CRISPR FDA approval

CRISPR-Based Gene Therapies: From Preclinical to Clinical Treatments"

In recent years, CRISPR/Cas9 gene editing technology has revolutionized genetic medicine, offering promising therapeutic avenues for inherited disorders affecting various organ systems, notably blood diseases and neuromuscular conditions like Duchenne muscular dystrophy (DMD). The article by Laurent et al. (2024) provides a comprehensive overview of how CRISPR-based therapies have transitioned from preclinical studies to clinical treatments, highlighting significant milestones, successes, and challenges in the field.

CRISPR/Cas9 functions by utilizing a single-guide RNA (sgRNA) to direct the Cas9 nuclease to specific DNA sequences, inducing double-strand breaks (DSBs) that can be repaired by the cell's natural mechanisms. These repair pathways include non-homologous end joining (NHEJ), which can introduce insertions or deletions (indels), and homology-directed repair (HDR), which uses a DNA template for precise correction. Advancements in CRISPR technology have led to the development of base editors (BEs) and prime editors (PEs), enabling precise genetic modifications without inducing DSBs. BEs can convert specific base pairs—cytosine to thymine (C•G to T•A) or adenine to guanine (A•T to G•C)—potentially correcting around 60% of pathogenic point mutations. PEs expand this capability by facilitating targeted insertions, deletions, and all 12 possible base-to-base conversions.

In the realm of hematological disorders, β-hemoglobinopathies such as sickle cell disease (SCD) and β-thalassemia have been significant targets for CRISPR-based therapies. These conditions affect approximately 400,000 births worldwide annually and result from mutations in the β-globin gene (HBB), leading to defective hemoglobin function. One therapeutic strategy involves reactivating fetal hemoglobin (HbF) by disrupting the B-cell lymphoma/leukemia 11A (BCL11A) gene, a repressor of HbF expression. Clinical trials using CRISPR/Cas9 to disrupt the erythroid-specific enhancer of BCL11A have shown remarkable success. Notably, Vertex Pharmaceuticals and CRISPR Therapeutics developed CTX001 (now known as Casgevy), which edits patient-derived hematopoietic stem/progenitor cells (HSPCs) ex vivo.

In clinical trials for SCD (NCT03745287) and transfusion-dependent β-thalassemia (TDT) (NCT03655678), patients achieved editing efficiencies of approximately 80% in HSPCs. This high efficiency led to significant increases in HbF levels—up to 40% of total hemoglobin in some cases. SCD patients experienced a complete elimination of vaso-occlusive events, while TDT patients became transfusion-independent. These groundbreaking results culminated in the FDA's approval of Casgevy, marking it as the first CRISPR/Cas9-based drug for severe SCD and TDT.

Other clinical trials have reported similar successes. For instance, Bioray Laboratories' BRL-101 therapy showed that patients exhibited an editing rate of 85% in the bone marrow, leading to clinically meaningful rises in HbF levels and transfusion independence over an 18-month follow-up. EdiGene's ET-01 also demonstrated promising outcomes, with patients achieving significant increases in HbF production.

An alternative approach targets the γ-globin gene promoters (HBG1/2) directly. By disrupting repressor binding sites using CRISPR/Cas9 or Cas12a nucleases, researchers have significantly increased HbF production. Clinical trials using Cas12a (e.g., NCT04853576 for SCD) have shown up to a 40% increase in HbF levels in vivo. Base editors have also been employed to introduce specific mutations associated with hereditary persistence of HbF. Beam Therapeutics is conducting a clinical trial (NCT05456880) using adenine base editors to modify HBG1/2 promoters, aiming to achieve sustained HbF production without inducing DSBs.

While reactivating HbF addresses the symptoms, correcting the underlying mutation in HBB offers a curative approach. Using HDR, researchers have attempted to correct the SCD mutation directly. However, challenges such as low efficiency and potential genotoxicity have limited clinical translation. Alternative strategies using base editors have shown promise. An engineered adenine base editor corrected up to 80% of SCD alleles in patient-derived HSPCs, resulting in up to 72% production of a non-pathogenic hemoglobin variant (Hb-Makassar) in differentiated erythroid cells.

Prime editing has also been explored for directly correcting the SCD mutation. Delivering optimized prime editors into SCD HSPCs achieved up to 41% allele correction. In mouse models transplanted with these edited cells, HbA expression was restored in 42% of red blood cells. This method holds potential for treating patients without inducing DSBs or relying on donor templates.

In primary immunodeficiencies, CRISPR-based therapies have targeted conditions like X-linked hyper-IgM syndrome (XHIM), RAG1 deficiency, chronic granulomatous disease (CGD), and Wiskott–Aldrich syndrome (WAS). By precisely inserting corrective cDNA sequences into the defective gene loci using CRISPR/Cas9 and AAV6 vectors, researchers have restored normal gene function. For instance, in XHIM, correction of the CD40L gene in patient-derived T cells and HSPCs led to restored protein expression and functionality. In WAS, gene editing restored WAS protein expression in HSPCs, leading to improved functionality compared to lentiviral transduction methods.

