Translating CRISPR to Practice in the Clinic: Transformative Therapy in Hemoglobinopathies and Emerging Applications in Malignancies.

LNP-mediated BCL11A Editing Corrects Sickling Phenotypes and Preserves HSPC Fitness Compared to Electroporation

Gene editing of hematopoietic stem cells: applications and advances.

Gene therapy approaches for inborn errors of immunity: from bench to bedside.

INTRODUCTION Inborn errors of immunity (IEI) are rare genetic defects that disrupt immune function, often resulting in life-threatening infections, malignancies, and immune dysregulation. Allogeneic hematopoietic stem cell transplantation (HSCT), a curative option for some diagnoses, is limited by donor availability and risks of graft-versus-host disease. This review explores the 30-year evolution of autologous gene therapy as a vital alternative to allogeneic hematopoietic stem cell transplantation for IEIs. AREAS COVERED Literature search using PubMed for gene therapy for IEI in the last 20 years. We trace the transition from early gamma-retroviral gene addition - which successfully restored immunity in severe combined immunodeficiency (SCID) but carried high risks of insertional mutagenesis and leukemogenesis - to the adoption of safer self-inactivating lentiviral vectors. The field is rapidly advancing beyond viral gene addition toward highly precise gene editing technologies, including CRISPR/Cas9, and base/prime editing, which offer targeted correction with minimized genotoxicity. EXPERT OPINION Recent milestones in diseases like Wiskott-Aldrich syndrome (WAS) and chronic granulomatous disease (CGD) highlight enormous scientific success, yet significant barriers to accessibility, manufacturing, and affordability remain. Overcoming this requires innovative regulatory frameworks and collaborative funding models. Streamlining development and ensuring equitable access are essential next steps to establishing gene therapy as a safe alternative.

Unlocking the Curative Potential of Gene Transfer and Editing for Hematopoietic Disorders.

Hematopoietic stem and progenitor cells (HSPCs) gene therapy may transform the therapeutic landscape for inherited hematological disorders and already offers potentially curative options for some diseases, including inborn errors of immunity and β-hemoglobinopathies. Its continued success relies on further refinement of gene transfer technologies, gene editing tools such as CRISPR-Cas, and optimized ex vivo HSPC manipulation protocols that ensure robust, long-term engraftment and clonal diversity with reduced-toxicity, non-genotoxic conditioning strategies. Here, we review recent developments and refinements in gene transfer and editing technologies for HSPCs, while also discussing the critical limitations and hurdles to clinical translation, as recently presented at the New Investigator Committee Gene Therapy webinar. Future directions must prioritize integrating technological innovation with the development of equitable and simplified models to reduce costs and ensure that these life-saving cellular therapies reach patients worldwide. Teaser abstract Hematopoietic stem and progenitor cells (HSPCs) gene therapy is advancing rapidly, with lentiviral gene transfer, genome editing, and emerging in vivo delivery approaches expanding the therapeutic landscape for inherited hematologic disorders. Recent clinical successes have demonstrated the potential for durable correction, while ongoing refinements continue to improve safety, efficacy, and feasibility. Key challenges remain in genotoxicity, conditioning toxicity, manufacturing scalability, and equitable global access.

Implementing a First-in-Human Clinical Trial of Prime-Editing Gene Therapy in p47-CGD: Clinical Data and Site Perspectives

