On The Potency Assurance Journey, Travelers Need A Road Map

By R. Mark Jones, Simon Walker, and Jeffrey J. Talbot

Capturing and quantitating the biological effect of a gene therapy candidate in vitro is the most direct and immediate way to support an in vivo clinical program. As such, analytical potency assessment is a universal requirement by global regulatory authorities to support the advancement of therapies from clinical development to registration.

In the realm of advanced therapy medicinal products (ATMP), specifically gene therapies, when and how to adequately and appropriately assess potency is a topic of ongoing debate within the biotech industry. Guidance from contemporary gene therapy documents includes:

• Potency Assurance for Cellular and Gene Therapy Products; Draft Guidance for Industry (FDA),
• Guideline on quality, non-clinical and clinical requirements for investigational advanced therapy medicinal products in clinical trials - Scientific guideline (EMA), and
• Cell and Gene Therapy Validation Challenges (BioPhorum).

These documents outline general expectations and key concepts and provide useful high-level recommendations. However, given the diversity of modalities and complex therapeutic biology, the field struggles to translate these general, high-level requirements into strategies that are reflected in a phase-appropriate basket of assays suitable to demonstrate lot-to-lot manufacturing consistency.

A potency road map devoted to guiding investigators through the challenges of developing, negotiating, and implementing a successful regulatory strategy is becoming necessary as these diverse gene therapies continue to gain traction in the medicinal landscape. Differing and inconsistent phase-dependency of regulatory expectations is difficult and confusing to navigate for both new and experienced potency investigators.

For example, the 2020 FDA guidance for industry on reviewing Chemistry, Manufacturing, and Control (CMC) for Human Gene Therapy Investigational New Drug Applications (INDs) specifically indicates validation of analytical methods is typically not required for Phase 1 submissions. In contrast, the 2025 EMA guideline on quality of advanced therapy medicinal products states “an appropriate degree of method validation should be applied at each stage” and that all safety-relevant assays must be fully validated.

Navigating Local Road Signs: Regional Divergences

While the guidance from these larger, well-established, and guidance-rich national regulatory agencies may be contradictory on the global scale, in direct interactions there tends to be a flexible and collaborative approach. Focusing locally, smaller neighboring and regional agencies often rely on these large agencies’ guidance verbatim, thereby unintentionally limiting the flexibility of the sponsor strategy.

Navigating the additive impact of these regional differences can quickly become a significant program burden, especially when sponsors are targeting ultra-rare indications where sufficient trial populations may not be available in a single region. How then should potency professionals work to create an integrated potency strategy that balances the needs of speed, budget, and a waiting patient population without compromising safety, accuracy, and meaningfulness of data? A strategic road map to visualize the processes may help speed tactical decisions and simplify strategic planning in multiple regions.

Check Your Mirrors: Reflecting On Common Strategy Questions

What should be considered at each step as a program progresses from preclinical development through to pivotal studies and filing, and can a unified strategy be put in place to make those deliverables consistent across the international regulatory spectrum?

One of the challenges investigators face is that each potency assay can be as diverse as the gene, the therapy, and respective disease it’s targeting. Without being overly prescriptive, developing a unified dialogue around the necessary milestones that can be strung together to produce a comprehensive potency strategy would greatly aid industry efforts to produce appropriate and effective potency methods.

Without this road map, the industry finds itself reflecting on how to best address persistent ambiguity around a repeating set of strategy questions.

Are there advantages or disadvantages to various methods?

For example, the commonly employed 50% tissue culture infectious dose (TCID₅₀) assay is used in early program phases to assess infectivity, replication, and then by statistical extrapolation, potency. This classic virology method is highly variable, not tissue-specific, and it is performed in a nonrepresentative reconstituted (supplementing with Rep/Cap) system. It is required by some, but not all, national regulatory agencies.

As a low hurdle to clear to have an early-phase potency assay of record, it is often employed by smaller or more time-constrained companies with the awareness that the data produced is of little value in assessing lot-to-lot variability, effectively kicking the proverbial potency can down the method life cycle road.

Is the strategy phase-appropriate?

Targeting mRNA or protein quantitation post-transduction both offer advantages in assessing potency during early development, especially when the product has a complex or unknown mechanism of action (MoA).

As stand-alone early-phase assays, strong scientific arguments can be made that either method can assess potency (and thereby infectivity more quantitatively than TCID₅₀) of candidate lots and, conceptually, should therefore be modular and interchangeable in their application based on supporting information provided by the investigator. However, the industry sees variable agency directives guiding one over the other or at times requiring both mRNA and protein quantitation to assess early-phase potency.

What surrogate method(s) basket is sufficient?

A surrogate assay(s) is used when an MoA-reflective assay is not achievable but a measurement more closely aligned to functional potency is required.

Surrogate assays are not always cell-based and could be physiochemical-based analytical method(s) if the measured property can be correlated to therapeutic activity relevant to the MoA. These surrogate assays tend to be more amenable to validation and performance in a quality control environment. As such, a well-supported basket of surrogate assays can nicely dovetail with existing potency assays to provide an overall potency strategy that is rigorous and well controlled.

Does the overall strategy appropriately interrogate the intended MoA?

An MoA-specific assay in a tissue-relevant cell line is the universal gold standard for GMP release and stability testing.

Early development cell-based assays that are MoA-relevant should be reviewed from a quality control perspective – meaning, is the assay relatively easy to perform and robust enough to produce consistent results in the stringent GMP operating environment? Reagents and cell lines used in early development as part of target selection may have use restrictions that have to be navigated prior to use in clinical and commercial supporting assays.

Additionally, understanding of the MoA may have changed as the program matured. Therefore, assay performance and criteria should be critically reviewed to ensure they can meet expectations needed for product release.

Setting The GPS: Aligning Strategy For The Road Ahead

Given the age and experience of the industry, there is space on both the regulatory and investigator sides to collaboratively evolve the regulatory landscape in such a way that patients are better, more efficiently served by more meaningful and robust potency data and methods, earlier in the development process.

The BioPhorum ATMP working group has been collaborating on this very challenge over recent months. Soon, they will unveil a Gene Therapy Potency Strategy Roadmap — an interactive aid designed to help developers through the complex decisions to aid planning and propose some simplifications for an ideal future state.

This publication will also spotlight two critical intersections: the reliance on infectivity measures like TCID₅₀ and the challenge of using representative versus nonrepresentative cell lines.

Having shared early insights at key industry conferences in 2025, the group has confirmed the importance of embedding potency strategy early in development planning. The forthcoming publication aims to pave a consistent path for gene therapy developers to follow and help create a foundation toward alignment and uniformity of expectations.

About The Authors:

R. Mark Jones, Jeffery Talbot, and Simon Walker are members of the BioPhorum ATMP group that developed the Gene Therapy Potency Strategy Roadmap.