Moving to neuromuscular disorders, Duchenne muscular dystrophy (DMD) has been a significant focus for CRISPR-based interventions. DMD affects about 1 in 5,000 male births globally and is caused by mutations in the DMD gene, leading to the absence of functional dystrophin protein. CRISPR/Cas9 strategies have aimed to restore dystrophin expression by deleting exons to restore the reading frame, correcting point mutations, or upregulating compensatory genes.

Preclinical studies have demonstrated the feasibility of these approaches. In mdx mice, a common DMD model, deleting exon 23 using CRISPR/Cas9 led to dystrophin expression levels reaching up to 92% in the heart and significant improvements in skeletal muscle function. In large animal models like dogs and pigs, similar strategies have shown therapeutic benefits, indicating potential for clinical translation. For example, in DMD dogs treated with CRISPR/Cas9 to delete exon 51, dystrophin levels ranged from 3% to 90% of normal, with up to 92% restoration in the heart.

Base editors have been employed to correct point mutations in the DMD gene without introducing DSBs. In one study, an adenine base editor corrected a nonsense mutation in exon 20 of the DMD gene in mice, resulting in dystrophin expression in 17% of skeletal muscle fibers. Another study achieved near-complete rescue of dystrophin in the heart and 15% rescue in skeletal muscle fibers ten months post-injection using an optimized base editor. These results demonstrate the potential for long-term therapeutic benefits.

Prime editing offers further potential by enabling correction of a wide range of mutations, including insertions, deletions, and all base substitutions. While still in early stages, prime editing has been successfully applied in vitro to correct mutations in DMD patient-derived cells, restoring dystrophin expression to 24.8–39.7% of healthy levels. This method could address various DMD mutations without inducing DSBs or requiring donor DNA templates.

Despite these advances, challenges remain in translating CRISPR-based therapies for DMD to the clinic. Systemic delivery of CRISPR components to muscle tissues poses significant hurdles, including immune responses to high doses of viral vectors and the need for efficient, tissue-specific targeting. The first CRISPR-based clinical trial for DMD (NCT05514249) aimed to upregulate a compensatory gene using CRISPR activation but faced safety challenges, underscoring the need for careful optimization of delivery methods and dosing.

Comparatively, the clinical translation of CRISPR therapies has progressed more rapidly in hematological disorders than in neuromuscular diseases like DMD. This disparity is largely due to the ability to perform ex vivo editing of HSPCs for blood disorders, allowing for controlled editing and screening before transplantation. In contrast, DMD requires in vivo delivery to muscle tissues, which is more complex and presents additional safety concerns.

In conclusion, CRISPR-based gene therapies have made remarkable strides from preclinical models to clinical applications, particularly in hematological disorders like SCD and β-thalassemia, where the first FDA-approved CRISPR therapy, Casgevy, marks a significant milestone. The success in these areas underscores the transformative potential of CRISPR technology. However, each field presents unique challenges. While blood disorders benefit from ex vivo editing of HSPCs, facilitating clinical translation, neuromuscular disorders like DMD face obstacles in in vivo delivery and long-term safety. Ongoing research aims to overcome these hurdles, with the development of more efficient delivery methods, high-fidelity Cas9 variants, and non-viral delivery systems. The progress in CRISPR-based therapies holds great promise for patients with debilitating genetic conditions, potentially offering cures where none existed before.

Editorial: First Regulatory Approvals for CRISPRCas9 Therapeutic Gene Editing for Sickle Cell Disease and Transfusion-Dependent b-Thalassemia

The landscape of gene therapy has reached a pivotal moment with the first regulatory approvals of CRISPR-Cas9 therapeutic gene editing for sickle cell disease (SCD) and transfusion-dependent β-thalassemia. On November 16, 2023, the UK Medicines and Healthcare Products Regulatory Agency (MHRA) became the first to approve Casgevy (exagamglogene autotemcel), a CRISPR-Cas9-based therapy, for patients aged 12 years and older suffering from these debilitating blood disorders. This groundbreaking approval was soon followed by the U.S. Food and Drug Administration (FDA) on December 8, 2023, which approved both Casgevy and Lyfgenia (lovotibeglogene autotemcel) for patients with SCD. The European Medicines Agency (EMA) also granted approval to Casgevy on December 15, 2023, marking a significant global milestone in gene therapy.

Sickle cell disease affects approximately 100,000 individuals in the United States, with a prevalence of 1 in 365 among African Americans. The disease is characterized by episodes of hemolytic anemia, severe pain, and organ damage due to misshapen red blood cells that obstruct blood flow. β-Thalassemia, on the other hand, has a global carrier prevalence of 80–90 million people, affecting up to 1.5% of the population. Patients with transfusion-dependent β-thalassemia suffer from severe anemia requiring regular blood transfusions, which can lead to iron overload and other complications.

