Gene Therapy Assays: Regulatory Compliance with Unified Immunogenicity

Introduction

Immunogenicity assessment is an integral component of gene therapy development as pre-existing immunity may limit the product’s therapeutic potential. The path from immunogenicity assay development to companion diagnostic (CDx) approval for adeno-associated virus (AAV) gene therapy is complex and time-consuming, especially if the assay is to be used in global clinical trials. To ensure that the right assay is ready at the right time at every stage of a clinical program, it is essential for gene therapy developers to plan ahead.

In this blog, we explore the regulatory frameworks for gene therapy assays in both the US and EU and share best practices for achieving compliance with a global regulatory strategy.

Importance of measuring immunogenicity before gene therapy

AAV vectors are the leading platform for transgene delivery and more than 50% of the general population has some degree of pre-existing immunity to serotypes of AAV. Consequently, the assessment of specific anti-AAV antibodies is a significant consideration in gene therapy development, even if the AAV has been modified or if the serotype used is not commonly seen in humans since there is significant cross-reactivity across serotypes.

The FDA recommends strongly considering the development of a CDx to measure antibodies to investigative gene therapy products. Notably, the agency has required immunogenicity testing, whether a laboratory-developed test (LDT) or a CDx, as a condition for the approval of the five most recently approved gene therapies.

Regulatory considerations for clinical trial assays and CDx in the US and EU

US regulatory framework 

In the US, there are three regulatory pathways available for clinical trial assays (CTAs), based on risk (see Figure 1):

1. Significant risk, which requires an investigational device exemption (IDE)
2. Non-significant risk, which falls under abbreviated IDE regulations where an IDE application is not required, but other requirements such as adverse event reporting and record keeping must be met
3. IDE exempt, which is typically reserved for assays used for research or exploratory analysis

Regardless of the route used for a CTA, institutional review board (IRB) approval is needed before proceeding with a clinical trial.

Figure 1. Risk assessment and regulatory requirements for CTAs

There are multiple factors that go into the determination of significant versus non-significant risk assessment, including:

• Is the assay being used prospectively or retrospectively?
• How is the assay being used? For research, stratification, or inclusion/exclusion?
• Has the assay ever been used in prior investigations with safety data?
• What type of sample is required, and does it involve an invasive procedure?
• If a fresh biopsy is required, is it a significant risk procedure?
• Are the test results used to determine whether the patient receives treatment? If so:
  • Would a false-positive result lead to a patient not receiving a known and effective therapy or standard of care (SOC)?
  • How does the safety profile of the experimental therapeutic compare to SOC?
  • Have patients exhausted all SOC options?

Importantly, the same rules for study risk determination (SRD) apply regardless of the clinical trial phase.

There are two routes for obtaining an SRD. The first is to go directly to the IRB and provide a risk-benefit assessment justifying why an assay is a non-significant risk. The second route is to do an SRD Q-submission with the FDA. The agency strongly recommends this approach since its thinking on gene therapy assays has evolved significantly over the past few years and IRBs may not be aligned with that thinking. In fact, there have been instances where trials have been put on clinical hold because the FDA disagreed with the IRB and deemed an assay to be a significant risk.

When preparing an SRD Q-Submission, it is important to include informed consent forms and a finalized clinical protocol, as changes to the protocol at a later date can invalidate the FDA’s decision on SRD. It is also important to keep in mind that the agency can take up to 90 days to issue an SRD, so this review period should be built into the overall development timeline.

In addition to the SRD Q-Submission, other FDA communications and submissions may include (see Figure 2):

• Pre-IDE Q-Submission, a voluntary pathway whereby the FDA provides feedback on analytical validation studies prior to submission of an IDE application to minimize the risk of rejection. The review period for this type of Q-Sub is 70-75 days, after which the agency provides written feedback. Based on that feedback, sponsors can determine whether to proceed with a meeting with the FDA.
• IDE Submission, which seeks approval for assay to be used in a clinical study.
• Pre-Submission, whereby the FDA provides input on analytical and clinical study designs for CDx development or addresses any other questions about assay strategy.
• Premarket Authorization (PMA), which seeks authorization from the FDA to market a CDx.

Figure 2. Potential interactions with FDA in CTA and CDx development

EU regulatory framework

The definition of CDx differs slightly in the EU compared to the US. If an assay or in vitro diagnostic (IVD) is used to identify patients who are likely to benefit from treatment or who are at an increased risk of adverse events, it would qualify as a CDx in both the EU and US. However, under the EU’s In Vitro Diagnostic Regulation (IVDR), a device that is used to monitor treatment with a medicinal product to ensure that the concentration or relevant substance in the human body is within the therapeutic window is not considered a CDx.[1] It is important to recognize this distinction, as it could impact the regulatory strategy for sponsors seeking therapeutic approval in both markets.

In the EU, a CTA is viewed to be used for medical management decisions of trial participants if it is used for inclusion and exclusion of subjects, treatment allocation, or monitoring of the safety or efficacy of the treatment during the study. In most cases, a CTA used for medical management decisions would be considered a CDx. However, if the assay is used for stratification or endpoint analysis it is not considered to have a medical purpose and would be subject to a lower level of regulatory scrutiny.

If an assay is being used to inform medical management decisions in a clinical performance evaluation, it is subject to numerous requirements, articles, and annexes under IVDR, including Annex XIII and Annex XIV performance study submissions. If the assay was already CE-marked under the In Vitro Diagnostic Directive (IVDD) and is being used within its intended purpose without any significant changes, it can be used in a clinical trial without additional submissions. If it is an in-house test developed and performed in an EU-based facility that qualifies as a health institution, it can be used if all requirements of Article 5.5 of the IVDR are met. In the EU, there is a very specific definition for a health institution, but if a lab qualifies, the test would be exempt from notified body submission under the IVDR.

