-24 month IND timelines our scientific leadership has historically delivered.
Frequently Asked Questions
How does Franklin Biolabs address the challenge of separating full versus empty AAV capsids during downstream processing?
We utilize advanced analytical ultracentrifugation (AUC) and anion-exchange chromatography (AEX) methods optimized for specific AAV serotypes. Our process development focuses on establishing precise gradient and elution conditions that maximize the separation of therapeutically active, full capsids from immunogenic empty capsids, a quality attribute scrutinized in ATMP submissions to European authorities.
What is the approach for scaling AAV purification from preclinical to GxP-compliant manufacturing?
Our strategy is built on a platform approach that uses scalable technologies from the outset. Methods developed for IND-enabling toxicology studies, such as tangential flow filtration (TFF) and column chromatography, are selected for their linear scalability. This ensures a consistent process and predictable product quality as we move from small-scale runs to the larger volumes required for clinical trials, supported by our >100,000 sq ft facility.
How do you ensure purification processes are suitable for novel or engineered AAV capsids?
A standard purification template does not exist for all AAVs. For novel capsids, our team conducts tailored process development, characterizing the vector’s unique isoelectric point and surface charge properties to customize chromatography resins and buffer systems. This bespoke approach is informed by deep vector biology expertise, ensuring high recovery and purity for next-generation AAV vectors.
Optimizing AAV Purity and Concentration for Clinical Success
The transition from upstream AAV production to a purified, concentrated drug substance is a multi-step process demanding rigorous scientific oversight. Each filtration and chromatography step must be optimized to remove specific impurities while maximizing the recovery of potent, full AAV capsids. Failure at this stage can compromise the safety profile and efficacy of a next-generation therapy.
Our approach to downstream process development is grounded in a deep understanding of AAV biology. The natural diversity and rapid evolution of AAV capsids in vivo (PMID: 12716974) informs our strategy, reinforcing the need for purification platforms that are robust enough to handle serotype variability while being precise enough to isolate the target vector. The approach requires a tailored strategy designed to de-risk clinical translation.
Core Downstream Technologies
A successful purification train for AAV vectors typically involves a sequence of clarification, capture, polishing, and formulation steps.
-
Clarification and Concentration: Initial harvest is clarified using depth filtration or centrifugation to remove cells and debris. Tangential Flow Filtration (TFF) is then employed for volume reduction and buffer exchange, preparing the product for chromatography.
-
Chromatography: Affinity chromatography is often used as a primary capture step for many AAV serotypes (e.g., AAV2, AAV8, AAV9), providing high selectivity. This is followed by polishing steps, such as ion-exchange or hydrophobic interaction chromatography, to remove remaining impurities like host cell proteins and empty capsids.
-
Sterile Filtration: The final step involves filtration through a 0.22 µm filter to ensure sterility before the drug substance is formulated for administration.
This meticulous process is fundamental to minimizing the risk of adverse immune responses from process-related contaminants. Separately, managing in vivo vector behavior is another component of a comprehensive safety profile. For example, research demonstrates that passive transfer of neutralizing antibodies can mitigate non-target tissue biodistribution for CNS-delivered vectors (PMID: 36320416). While distinct from purification, this highlights the level of biological control required for clinical translation. Our team’s “vast knowledge in all aspects of vector production and analytics” ensures that both the purity of the final product and the overall development strategy align with program-specific safety goals.
The Franklin Biolabs team, whose core scientists and leadership maintained a 100% successful IND rate since 2019 prior to our formal 2024 launch, has refined these techniques across hundreds of vector lots. This historical expertise is applied to every program, ensuring that the downstream process is not only scientifically sound but also compliant with global regulatory expectations from the FDA, EMA, and MHRA. For a deeper look at our vector development philosophy, view our recent webinar on initiating successful AAV programs.
This specialized work is part of a larger, cohesive development pathway. For more information on our overarching manufacturing capabilities, please see our parent hub page on Large-Scale AAV Manufacturing and Process Development.
Proven Intelligence in AAV Downstream Processing.
This content is for informational purposes. For guidance specific to your therapeutic program, please contact our team for a consultation.