Casgevy utilizes CRISPR-Cas9 technology to edit the BCL11A gene in patients' hematopoietic stem and progenitor cells (HSPCs). By disrupting the erythroid-specific enhancer region of BCL11A, the therapy reactivates the production of fetal hemoglobin (HbF), which can compensate for the defective adult hemoglobin in these patients. In clinical trials involving 31 patients with SCD, 93.5% (29 patients) achieved the primary efficacy outcome of freedom from severe vaso-occlusive crises for at least 12 consecutive months during a 24-month follow-up period. Importantly, all 31 patients achieved successful cell engraftment without any cases of graft rejection or failure.

The safety profile of Casgevy was generally favorable. Commonly reported side effects included reduced platelet counts, decreased white blood cell levels, nausea, mouth ulcers, musculoskeletal pain, abdominal pain, and febrile neutropenia. These side effects are consistent with those expected from myeloablative conditioning and stem cell transplantation procedures.

Lyfgenia, approved concurrently by the FDA, is a cell-based gene therapy that employs a lentiviral vector to introduce a modified β-globin gene (HbAT87Q) into patients' HSPCs. This modified hemoglobin functions similarly to normal adult hemoglobin, reducing red blood cell sickling and improving vascular blood flow. In a study of 32 patients with SCD aged between 12 and 50 years, 88% (28 patients) achieved complete resolution of vaso-occlusive events between 6 and 18 months post-infusion. The most common side effects mirrored those of Casgevy, including stomatitis, reduced platelet and white blood cell counts, and febrile neutropenia.

However, there have been reports of hematologic malignancies in patients treated with Lyfgenia, leading to a black box warning regarding this risk. This underscores the importance of long-term monitoring for patients receiving gene therapies, as the integration of viral vectors into the genome may have oncogenic potential.

The approval of these therapies represents a culmination of decades of research in gene editing. In 2020, Emmanuelle Charpentier and Jennifer Doudna were awarded the Nobel Prize in Chemistry for their discovery of the CRISPR-Cas9 system, a tool that has revolutionized the field of genetic engineering. CRISPR-Cas9 allows for precise, targeted modifications of DNA, offering the potential to correct genetic defects at their source.

Despite the excitement surrounding these advancements, challenges remain. Ethical concerns persist regarding gene editing, particularly in germline cells, which could have heritable effects. In somatic cells, safety concerns focus on potential off-target effects and the long-term consequences of genetic modifications. The reported cases of hematologic malignancy highlight the need for vigilant post-therapy surveillance.

The high cost and complexity of these treatments also present barriers to widespread accessibility. Myeloablative conditioning and stem cell transplantation require significant medical resources, limiting availability to patients in resource-rich settings. Additionally, the personalization of these therapies, which involve editing each patient's own cells, adds to the logistical challenges.

In response to these challenges, attention is turning towards RNA-editing therapies as a potentially safer alternative. Unlike DNA editing, RNA editing does not introduce permanent changes to the genome, reducing the risk of unintended long-term effects. In February 2024, RNA-editing treatments for alpha-1 antitrypsin deficiency (AATD) and Stargardt disease received approval for clinical trials. These therapies utilize mechanisms such as adenosine deaminase acting on RNA (ADAR) to make single-base edits or exon editing to correct multiple mutations at the RNA level.

RNA-editing approaches may also have applications in oncology, where they could be used to produce proteins that inhibit tumor growth or to replace RNA sequences associated with cancer progression. The transient nature of RNA edits offers a balance between therapeutic efficacy and safety, potentially circumventing some of the risks associated with DNA editing.

The recent regulatory approvals signify a pivotal shift from experimental research to clinical application of gene editing therapies. For patients with sickle cell disease and β-thalassemia, these therapies offer hope for a cure rather than lifelong management of symptoms. The long-term efficacy and safety of Casgevy and Lyfgenia will continue to be evaluated through ongoing studies and patient monitoring.

As the field progresses, it will be essential to address the ethical, safety, and accessibility issues inherent in gene editing therapies. Collaborative efforts between researchers, clinicians, regulatory bodies, and bioethicists will be crucial in guiding the responsible development and implementation of these technologies. The potential to alleviate suffering caused by genetic diseases is immense, but it must be pursued with caution and a commitment to patient welfare.

In conclusion, the first approvals of CRISPR-Cas9 gene editing therapies mark a significant milestone in medicine, heralding a new era of precision therapies for genetic diseases. While challenges remain, the progress achieved thus far lays a strong foundation for future innovations. The integration of gene and RNA-editing technologies may expand therapeutic options, offering safer and more accessible treatments for a range of genetic conditions.