Immune cell function restored in two patients in first-in-human prime editing trial… a review examines achievements and challenges
Precision gene editing advances to early clinical testing
The first-in-human trial of prime editing (PE) reported restored immune cell function in two patients with chronic granulomatous disease. Function was maintained over follow-up periods of 6 and 4 months, respectively, but it is too early to assess long-term effects or a cure. Published in Molecular Therapy, the paper is a review that synthesizes how prime editing works, technical improvements, preclinical research, and early clinical results; it is not an original research article presenting new clinical findings.
A nick in one DNA strand and an RNA template change the sequence
Conventional CRISPR-Cas9 cuts both DNA strands and then relies on the cell’s repair pathways. The small insertions and deletions produced in this process are useful for inactivating genes, but this approach has limitations when precisely correcting disease variants to a desired sequence.
Basic prime editing combines a Cas9 nickase, which cuts one DNA strand, with a reverse transcriptase. The prime editing guide RNA (pegRNA) both directs the system to the target site and provides a template for the corrected sequence. The reverse transcriptase uses this RNA template to synthesize new DNA, and the corrected sequence is incorporated through the cell’s repair process.
Prime editing offers greater flexibility in where editing can occur, but it does not eliminate the constraints imposed by the PAM, an adjacent sequence recognized by the Cas protein. PE3, which adds a nick in the opposite strand, increases efficiency but may also increase unwanted insertions and deletions. Some derivative techniques use double-strand breaks, so not all prime editing can be described as a technology without double-strand breaks.
In the first clinical trial, patient cells were edited outside the body
The clinical cell product PM359, described in the review, involves correcting the delGT variant in the NCF1 gene ex vivo in a patient’s own CD34+ hematopoietic stem and progenitor cells, which are then reinfused. In chronic granulomatous disease, the function of NADPH oxidase, which is needed to eliminate microbes, is impaired. The trial assessed functional recovery by directly correcting the relevant gene variant.
During manufacturing, electroporation was used to deliver mRNA encoding the editing enzyme and chemically modified guide RNAs into the cells. This was not a trial in which nanoparticles or split viral vectors were administered to patients. Busulfan conditioning was also given before cell infusion.
On day 30 after infusion, the proportion of positive cells in the dihydrorhodamine (DHR) assay, which assesses the oxidative function of neutrophils, was 69% and 83% in the two participants, respectively. Neutrophil NADPH oxidase activity was maintained for 6 and 4 months, respectively, and no serious adverse events attributed to PM359 were reported. These findings are early evidence of functional recovery; they do not mean that there were no adverse events at all or that long-term safety has been established. Interpretation is also limited by the small number of participants (2) and short follow-up.
Studies in other diseases show potential and its conditions
In a sickle cell disease study included in the review, about 40% of alleles in patient-derived hematopoietic stem and progenitor cells were corrected. This was an ex vivo cell study result, not a treatment outcome in patients. In a mouse study of hereditary tyrosinemia type 1, corrected liver cells expanded selectively, leading to functional improvement even with a low initial correction rate. However, the same selective growth advantage cannot be assumed in the kidney or nervous system, so results in the liver alone make it difficult to assess the potential for systemic treatment.
The researchers identified miniaturizing the editing enzyme, stabilizing guide RNAs, and improving tissue-specific delivery methods as key challenges for expanding clinical applications. Low editing efficiency in non-dividing tissues also needs to be addressed. Safety assessments should cover not only off-target editing but also unwanted editing products at the target site, large-scale genomic structural changes, and immune responses. The scope of application will depend on validation of efficiency, delivery, and safety for each disease and cell type.
For details, see the full text of the Molecular Therapy review “A primer on prime: A prime editing update from advances to first-in-human trial”.
The advent of CRISPR systems has transformed genome editing, offering unparalleled efficiency and versatility with wide therapeutic potential. However, conventional CRISPR systems face key limitations, including unpredictable and imprecise outcomes during repair of double stranded breaks and reliance on specific protospacer adjacent motif sequences. In response, prime editing (PE) has emerged as a powerful alternative, enabling precise custom edits using a fusion of Cas9 nickase and an engineered reverse transcriptase (RT) together with a prime editing guide RNA (pegRNA) that encodes the desired repair template. PE enables edits to be installed at or downstream of the target site, expanding the range of targetable sequences. Since its inception, PE has undergone extensive optimisation, including Cas variant selection, RT engineering, and pegRNA improvements. In parallel, advances in delivery, including nanoparticles and split viral systems, have accelerated translation across preclinical disease models. Notably, PE has now entered the clinic, with the first in-human study reporting functional restoration with a promising safety profile to date. Here, we summarise recent mechanistic insights, architectural innovations, and therapeutic applications of PE, and discuss the remaining challenges in efficiency, delivery, and safety that will shape broader clinical impact.
The practical value of prime editing lies in its ability to go beyond inactivating genes by directly correcting disease-causing sequences or restoring a protein’s reading frame. Researchers need to assess the proportion of precise correction products, functional recovery, unwanted editing products, and which tissues can be reached, alongside editing rates. In particular, early clinical results in hematopoietic cells that can be collected and reinfused cannot be generalized directly to tissues within the body, such as muscle or the retina. A separate animal study of liver disease also shows that selective expansion of corrected cells may determine the effect. The PM359 results provide early evidence supporting the feasibility of ex vivo prime editing at clinical scale; long-term follow-up and disease-specific validation will determine its future applications.