FDA's Updated Guidance On Human Genome Editing: New Implications & Remaining Questions

By Brent Morse and Nate Manley, Dark Horse Consulting

In March 2022, the U.S. FDA released the draft guidance for industry Human Gene Therapy Products Incorporating Human Genome Editing. The purpose of this draft document was to share preliminary FDA thoughts about proper implementation and control of human genome editing (GE) in therapeutic product development and to invite commentary from the scientific community about potential improvements to the guidance. Numerous scientific organizations and interested parties, including the American Society for Cell and Gene Therapy and the International Society for Cell Therapy, among others, provided feedback to the FDA about how to improve the clarity and utility of the guidance. On Jan. 29, 2024, the FDA released the final version¹ of the guidance document.

The now finalized guidance document is an important resource for the industry as it provides an overview of FDA expectations for GE products, whether they be ex vivo modified cell-based products or in vivo targeted gene therapies. The guidance provides general recommendations for chemistry, manufacturing, and controls (CMC), including analytical testing expectations for GE components and resulting drug product (DP), and what GE component/DP manufacturing information should be included in a sponsor’s IND submission. The second section of the guidance highlights some key considerations for nonclinical development, including test article selection for IND-enabling nonclinical studies and key endpoints for assessment of product activity and safety. The final set of recommendations focuses on clinical development and includes considerations pertaining to study population selection, DP dosing and delivery, trial design, safety monitoring, and study endpoints. Collectively, this guidance provides an informative set of recommendations that span the GE product development trajectory.

So, what changed between the draft and now final version of the guidance and, perhaps more importantly, what open questions remain for the industry and scientific community to help resolve? In the next section of this article, we highlight key updates introduced in the finalized guidance and their implications for the industry. Subsequently, we delve into areas of GE product development not explicitly covered by the guidance, what we can learn from current industry best practices and trends, and remaining challenges for the field.

What’s New In The Final Guidance?

Place the draft guidance and final guidance side-by-side, and generally they appear very similar with respect to structure and content. However, closer scrutiny reveals some interesting changes in the final guidance that are impactful to the cell and gene therapy (CGT) industry. This section highlights key changes found in the final GE guidance and considers their potential implications for both developers of GE products and suppliers of GE components. Arguably, the more substantive updates reside in the CMC and nonclinical sections of the GE guidance, and thus they are the primary focus of this section.

Within the CMC section of the guidance, the first noteworthy change is added language that distinguishes between gene editing performed as part of routine DP manufacturing (i.e., performed for each new lot of resulting DP) versus gene editing performed once, upstream of a cell bank that will serve as the cellular starting material for DP manufacturing. Specifically, the final guidance indicates that for one-time genome editing (as would be done to generate a GE master cell bank (MCB), for example), it may be acceptable to utilize GE components with reduced manufacturing information and reduced release testing. The implications of this are particularly relevant to GE products derived from pluripotent stem cells (PSCs), for which an increasingly popular approach is to perform the GE step prior to generation of a PSC seed bank that is subsequently used as starting material for MCB production; the MCB then serves as the starting point for working cell bank (WCB) production or is used directly for DP manufacture. To maximize cost- and time-saving benefits, GE seed bank production often will utilize GE components not manufactured in accordance with good manufacturing practice (GMP) guidelines and will be performed in a non-GMP lab environment. The success of this approach generally relies on three main factors, namely 1) implementation of sufficient control strategies for the non-GMP GE components and lab environment, 2) downstream production of a clonally derived MCB in a fully GMP-compliant environment, and 3) comprehensive MCB testing for safety and genome integrity to ensure its suitability as WCB or DP starting material.² Acknowledgement of this approach is important for GE component suppliers, as it indicates that there likely will continue to be demand for small batch, non-GMP GE components, and it is a clear win for developers of GE PSC-based products. As noted in the guidance, sponsors interested in this approach are highly encouraged to seek FDA buy-in via early engagement.

Within CMC, a second noteworthy set of changes is found within the guidance’s updated analytical recommendations. First, the new guidance presents a potential phased approach to potency testing, specifically with respect to the genetic modification. In early phases, demonstration of the sequence modification may be an adequate measure of potency, whereas for late-phase studies, a functional consequence of the sequence modification may be required. This is how many developers have approached potency for gene edited products thus far, but an explicit acknowledgement of the acceptability of this approach should serve to reduce ambiguity among sponsors and also nicely aligns with the concept of a potency assurance strategy as articulated in the 2023 update to the cell and gene therapy potency guidance.³ Another key change in the final guidance concerns testing for off-target editing. While the draft guidance includes the suggestion that drug product testing of gene edited products “should include” analysis of off-target editing, the final guidance suggests that drug product testing “may include” off-target analysis, dependent on the result of nonclinical studies to assess the potential impact of off-target editing. Although seemingly subtle, this is an important change to the language in the guidance, as it may enable a reduction in analytical testing burden provided there is sufficient nonclinical characterization of the prevalence and potential impact of off-target editing. Again, this change reflects how many developers are already approaching this issue, so it is great to see further alignment with regulators.