In April 2025, CHU Sainte-Justine performed the world’s first gene therapy using prime editing. The patient was an 18-year-old male with autosomal recessive p47phox chronic granulomatous disease (CGD). Infectious history included osteomyelitis, abscesses, and Burkholderia cepacia pneumonia. He also had an active colitis with intermittent oral ulcers, managed with Pentasa. An allogeneic hematopoietic cell transplantation (HCT) had been dismissed during childhood in the absence of a compatible donor. At fall 2024, the opportunity arose for the patient to enroll in this gene therapy clinical trial to achieve a permanent cure for his disease. After completing the screening, apheresis, and busulfan conditioning phases, the patient received the investigational product, PM359, manufactured from autologous CD34+ cells in which the GT deletion in NCF1 was corrected using prime editing. Aside from mild adverse events related to busulfan conditioning, no serious events occurred. Neutrophil engraftment was achieved on Day 16 and platelet engraftment on Day 19. NADPH oxidase activity in neutrophils, measured by dihydrorhodamine (DHR), increased from less than 1% pre-infusion to 69% by Day 30. To this day (9 months post-infusion), the DHR remains stable at 77%, and the patient has no CGD manifestations and is no longer on medication. Another 57-year-old patient with a long history of infections of lung, liver, and lymph nodes plus inflammatory bowel disease (IBD) received the same treatment at the National Institutes of Health (NIH), and the outcome was also excellent and uneventful. The DHR normalized for 83% of neutrophils, and this result was stable after 6 months. He is no longer on prophylactic antibiotics, and his IBD has significantly improved. While our patient’s clinical course was simple, setting up the clinical trial required acrobatics and coordination worthy of a hyperactive octopus. Implementing a first-in-human therapy for patients whose lives are not immediately at risk represents a high degree of complexity. This success required nearly two years of preparation, from initial contact with the trial sponsor to PM359 administration. Through the preparation of the site qualification and audit days, it was essential to surround ourselves with the right resources—within the cell therapy laboratory, the apheresis unit, and the quality assurance team. Beyond having those resources in order, one must know how to present them, clarify them, and adapt to the sponsor’s requests, while ensuring compliance with existing procedures. We orchestrated the treatment plan with our clinical teams to align as closely as possible with standard practice while anticipating tools to mitigate uncertainty when deviations were necessary and making sure no one was overlooked: nurses, physicians, nutritionists, laboratories, pharmacy, imaging, and more. In parallel, we had to prepare a robust ethics submission and a consent form that says everything—without being overwhelming. Finally, it culminated in presenting the trial to the patient and his parents and guiding their decision-making, leading to the results presented above. Our objective is to present the outcome of those patients together with the description of all the steps of the clinical trial, likely including a video testimony of our patient.

Non-genotoxic transplantation and in vivo selection through epitope editing.

The short-term and long-term effects of genotoxic pre-transplant conditioning remain barriers to the broader application of haematopoietic stem/progenitor cell (HSPC) transplantation and gene therapies1-4. Although monoclonal antibodies targeting KIT have been proposed as alternatives to chemotherapy or radiotherapy5-7, their pharmacokinetics hinder clinical applications owing to the risk of depleting transplanted HSPCs. Here, to address this issue, we identified amino acid changes in the extracellular domain of KIT that disrupt the binding of two therapeutic monoclonal antibodies8,9, which impair stem cell factor (SCF)-mediated signalling without affecting KIT expression or functionality. We exploited adenine base editing10 or prime editing11 to efficiently introduce these mutations in HSPCs and combined them with the disruption of the BCL11A erythroid enhancer to promote expression of fetal haemoglobin (HbF)12,13, a therapeutic approach for several haemoglobinopathies. This strategy enables in vivo co-selection of gene-engineered cells to reach the threshold required to provide therapeutic benefit in patients affected by sickle cell disease and β-thalassaemia. We show progressive enrichment of KIT plus BCL11A multiplex-edited haematopoiesis under selective pressure with KIT monoclonal antibody, in vitro and in vivo. We report that extended treatment with anti-KIT regimens leads to superior in vivo enrichment while avoiding clonal selection, as assessed by a lentiviral barcoded library. Finally, by overcoming the limitations of monoclonal antibody pharmacokinetics, epitope editing enables novel haematopoietic replacement regimens that are not limited by on-target graft elimination, allowing prolonged immune-based conditioning that maximizes haematopoietic niche clearance without chemo-radiotherapy or monoclonal antibody wash-out.