By PD Dr. Arno Kromminga
Gene therapy has come a long way since it was first suggested as a potential treatment for genetic-based diseases in 1972. Completion of the Human Genome Project and other scientific achievements paved the way for the first gene therapy approval in China in 2003, which led to subsequent approvals in Europe and the United States in 2012 and 2017, respectively. However, as this exciting class of therapy enters a new age, so do the bioanalytical challenges it brings, including those related to immunogenicity assessment. Immunogenicity evaluations inform on the safety and efficacy of a given therapy over time and are necessary for regulatory approval.
Because of their inherent complexity, there are a number of considerations to take into account when performing immunogenicity testing of gene therapy compounds. In our recent webinar, we sought to address some of the most pressing questions regarding this rapidly progressing field, which include the following:
What are the current challenges in immunogenicity assessment of gene therapy compounds?
Gene therapies are multicomponent drugs; they do not simply consist of one molecule, or even one fusion or multidomain molecule. Each part of the drug—the delivery vehicle (or vector), the delivered nucleic acid (or cargo), and the therapeutic protein encoded by the nucleic acid—can trigger a distinct immune response. Consequently, safety and efficacy issues can originate from multiple structural elements that comprise the gene therapy compound.
Gene therapies can also initiate a number of humoral and cellular responses, which demand bioanalytical scientists to assess more than just antibody production. To add another layer of complexity, multiple bridges do exist between innate and adaptive immunity. For instance, activated macrophages can act as antigen representing cells (APC) once they have attacked the phagocyte invader.
Which immunogenicity assessments are recommended from the scientific point of view?
A number of assessments can provide insights into a gene therapy’s overall immunogenicity potential; however, approaches should be specifically tailored to the compound and to the immune responses being evaluated. When assessing cell-mediated immune responses, it is valuable to assess cytokine synthesis, which can be performed with an AID EliSpot Reader platform, which provides a read-out of the activation or proliferation of T cells. This technique can also help identifying the sequence of presented antigen peptides to ultimately guide mutagenesis to de-risk the compound’s immunogenicity potential. Another measurement that can be used for assessing cell-mediated immune responses is immunophenotyping, which can be performed using a flow cytometry platform like the BD FACSLyric™. Differences in immune cell population ratios before and after administration of the compound can be detected by this approach.
A tiered approach is commonly applied when assessing antibody-mediated immune responses. The classical tiered approach includes a screening assay, confirmatory assay, titration, and then characterization of neutralizing activity, isotyping, and epitope mapping. By following this approach, key questions can be answered such as whether the gene therapy has neutralizing antibody activity that can block the binding activity of the drug and impact its effect as well as what the isotypes of the antibodies are. While this classical tiered approach serves as a good general model to follow, it is one of several methods and may not be well-suited for all gene therapeutics.
What immunogenicity risk assessment factors need to be considered?
The route of administration, the transgene, the therapeutic protein, and the vector can all factor into the compound’s likelihood of inciting an immune response. The impact of the route of administration on immunogenicity can vary since the drug formulation changes with the route of administration. Some data has pointed out that some routes are more immunogenic as compared to other routes (such as inhalation for therapeutic monoclonal antibodies for instance).
Nucleic acid cargos can also be immunogenic. Some nucleic acids can constitute danger-associated molecular patterns (DAMPs) and pathogen-associated molecular patterns (PAMPs) and activate the innate immune system. Gene therapies—which contain nucleic acids that encode for proteins—have a much higher immunogenicity risk than those that exert an inhibitory function on gene expression, such as siRNAs. There is also varying immunogenicity among proteins encoded by the transgene, depending on the presence of an endogenous counterpart.
Viral vectors like the adeno-associated virus (AAV) have also shown immunogenicity issues since neutralizing antibodies (NAbs) prevail in a large fraction of the human population, and can impair the AAV’s transduction efficacy. However, strategies can be implemented to circumvent this issue.
Among all of these factors, it is important to abide by the regulatory standards of conducting long-term follow-up (LTFU) for patients administered with gene therapy products. Both the U.S. Food and Drug Administration (FDA) and European Medicines Agency (EMA) outline LTFUs in their respective regulatory protocols. These assessments ensure patients are monitored for delayed adverse events, such as delayed toxicity, and help to ensure patients are kept safe and treated optimally over the long-term.
Thank you again to all of you who joined me for the live session and asked questions. Feel free to reach out to me directly with any additional inquiries about immunogenicity and BioAgilytix’s related capabilities. Our team of expert bioanalysis scientists is ready to support your next project.
Download the On-Demand Bioanalysis Webinar
To access the full recording of the live webinar and Q&A session, click here.