GeneLancet Biosciences has filed a public comment on FDA’s guidance Human Gene Therapy Products Incorporating Human Genome Editing. The comment is now available on Regulations.gov.
In a 29 February 2024 CBER webinar, Agency staff recommended that,
“For high-performance liquid chromatography (HPLC) and mass spectrometry purity analyses … we recommend the purity of gRNA full-length product to be greater than or equal to 80% and that you identify any impurities that are present at greater than or equal to 1%. However, we do acknowledge that the acceptance criterion can be dependent on the sensitivity of the method that’s used to determine purity. There are multiple different HPLC and mass spectrometry methods that can be used. Therefore, if you’re unable to obtain gRNA
purity of greater than or equal to 80% due to this method sensitivity, we ask that you provide further justification for the proposed acceptance criterion. In your justification, we’d like you to include data on the impurities present and a risk assessment on how these impurities may affect the safety of your product as it pertains to off-target editing.”
However, changing the HPLC method so the main-peak area prints 80% does not create 80% intended guide. It reports an unresolved envelope as full-length product.
Guide RNAs are synthesized 3′ to 5′, scaffold first and spacer last. Truncations and related defects in the spacer are therefore the products of late stage more error-prone synthesis. Many of those species remain long enough to direct Cas9, and they are often contained inside the reported main HPLC peak. A passing full-length number can therefore include sequences that are not the intended guide leading to unpredictable off-targets.
For a genome-editing product, that gap is not only analytical. A sequence defect in a guide RNA drug substance can cause a sequence defect in a patient’s genome. Off-target studies built on the intended spacer will not find cuts programmed by a different spacer in the same vial.
The comment asks CBER to correct the webinar language and to treat spacer-resolved purity as a quality attribute, not as a method-sensitivity footnote.
Read the comment: https://www.regulations.gov/comment/FDA-2021-D-0398-0044
Note: FDA’s posting of the letter is not an indication that the Agency has adopted our position.
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