Krabbe disease or globoid cell leukodystrophy (LDG) is a genetic leukodystrophy caused by mutations in the gene encoding galactocerebrosidase (GALC). Both healthy parents of an affected person carry one copy of a pathological mutation in the GALC gene.

When a person inherits two copies of a mutated gene, they exhibit very low GALC activity, which results in a significant build-up of galactose-containing lipids, which are mainly found in the white matter or myelin of the central and peripheral nervous system. (SNC and SNP). This pathology, in addition to the abnormality in the lipid composition of myelin, also has an inflammatory component.

Although Krabbe disease primarily affects infants, the diagnosis is also made in older patients. In some patients, the diagnosis may not be made until late, when certain symptoms lead to genetic testing. However, this late diagnosis may limit the success of any treatment aimed at preventing or repairing damage to the nervous system.

In order to allow faster diagnosis, some states in the United States have introduced newborn screening for Krabbe disease. Some subjects identified in this screening because of reduced GALC activity have disease-causing mutations and certain changes called polymorphisms (normal changes in the gene that can lower the level of activity measured without causing disease).

Being able to determine when and if the subject will have Krabbe disease is critical to the success of the program. Clinical evaluation and neurodiagnostic studies are essential to determine when to initiate treatment. Treatment options are currently limited. Hematopoietic stem cell transplantation (HSCT), when performed in infants who are very mildly affected or before symptoms appear, can prolong life. However, these patients experience significant expressive language problems and increasing problems with walking. Although this treatment is now considered the norm, it is evident that more effective treatments are needed.

Several animal models also show low activity of GALC. They are used to test different treatments, to make sure they are both safe and effective, before they are evaluated in clinical trials in patients.

Numerous studies have been carried out on the mouse model called twitcher (twi), since 1984. These studies have focused on:

• bone marrow transplant (GMO),
• gene therapy using different viral vectors to provide a correct copy of the defective GALC gene,
• enzyme replacement therapy aimed at compensating for the deficiency in GALC activity,
• medicines aimed at slowing the synthesis of galactolipids, reducing the immune response or correcting the synthesis of the mutant GALC protein,
• neuronal stem cell therapy to stimulate remyelination
• and combinations of these treatments. Some treatments have resulted in only a small lifespan, others have not been deemed safe for use in patients Some studies have involved injecting different drug carriers directly into the brains of twi mice, sometimes in combination with other treatments. With some drug carriers, the gains have been modest.We tested an AAVrh10-type drug vector containing the GALC gene, called AAVrh10-GALC. The injection of this vector into the brain and into a blood vessel (intravenous, IV) in 2-day-old mice (PND2) has been shown to be promising: it has significantly extended the lifespan of the treated mice (approximately 40 days to 150 days and more). These mice are fertile and show few signs of the disease until very old age.

In order to streamline the treatment procedures, we decided to inject the vector AAVrh10-GALC IV into 10-day old mice. We chose this age because at this point in the development of the mice, myelination of the nerves begins and it is possible to inject a larger volume of the vector compared to younger mice. The single injection prolonged the life of the mice by an average of 25 to 35 days compared to untreated mice, although some lived longer than 150 days.

In addition, this single injection ensured GALC activity in the brain and medulla, as well as very strong activity in the sciatic nerve, a critical tissue not treated by other procedures. These mice remained fertile, exhibited normal mobility, did not suffer from tremors, and their weight gain was normal until a few weeks before they weakened and died. Myelination of the brain, spinal cord and sciatic nerve has been shown to be normal.

Since HSCT is the standard treatment in patients with Krabbe’s disease, we decided to combine GMO with a single injection of the AAVrh10-GALC vector given the next day in mice aged 9-10 days. These mice have normal weight, normal movements, and some currently live more than 200 days.

This approach should be the subject of further studies in order to determine the optimal time between GMO and the injection of the AAV vector and in order to determine the minimum dose of the drug vector necessary to deliver the gene into the target tissues. All of this is being evaluated in the mouse model.

Studies are also underway in the dog model, before being able to validate these procedures to be able to offer a clinical trial to patients with Krabbe disease.