Challenges of developing ADA and concentration (PK or PD) assays for lysosomal transgene enzymes on an automated immunoassay platform

Gene therapy can be used to permanently correct genetic disorders by delivering a functional copy of the gene into the nucleus of affected cells or alternative cells, which then release the functional protein into systemic circulation. 

One of the biggest challenges faced by the bioanalytical sector is the lack of a true reference material. As a result, in most assays, an alternative commercially available therapeutic (for example, an enzyme replacement therapy (ERT) product) can be used as a surrogate for the transgene enzyme. This webinar discusses approaches taken in order to monitor the concentration of the expressed transgene product and any associated immunogenicity.

Transgenes are expected to have very low levels of immunogenicity; therefore, immunogenicity assays against transgenes themselves are not widely developed. Nonetheless, it is very important to monitor any potential case of immunogenicity. Current regulatory guidelines are not applicable to transgene products, which adds to the challenge. This case study describes the considerations and challenges encountered when developing and validating an assay for transgene lysosomal enzyme products. Since lysosomal enzymes are sensitive to physiological pH and salt, which is required for the detection of anti-drug antibodies (ADA), specific buffers were required to maintain their optimal configuration. 

The same level of challenges was also encountered during the development of an assay to determine the transgene product concentration, mainly due to the lack of a true reference standard and the presence of the equivalent endogenous molecule in healthy and disease matrices (with the potential that the defective enzyme could still be detected in the ligand-binding assay). Therefore, measurement of the transgene concentration could not be described as a true PK assay and does not conform to current the PK guideline/guidance, requiring an approach more aligned with a PD assay with a well defined context of use.

Attendees will learn about:

• Challenges associated with enzyme immunogenicity assessment
• Measurement of the transgene product - PK or PD?
• Development considerations for non-stable enzyme analytes
• Immunogenicity assay development on the Gyrolab® platform

Issa Jyamubandi

Principal Scientist, Alliance Pharma

John Chappell

Director of Scientific Support - EMEA and Asia Pacific, Gyros Protein Technologies

 

John Chappell has approximately 25 years of experience in the Contract Research industry supporting both preclinical and clinical drug development. He has specialized in supporting biological compounds from an analytical perspective e.g. Pharmacokinetic, Immunogenicity and Biomarker analysis. He is particularly interested in validation requirements and ensuring that data generated will be acceptable to the regulatory authorities.

He has spoken at many international conferences on various topics including Oligonucleotide analysis, Biomarker Analysis, Immunogenicity and the analytical support of Biosimilar programs. He now leads the Application Support and Service teams for Gyros where he is responsible for customer service and technical support in Europe and the Asia Pacific regions. John has been a user of the Gyros system for over 10 years so will use this experience to help customers.

He is a Fellow of the Royal Society of Chemistry and was involved in the American Association of Pharmaceutical Scientists (AAPS) Biosimilar Committee that has prepared papers on Pharmacokinetic and anti-drug antibody assays.