Cure Rare Disease’s mission is to develop advanced therapeutics to treat individuals with rare and ultra-rare genetic diseases including Duchenne muscular dystrophy (Duchenne). 

Steps toward a clinical trial

Cure Rare Disease has developed a framework which includes 5 primary stages of development for a therapeutic. Phase 1 focuses on developing a cell line, which is a culture of cells that are derived from a patient harboring the mutation or disease in question. The cells are obtained through a tissue biopsy. The goal here is to understand exactly what genetic mutation is present and how it is impacting the production of the RNA and protein, which in the case of Duchenne is dystrophin.

In phase 2, the therapeutic is prototyped by selecting the appropriate CRISPR system along with the guide RNA and muscle promoter that will allow for full-length (or near full-length) dystrophin to be produced. The guide RNA tells the therapeutic where to target on the patient’s genome while the promoter allows the therapeutic to express in the targeted tissue type (ie: cardiac muscle). The prototyped therapeutic is tested and optimized in the cell line prior to starting rodent efficacy studies. 

Phase 3 shifts testing from in-vitro (in cells) to in-vivo (in life, or, more practically, in rodent models) using rodent models. For CRISPR, the therapeutics are mutation specific and human specific. A common method of conducting these in vivo studies is through mice that are engineered to harbor the specific mutations in question and contain a humanized dystrophin gene (remember: CRISPR therapeutics are generally specific to human mutations). While rodent models offer a helpful perspective, rodent models pose a challenge of using CRISPR for Duchenne since there are many mutations that result in Duchenne and consequently, a model needs to be engineered, generally, for each mutation that is being treated. Animal model generation is a long process that generally takes 2 years or more to complete. Nonetheless, once in-vivo testing is complete and if efficacy is shown, the project proceeds to the preIND stage. This stage is a critical opportunity to get the FDA’s feedback regarding the completed and proposed development plan in an effort to avoid expensive mistakes. 

After efficacy is established and the preIND meeting is complete, phase 4 commences. During phase 4, the human grade therapeutic is manufactured and the pivotal toxicology study is conducted. Manufacturing gene therapies is one of the biggest challenges in the age of gene therapy. In terms of cost and complexity, this is an area where many challenges arise - can the therapeutic be made at scale? The sheer cost of this step is a challenge as well - typically clinical grade manufacturing costs $2M per drug. For this reason, many programs fail at this stage or are unable to proceed due to cost constraints. Advancing past manufacturing and toxicology studies leads to the final preclinical step: IND preparation and submission to the FDA. 

Once the IND is submitted, the FDA has up to 30 days to provide feedback - whether that is approval to dose patients or additional questions. Once the FDA is satisfied, the IND is approved. The hospital that hosts the clinical trial also has internal approval processes via the Institutional Review Board or IRB. The IRB is responsible for oversight within the hospital along with the FDA. 

Phase 5 is the final stage, in which the clinical trial is conducted. For gene therapy treatments, immune suppression is an important step to help minimize the chances that there is a severe immune response to the AAV-based therapeutic. Once the immune suppression is completed, the therapeutic is administered and the patient is monitored for a period of time in the hospital before being released. Once released, the clinician will continue to follow up with the patient over a period of 15 years. The 15 year time period is derived from FDA guidance and is specific to genome editing therapeutics. Other types of therapeutics will entail different periods of follow up. 

Where is this done?

Many institutions and researchers collaborated in order to make these treatments a reality. CRD works with the Yale School of Medicine in order to develop the drug itself. Recently, CRD invited families to Yale University to learn more about the research being done there. Once developed, CRD, Yale and Charles River works together to test the drug during the preclinical stages. Finally, the patients are dosed with the drug at UMass Medical School. Drug development is a team effort and through collaboration we can expedite the development process. 

What can CRISPR do?

Because each person with a genetic disease can have different genetic mutations causing the disease, treatment plans must be focused on the targeted mutation - whether that mutation impacts 10 people or 10,000 people. In DNA, an exon is a portion that codes for a protein. If there is a duplication, CRISPR can be used to cut out the extra exons in a process called deletion. With repression, genes that cause disease can be turned off so that they are no longer expressed and the problematic protein is no longer produced. On the other hand, activation is the process by which protein production can be increased. 

Neutralizing Antibodies

A type of virus called AAV, or adeno-associated virus, is utilized for many gene therapies today. Unfortunately, a percentage of people have neutralizing antibodies (NAbs), which are naturally occurring antibodies against this virus. CRD conducted a study to analyze a subset of the population to better understand the rate at which people have NAbs to commonly used AAV types (serotypes). You can read more about those findings here. Additionally, AAV-delivered therapeutics will result in the formation of more NAbs, which is why this method can currently only be used one time per patient. CRD is working to remove this barrier to enable redosing, where necessary, of life-saving treatments to people impacted by Duchenne and other rare, neuromuscular disorders.