Development of gene therapies and gene-edited cell therapies has made significant progress in the past decade. Some processing challenges remain, however. Specifically, purification of viral vectors (lentivirus in the case of ex vivo therapies and adenoassociated virus (AAV) for in vivo products) is a complex process because titers are small compared with those of traditional biopharmaceuticals. Moreover, contaminant removal requires multiple filtrations. Such challenges ramp up process costs and extend timelines. Consequently, biomanufacturers need to optimize and streamline their processes. In this Featured Report, BPI’s managing editor presents highlights from the gene therapy tracks at the Cell and Gene Commercialization conference. Authors from PharmaLex focus on manufacturing challenges and regulation issues of advanced-therapy medicinal products (ATMPs), including those raised when reducing chemistry, manufacturing, and controls (CMC) timelines. Authors from BIA Separations (Sartorius) present an approach for simplifying the purification of AAV drug products. And authors from Charles River Laboratories present findings from a study using analytical ultracentrifugation for recombinant AAV therapies.

Introduction: Highlights from Cell and Gene Therapy Conference
by Maribel Rios
Presenters at this year’s Cell and Gene Therapy Bioprocessing and Commercialization conference focused on gene-edited ex vivo therapies and in vivo gene therapies. Topics included data-driven strategies for downstream processing of plasmid DNA, raw material and supply chain strategies, recombinant AAV titration by HPLC, and analytical method development for gene therapies.
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Streamlining Industrial Purification of AAV
by Pete Gagnon, Maja Leskovec, Blaz Goricar, and Aleš Štrancar
AAV is a preferred vector for gene therapy. But unlocking its full potential still poses challenges, many of which are associated with purification. The first involves the transition from upstream to downstream. AAV-bearing lysates are laden with debris that foul filtration media and limit or prevent concentration. Another involves reduction of soluble host-cell DNA, which is complicated by its strong association with nucleoproteins. A third involves elimination of empty capsids. Ultracentrifugation presently meets that need, but scale-up issues make chromatographic alternatives attractive. A fourth challenge involves the need for rapid, accurate, and revealing analytical results to guide process development, support validation, document control and reproducibility of manufacturing processes. The authors share experimental data showing how those challenges can be addressed to advance the evolution of gene therapy with AAV drug products.
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CMC Challenges in Cell, Tissue, and Gene Therapy Development and Regulatory Support
by Anjali Apte, Adeyemi Afuwape, Zaklina Buljovcic, and Zeb Younes
Cell, tissue, and gene therapy (CGT) medicinal products have made tremendous progress in the past decade. These products are considerably different from small molecules or biologics because of their inherent complex nature and variability. Although many unknowns remain about CGT development, the clinical success that they have achieved has enabled this field to advance rapidly. Standard biologics guidelines are not always applicable to CGT products, and guidelines are being actively reviewed and updated by both the FDA and EMA to ensure that companies have more relevant and applicable guidance to help them during the drug development process. The development of CGT products often is under short timelines and require high cost. Many complex but novel approaches to nonclinical and clinical programs have been developed. Along with the clinical development, product quality related to the chemistry, manufacturing, and controls (CMC) must be maintained. The authors highlight challenges encountered in the manufacturing of CGT products and the key regulatory support required to facilitate bringing them to market.
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Ultracentrifugation for Recombinant Adenoassociated Virus Therapies
by Christopher Sucato and Cynthia Swanson
Analytical ultracentrifugation (AUC) assay for characterization of recombinant AAV drug products provides excellent resolution of empty, full, and other AAV product variants. It can meet criteria for a validated method, including precision, linearity, and robustness parameters and can comply with data integrity standards. The authors provide examples demonstrating how AUC assays can be used to determine such parameters for all AAV drug products, irrespective of serotype and transgene size. They also emphasize that contract research organization can leverage resources to provide an AUC service supported by multiple instruments and dedicated personnel.
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