Recent Advances in Gene Therapy

Gene Therapy is the use of genetic material to treat or prevent diseases—often rare, inherited disorders that can be life-threatening or debilitating and have limited treatment options. The goal is to develop a one-time therapy to correct the underlying genetic cause of the disease.

The earliest methods involved introducing genetic material, such as functional genes, into cells to replace, silence or manipulate specific genes. More recently, scientists are also developing gene-editing techniques, including the introduction of molecular tools to remove or correct existing DNA segments at precise locations in cells.

The success of gene therapy relies on the safe and efficient delivery of genetic material into affected cells using suitable vectors. The vector can either be delivered directly to the body to be taken up by individual cells, or it can be delivered to a patient cell sample in the laboratory and then returned to the body. Certain viruses are often used as vectors for gene therapy due to their ability to infect cells. Some types of viruses, such as retroviruses, integrate genetic material, including new genes, into the chromosomes of cells. Other viruses, such as adenoviruses, transfer their DNA into the nucleus but do not integrate with the cell's chromosomes. In addition, viruses can deliver gene editing tools into the nucleus.

To date, 23 gene and cell therapy products have been approved by the U.S. FDA, including game-changing treatments for neuromuscular diseases, inherited blindness, certain blood cancers, and more. Below, we discuss some of the latest innovations in gene therapy technology that are laying the groundwork for significantly expanding the impact of this cutting-edge therapy. (by: Dr. Alison Halliday)

1. Virus-free delivery system

A major challenge in gene therapy is how to safely and efficiently deliver genetic material into cells. While gene therapy using viral vectors has been successful, it does come with risks. Sometimes viruses trigger dangerous immune responses—plus, vectors that integrate into a cell's genome can cause errors that can lead to cancer. Therefore, researchers are developing virus-free delivery systems, such as nanoparticles, to deliver genetic material or gene-editing tools without the use of viruses and target them to specific cells.

In a study published in Cell in January, a team of researchers from Harvard and MIT Broad Institute developed a new drug delivery system that uses engineered DNA-free viruses Like particles (eVLPs) that deliver therapeutic levels of specific gene-editing proteins into mouse models of disease. The research team used the eVLP system to successfully edit the Pcsk9 gene in mouse liver cells, which is associated with high cholesterol levels.

In addition, the research team also used a single eVLP injection to correct point mutations in a mouse model of genetic blindness, resulting in partial restoration of vision. This new platform offers a promising new technique for the delivery of gene editing tools with a lower risk of off-target editing or DNA integration compared to viral vectors.

Other researchers have recently developed peptide-based materials as effective vehicles for delivering gene therapy. In a study published in Biomaterials Science, a new platform developed by Sally-Ann Cryan's research team at the University of Dublin, Ireland, produces custom star-shaped polypeptide nanoparticles capable of efficiently delivering a range of therapeutics into cells.

To demonstrate its potential, the team successfully delivered a gene therapy that regenerated bone. In experiments conducted in the lab, they showed that an implantable scaffold loaded with DNA molecules that promote bone and blood vessel growth accelerated bone tissue regeneration and increased new bone formation compared to scaffolds not loaded with DNA molecules. 6 times.

2. Fine-tune gene expression

Once gene therapy is successfully delivered into target cells, it is difficult to control its expression levels. If the gene or DNA editing tool is expressed at high levels, it may increase the likelihood of off-target effects, and if it is too low, the treatment may not be effective.

In a major advance in the field, scientists from institutions including the Children's Hospital of Philadelphia have developed a "dimmer switch" system, X on, that can control gene therapy like a light. Vector-expressed protein levels. The delivery system is based on an alternative RNA splicing (AS) technology regulated by oral small-molecule drugs, which can effectively function throughout the body tissue. Alternative RNA splicing allows a single gene to encode multiple proteins, depending on how the RNA is spliced.

With the X on system, the "cargo" carried by the gene therapy vector is inactive until an oral small molecule drug is administered, which then drives the splicing of the desired corrective gene into its active form. The team has successfully used this system in mice to regulate levels of erythropoietin (EPO), a glycoprotein hormone that is used to treat anemia associated with chronic kidney disease.

The team also showed that, when expressed at high levels, the X on system can also be used to control the expression of gene products that are toxic to the brain. This innovative system provides an unprecedented opportunity to refine and customize gene therapy for use in humans.

3. Precise targeting

Significant progress has also been made in engineering lipid nanoparticles (LNPs) as gene delivery systems in recent years. However, intravenously delivered nucleic acid-containing LNPs primarily target the liver and spleen, limiting their therapeutic applications to diseases affecting these organs. Currently, scientists are investigating other ways to target LNPs to a range of other specific tissues and organs to expand their potential clinical use and minimize the risk of toxic side effects elsewhere in the body.

A team of scientists from Tufts University and other institutions has engineered LNPs that carry mRNA encoding the Tsc2 gene to target lung tissue. Patients with lymphangioleiomyomatosis (LAM) have a defect in the Tsc2 gene, which causes smooth muscle cells to grow out of control and form cysts in the lungs, causing difficulty breathing. In a mouse model of the disease, LNPs carrying a normal copy of the Tsc2 gene were delivered directly to the lungs, resulting in a significant reduction in cysts. By altering the different components of lipids and other molecules, researchers have also successfully developed LNPs that target the brain, immune system, spleen, liver and even specific cell types.

The precise targeting of specific immune cells by LNPs could help improve a class of gene therapy already approved for certain blood cancers. This gene therapy involves isolating T cells from a patient's blood and introducing a chimeric antigen receptor (CAR) gene that helps the T cells better recognize cancer cells. Genetically modified CAR-T cells are reinjected into patients to boost the body's immune system's ability to fight cancer.

In a recent study published in Science, scientists from the University of Pennsylvania encapsulated a modified mRNA encoding a CAR in a T-cell-targeting LNP, and produced transient CAR-T in vivo by injecting this LNP. cell. This ability to generate CAR-T cells in vivo has the potential to be a therapeutic platform for the treatment of multiple diseases.

4. Safer gene editing

CRISPR-Cas9 is a gene-editing system that consists of two molecules: the Cas9 enzyme that cuts both strands of DNA, and a piece of RNA that guides it to its correct location in the genome. The cell then recognizes the DNA damage and tries to repair it, providing a way to remove, add, or alter segments of the genome. While the technology has great potential for treating human disease, researchers are currently focusing on fine-tuning its application in preclinical studies. Much of this work was done to eliminate "off-target" effects that could accidentally introduce potentially dangerous mutations.

In a study published in Nature in March, scientists from the University of Texas at Austin created a new Cas9 enzyme, SuperFi-Cas9, that remains the same as the original Cas9 enzyme While editing is efficient, it is thousands of times less likely to target the wrong DNA segment, making it potentially safer.

While other labs have previously redesigned Cas9 to reduce off-target interactions, these versions of Cas9 sacrifice editing speed for accuracy. But SuperFi-Cas9 is 4,000 times less likely to cleave off-target sites, yet as fast as the natural enzyme. These results provide a molecular blueprint for designing next-generation high-fidelity Cas9 variants that reduce the risk of off-target effects while maintaining editing efficiency.

5. Future prospects

After decades of hard work, the future of genetic medicine is looking increasingly promising. The success of recent years has led to regulatory approval of several gene therapies that are transforming patients' lives, and many more are in development. Thanks to recent technological advancements, scientists are achieving unprecedented levels of control over nucleic acid delivery and precise manipulation of the human genome, unlocking exciting opportunities to unleash a new wave of transformative genetic medicines.