The second set of significant updates to the GE guidance resides within its coverage of nonclinical development. In line with the FDA’s 2013 CGT preclinical guidance,⁴ the GE guidance walks through the essential components of a GE product’s nonclinical package, including 1) safety studies to identify potential treatment-associated toxicities, 2) a biodistribution study to characterize the product’s tissue/organ distribution over a time frame representative of its expected persistence in humans, and 3) a proof-of-concept (PoC) efficacy study demonstrating the product’s potential therapeutic benefit. Potentially impactful changes are found within the FDA’s discussion of each of these nonclinical development objectives and are summarized hereafter.

Within the context of assessing product safety, the finalized guidance indicates that studies should include characterization of off-target and on-target editing capacity of GE products (previously only calling out off-target editing). The purpose of this addition likely is to acknowledge the difference between productive and non-productive on-target edits, which can be impactful to product safety or activity. For example, a GE strategy that is meant to knock-out a deleterious gene may be on-target in terms of occurring at the intended genomic loci but may result in varying frequencies of productive gene silencing, non-productive editing (e.g., silent mutations), or even deleterious changes in gene function or immunogenicity. Understanding the frequency of these different on-target events may inform product dosing or safety monitoring. Specific to in vivo GE products, the recommended analysis of on- and off-target editing also has been expanded to include all major cell types in which editing is detected, as informed by biodistribution studies.

Two elements of in vivo GE product characterization previously captured under safety assessments, namely kinetic assessment of GE component expression/activity and potential for inadvertent germline modification are now covered under assessment of biodistribution. The relocation of inadvertent germline editing from safety to biodistribution is particularly interesting given the recent, although ultimately assuaged, concerns about potential germline editing of Intellia’s NTLA-2002.⁵ While this change suggests that FDA may be less concerned about potential germline modifications than it was in 2022, it also is a safe bet that if a GE product’s biodistribution profile includes localization to germline tissues, an associated on- and off-target editing safety package will be requisite. Lastly, in the context of biodistribution, the guidance now acknowledges that biodistribution assessments may be included as part of PoC efficacy studies or safety studies and, as a result, are not explicitly required to be conducted in accordance with Good Laboratory Practice (GLP) guidelines. While this represents a potential cost-saving opportunity for sponsors (i.e., avoiding costs associated with GLP compliance), it is important to note that biodistribution studies must be properly controlled/documented and utilize a validated and quantitative analytical method with appropriate sensitivity.

Perhaps the most curious update can be found within the section on Assessment of Activity, which no longer recommends that sponsors assess for potential relationships between GE activity and genetic variation within their intended target population. Considering that this was a key focus of last year’s FDA advisory committee meeting for Vertex’s now approved CASGEVY, it is interesting that the FDA has decided not to include this recommendation in the finalized guidance. Although human genetic variation may not be expected as part of a GE product’s nonclinical package, the agency may continue to look for such data sets as more GE products gain clinical experience.

A final set of noteworthy updates to the nonclinical section relates to proper selection of drug product test article for nonclinical studies. Long has the mantra been that test article used for IND-enabling nonclinical studies should be appropriately representative of the intended clinical drug product. To the surprise of many (including these authors), the draft GE guidance specifically called out a need to demonstrate comparability between nonclinical and clinical DP. This term has been removed from the finalized guidance, indicating a welcome return to prior regulatory expectations. In addition, the nonclinical section contains even further evidence of the evolving regulatory mindset with the acknowledgement that sponsors can consider the use of in vitro or even in silico studies in lieu of in vivo/animal studies with appropriate regulatory buy-in (e.g., through INTERACT or pre-IND regulatory meetings). Lastly, and most wisely, this section encourages sponsors to save retains of test articles used in IND-supporting nonclinical studies for potential future analyses. As discussed in the next section, the field of GE is fast-paced and ever-expanding. As new discoveries unfold and more powerful analytical techniques emerge, it is more or less a given that we will need to continuously refine how we use and assess GE products.

But before we move to the topic of what is not addressed by the GE guidance, there is one key addition to the clinical section that deserves recognition. Based on input from the scientific community, the last clinical subsection on Study Endpoints now affirms that human GE products may be eligible for accelerated approval, providing the associated requirements are met. Representing an opportunity to significantly reduce the clinical development timeline, this is another important win for the field.

Unanswered Questions & Remaining Challenges

The finalized guidance expands upon and clarifies many of the key topics of interest to developers and addresses several questions that have come up since the draft guidance was published in 2022. However, there are questions that remain regarding application of the final guidance to many of the complex situations that are encountered by developers at the cutting edge of gene editing technology.