In the EU, a study that involves simultaneous investigation of a medicinal product and an IVD is called a combined trial and is subject to the requirements of both the Clinical Trial Regulation (CTR) and IVDR. The performance of the assay in a combined trial is called a Clinical Performance Study (CPS) and must meet all the IVDR requirements for a CPS. Unfortunately, there is currently no harmonized procedure for combined submissions for the drug and the assay in a combined trial.

Of note, national competent authority (CA) and ethics committee (EC) review processes and timelines vary widely across the EU (see Figure 3), so it is important for both diagnostic and drug sponsors to work closely together as approvals may take 6-14 months.

To further complicate the assay planning process in the EU:

• Validation requirements are not clear. IVDR adopts state-of-the-art level of assay validation per phase of the clinical study, which is typically seen as Clinical and Laboratory Standards Institute (CLSI). However, in the absence of a pre-submission process, sponsors are not able to get regulatory feedback on the analytical validation of a Clinical Trial Application (CTA).
• EUDAMED is not yet in place. This database is intended to harmonize submissions across all EU countries. Until the clinical module is functional, which could be another few years, the submission process remains decentralized, and sponsors must submit individual country submissions to competent authorities (CAs), each with their own nuances.
• Long timelines for Annex IV submission review and approval. National CA and ethics committee (EC) review processes and timelines vary widely across the EU (see Figure 3). In some cases, EC and CA submissions must be done sequentially, rather than in parallel, further prolonging the timeline.

Figure 3. National CA and EC review timelines

Considerations for assay development

Gene therapy assay development can be costly since there are few, if any, off-the-shelf solutions. As the field evolves with novel capsid types and tissue targeting strategies, assays must be designed to be fit for purpose. For sponsors who have a platform capsid that is being investigated across multiple indications, early consideration of how to build an assay that can be pivoted to support other indications may eliminate the need for redevelopment or revalidation.

Assay design

Key questions to ask when designing an assay include:

• What type of assay is needed, a neutralizing antibody (NAb) assay or a total antibody (TAb) assay?
• Is a clinical cutoff needed, or will all patients with detectable pre-existing immunity be excluded?
• If a clinical cutoff is needed, how will the necessary samples be acquired to determine and validate the cutoff?
• Is pre-existing immunity a safety or efficacy issue, or both?
• Will assay performance testing with concomitant medications be needed?

Another critical question is when that assay may need to become a CDx. If an assay will be used to determine whether or not a patient can enroll in a study, it is likely that it will need to start moving along the diagnostic pathway as early as phase one. In addition, if it is expected that immunogenicity will affect efficacy, a CDx will likely be required. Early conversations with regulatory authorities will help clarify the level of assay validation needed at each stage of development.

Assay validation

Keep in mind that, in the US, Clinical Laboratory Improvement Amendments (CLIA) requirements may not be robust enough to support an IDE application. Often, studies included in an IDE application for a gene therapy CTA require more samples or replicates, more rigorous assessment of endogenous and exogenous interference, and evaluation of sample and control stability.

The EU may generally accept the same level of assay validation accepted by the FDA, though there are terminology differences in the types of validation required during the development process (see Figure 4).

Figure 4. US vs EU assay validation

Assay application in a phase 3 trial

Using a final locked CTA in the phase 3 registrational trial maximizes the likelihood of drug/CDx co-approval and eliminates the need for bridging studies, which can cause significant delays.

Best practices for a global regulatory strategy

Validating an assay is expensive and time-consuming, so it is critical to plan early, to think through all the scenarios, and to talk to regulatory agencies about the strategy. Performing well-designed animal studies will help to inform human assays and saving samples from all stages and studies with the appropriate consent to allow additional testing can help to mitigate risk.

When developing a global regulatory strategy for a gene therapy assay:

• Identify the markets in which clinical studies will be conducted
• Develop an assay from the start that would meet both US and EU requirements
• Consider how the assay is being used in the clinical study and, where feasible, consider an all-comers trial which offers the potential to be relieved from additional regulatory burden
• Tailor the level of validation to study phase
• Understand the exportation laws of human samples in the countries where the study will be conducted
• Plan ahead if the CTA will eventually need to become a CDx
• Interact with regulatory authorities early and often

Conclusion

Planning early and planning well for assay development is essential, and the importance of having an embedded regulatory team cannot be overemphasized. Having regulatory input at the very earliest stages of assay planning helps ensure that the assay is fit for its intended purpose, streamlining development. However, any assay development strategy must be flexible, as it may be necessary to change assay format, clinical cutoff, or even level of validation as study data is generated. Thus, having the right plan at the right time is key.

Precision for Medicine offers a breadth of solutions for accelerating therapeutic and diagnostic development, starting with specialty assays and immune monitoring that can be executed in global clinical trials. We also provide regulatory consulting and quality biospecimens for assay development and validation and have managed more than 200 IVD and CDX submissions. Precision for Medicine has supported more than 15 AAV-focused gene therapy companies and their projects across AAV serotypes, assay types, and therapeutic areas, including rare diseases. We also developed and validated the LDT and CE marked assay for Hemgenix® and serve as the sole testing site.

Learn more about how Precision for Medicine can help with global strategies for immunogenicity assays in AAV gene therapy.

References

[1] Official Journal of the European Union. REGULATION (EU) 2017/746 OF THE EUROPEAN PARLIAMENT AND OF THE COUNCIL of 5 April 2017 on in vitro diagnostic medical devices and repealing Directive 98/79/EC and Commission Decision 2010/227/EU. Available at https://eur-lex.europa.eu/legal-content/EN/TXT/PDF/?uri=CELEX:32017R0746.

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