Vector Copy Number Analysis

Vector copy number (VCN) represents the quantity of transgene DNA in genetically modified cells or viral gene therapy products before administration. The FDA raises concerns about potential oncogenicity due to the risk of pathological insertions associated with vector-mediated genetic modification. To address these safety concerns, measuring transgene copies within cell therapy products and quantifying the VCN is required. More integrated transgene copies are believed to enhance product potency as well as increase the oncogenic potential by random vector integration into the host genome. Therefore, the objective is to balance an effective VCN with minimizing associated risks.

FDA approval for clinical use mandates limiting CAR-T VCN to fewer than five vector copies per transduced cell, reducing the number of vector insertion sites in the host genome.

For in vivo administered viral gene therapy products, accurate quantification of the viral genome titer is crucial for dosing and potency analysis. To assess VCN persistence in cell therapies and in vivo administered viral gene therapies, various techniques such as qPCR, dPCR, or NGS are employed to amplify integrated viral vector sequences in genomic DNA obtained from biospecimens. These analyses contribute to gene therapy products’ overall safety and efficacy assessment.