One prominent example is the case of gene editing reagents intended for one-time use, such as in the creation of an MCB. While the language in the final guidance helps relieve a significant amount of burden on developers by providing options for reduced manufacturing information and release testing, the question of what amount of information is “sufficient to support the quality of the GE component” can be a daunting one for developers. For example, although the reagents used for the MCB may not need to be manufactured under GMP, a minimal level of manufacturing control will be expected and, therefore, sponsors will need to think carefully about what “minimal control” means within the context of their manufacturing process. For example, the sponsor should still scrutinize the source of raw materials used in the editing reagent manufacturing processes, and even though the reagents may be produced in a development environment, rather than a GMP suite, the sponsor should consider the additional uses of that environment (e.g., what else is being or has been cultured in the biosafety cabinets, and what else has been stored in the incubators? What impact might the answer to these questions have on exposure to pathogens?). Likewise, exhaustive testing of the MCB will allow the sponsor to reduce release and characterization testing of gene editing reagents. However, the details of how this reduced testing regimen is implemented will be critical. Sponsors may consider reducing the breadth of testing (e.g., dropping the use of some assays), the depth of testing (relaxing qualification requirements and/or acceptance criteria), or both. These decisions should be guided by a comprehensive risk assessment that considers potential effects of a reduced testing regimen to the drug product and to the patient and how the proposed MCB testing may mitigate those risks.

Assessing the off-target editing risk will likely continue to be a challenge for developers, even with the refinement presented within this guidance. Although a reasonable conceptual framework for identifying and confirming off-target editing events has emerged, implementing this in a real-world situation can be complex. Basic questions regarding the expected depth and breadth of off-target testing remain front and center, especially as technological developments continue to extend the capability to interrogate potential for both off-target genomic events and unintended on-target genomic events⁶ at a rate that may outpace our ability to assess their risk. This is further exacerbated by a rapidly evolving landscape in the technological development and potential use of gene edited products. Recent innovations such as base editing,⁷ PRIME editing,⁸ and gene writing⁹ technologies may employ multiple targeting approaches and enzymatic activities that must be accounted for. Likewise, as the field is expected to move into riskier technological and therapeutic areas, e.g., in vivo editing applications, broader patient populations, or into first- and second-line therapies in the case of oncology, the risk-benefit calculus will likely need to be continually reevaluated.

Summary

The final genome editing guidance represents a step forward as a useful tool for developers to understand the agency’s thinking regarding development of GE products and presents a solid framework for developers to initiate key activities such as evaluation of off-target editing and development of potency assays. However, the dynamic nature of GE technology as well as its expected expansion into larger therapeutic populations will continue to present challenges for developers beyond the now refined guardrails of the GE guidance and demand some degree of bespoke development strategies for some time yet to come.

References

1. Guidance for Industry: Human Gene Therapy Products Incorporating Human Genome Editing. Center for Biologics Evaluation and Research. 2024.
2. Considerations for development of gene-edited PSC-based therapies. Morse, Brent and Mack, Amanda. 9, 2023, Considerations for development of gene-edited PSC-based therapies, Vol. 9, pp. 1341-52.
3. Draft Guidance for Industry: Potency Assurance for Cellular and Gene Therapy Products. Center for Biologics Evaluation and Research. 2023.
4. Guidance for Industry: Preclinical Assessment of Investigational Cellular and Gene Therapy Products. Center for Biologics Evaluation and Research. 2013.
5. https://ir.intelliatx.com/news-releases/news-release-details/intellia-therapeutics-announces-fda-clearance-investigational. Accessed February 20, 2024
6. Off-Target Analysis in Gene Editing and Applications for Clinical Translation of CRISPR/Cas9 in HIV-1 Therapy. Atkins, Andrew, et al. 2021, Frontiers in Genome Editing, Vol. 3, p. 673022.
7. CRISPR-Cas9 base editors and their current role in human therapeutics. Lahr, Walker S, et al. 3, 2023, Cytotherapy, Vol. 25, pp. 270-76.
8. Prime editing for precise and highly versatile genome manipulation. Chen, Peter J and Liu, David R. 2023, Nature Reviews Genetics, Vol. 24, pp. 161-77.
9. Drag-and-drop genome insertion of large sequences without double-strand DNA cleavage using CRISPR-directed integrases. Yarnall, et al. 2022, Nature Biotechnology, Vol. 41, pp. 500-512.P

About The Authors:

Brent Morse, M.S., is a principal consultant at Dark Horse Consulting Group with more than 20 years of experience in the biotech industry, including eight years of experience in cell and gene therapy. Earlier in his career, he led analytical development at CRISPR Therapeutics and was VP of process and analytical development at Vor Biopharma. At Dark Horse, Morse’s areas of focus include analytical development, process development, external manufacturing, and CMC strategy.

Nate Manley, Ph.D., is a senior principal consultant and head of nonclinical at Dark Horse Consulting Group with 11 years of industry experience and over 20 years of experience in the field of cell and gene therapy. Prior to joining Dark Horse, he led research and preclinical development efforts at Asterias Biotherapeutics and trained as an academic researcher in the departments of biology and neurosurgery at Stanford University. At Dark Horse, Manley specializes in nonclinical and analytical development strategy, regulatory submissions, and path to FIH operational planning.