Gene editing for CGD

Dr Suk See De Ravin and Dr Kol A. Zarember, from the National Institutes of Health, USA, describe new approaches to fix the faulty genes that cause X-linked CGD.

Since the 1950s, when chronic granulomatous disease (CGD) was first described, there have been tremendous improvements in clinical support for patients with CGD but definitive cures have been elusive. Bone marrow transplant offers the closest thing to a cure for CGD although finding appropriate donors and the risks of transplant-related problems underscore the need for further research.

Gene therapy for CGD offers one way to get round the problem of donor availability and transplant because it uses the patient’s own cells as a part of the cure. Until very recently, the only experimental approaches to gene therapy in patients with CGD have used viruses to carry healthy copies of CGD genes into patient stem cells. These so-called haematopoietic stem cells (HSCs) are harvested from each patient, treated with the engineered virus and returned to the patient, where they give rise to healthy, corrected blood cells that can fight infection. Although careful design of these engineered viruses has resulted in some limited clinical successes and improvements from earlier gene therapy efforts, the viruses tend to integrate more or less randomly into DNA, so there is a risk that the virus may damage other genes or that the healthy gene may be turned on in the wrong place at the wrong time. Over the past few years, several cutting-edge approaches to reduce these problems have been moving closer to experimental clinical trials.

Target-specific approaches

Zinc finger nucleases

One approach to get round random insertion of early generation gene therapy viruses into the patient’s DNA is to target a specific site in a cell’s genetic instructions (the genome) that is known to tolerate insertions without problems. This is called a ‘safe harbour’ site. Recently, we and others (for more information, see De Ravin et al. 2016 Nature Biotechnology 34(4): 424–9) have used zinc finger nuclease, an enzyme that can be engineered to target specific DNA sites, to help deliver a healthy copy of the entire gene that codes for the gp91phox gene (encoded by the CYBB gene) into HSCs from a patient with X-CGD. After transplant into mice, these corrected HSCs gave rise to blood cells that produced gp91phox protein and were able to make superoxide anion, a reactive oxygen radical that helps white blood cells kill pathogens. Even though the healthy gene is in the wrong place in the genome (the safe harbour site), we showed that it was still turned on and off normally by the cell. This has significantly less risk of damaging other genes.

The CRISPR technique

Another new approach is to use the CRISPR system that can be tailor-made to target specific sequences in the DNA for repair. We and others (for more information, see De Ravin et al. 2017 Science Translational Medicine 9(372)1: 10) recently showed that CRISPR could be used to correct the most common mutation in X-CGD. HSCs from patients with this mutation were treated with CRISPR, resulting in correction of the CYBB gene itself without detectable changes elsewhere in the genome. These corrected HSCs were transplanted into mice and differentiated into normal, healthy blood cells, showing that the corrected, healthy gene could be turned on in the right place at the right time.

Looking forward

Although much work remains before these approaches will translate from animal models into human clinical trials, the potential to definitively cure HSC progenitors, and the blood cells that arise from them, is an exciting development.

Authors: Suk See De Ravin, MD, PhD, of the Gene Therapy Development Unit of the Genetic Immunotherapy Section, and Kol A. Zarember, PhD, of the Clinical Pathophysiology Section of the Laboratory of Host Defenses, National Institutes of Allergy and Infectious Diseases, National Institutes of Health, USA.

Posted February 2017