Development of Base Editing for Gene Therapy of ELANE-Mutated Severe Congenital Neutropenia
Abstract
We propose to develop a novel approach to gene therapy for severe congenital neutropenia (SCN), a life- threatening bone marrow failure syndrome. Patients with this disorder lack sufficient numbers of neutrophils, the form of white blood cells necessary to protect against bacterial infections. Affected children present early in life with recurrent, often very severe infections; without treatment, most die by age 2 years. Current treatment with lifelong drug injections has dramatically improved survival, but at the cost of progression to leukemia, with a 20%-40% risk over 15 years of therapy. Also, approximately 10% of patients do not respond to therapy and are thus at continual risk of fatal infection. Bone marrow transplantation is the only currently curative treatment, but is limited by donor availability and by the risk of severe and sometimes fatal transplant-related complications. Thus there is an unmet need for safe and effective therapy for SCN. Approximately 40% of SCN cases derive from inherited mutations in ELANE, the gene that produces a protein termed neutrophil elastase. All of these mutations change the structure of the elastase protein resulting in an abnormal protein that causes death of the bone marrow cells that would normally develop into mature neutrophils for release into the blood. We propose to develop methods to change the ELANE gene’s genetic code (i.e., edit the DNA) to keep it from making the abnormal protein. The absence of neutrophil elastase is unlikely to interfere with the development or function of neutrophils produced by cells with the edited DNA, as indicated by the absence of any increased risk of infection in individuals with an inherited disorder, Papillon Lefevre syndrome. Neutrophils from patients with this syndrome lack not only elastase, but also three other similar proteins; they have bad gum disease, but no problems with infections elsewhere. Although methods are in development by our group and others to knock out the ELANE gene using the CRISPR/Cas9 system, that method breaks the DNA, resulting in a risk of removing or altering genetic material in ways that could cause other unwanted mutations or cancer. We propose to edit the ELANE gene using the new technology for DNA base editing, which causes pinpoint changes in the genetic code without breaking the DNA. Also, rather than targeting the part of the gene that contains the genetic code for the protein, our approach will alter the code that regulates activity of the gene, causing it to be silenced. We will first test and optimize the editing system in cells that easily accept the introduction of materials such as the base editing components. We will also find the optimal conditions to minimize editing at undesired sites in the cells’ DNA. We will further refine these processes in normal blood stem cells. These cells will then be induced to develop into functional neutrophils in the laboratory, so we can assess the effects of editing on the functions of these cells, particularly those functions necessary for the uptake and killing of bacteria. The results of these experiments will provide the basis for future research to translate its application from the laboratory to human gene therapy for curative therapy of ELANE-mediated SCN. Analysis of neutrophil function after ELANE silencing will also provide basic insight into which functions of neutrophils are dependent or independent of elastase activity.
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Dec 28, 2022
- Source ID
- W81XWH2210440
Entities
People
- Peter Newburger
Organizations
- United States Army
- University of Massachusetts Medical School