Givinostat – How does it work and what do we know so far for Duchenne?

By Prof Annemieke Aartsma-Rus. This version was generated on July 8, 2024.

Givinostat has been approved for the treatment of Duchenne muscular dystrophy patients aged 6 years of age and older by the FDA and evaluation by the EMA is ongoing. In this document, I try to explain how givinostat works.

 

 

Illustration on Givinostat: How does it work?

 

 

dystrophin and Duchenne

Duchenne is caused by a lack of dystrophin. Muscles consist of muscle fiber bundles, which contain a protein skeleton that can contract. Each muscle fiber is surrounded by connective tissue that connects to other muscle fibers, so contraction is coordinated. Muscle fiber contraction causes stress for the muscle fibers. Normally, the dystrophin protein stabilizes contracting muscle fibers by forming a link between the protein skeleton and the connective tissue surrounding muscle fibers. Without functional dystrophin, muscle fibers will be damaged during contraction.

 

Normal muscle repair

Muscle damage will normally be repaired in a well-orchestrated and synchronized series of processes:

1. The immune system will clear the damage and debris; during this process the immune system will inhibit the repair system (you wait with redecorating your house until after the demolition team is done)
2. Once the damage is cleared, fibro-adipogenic stem (FAP) cells are activated to repair the extracellular matrix – the connective tissue shell that surrounds each muscle fiber
3. Then, the muscle stem cells are activated to proliferate and differentiate into mature muscle either by fusing with the remaining muscle fiber or by forming a new muscle fiber within the shell of connective tissue

 

Failed muscle repair in Duchenne

In Duchenne, muscle damage is chronic and therefore there is a desynchronization of the repair processes that leads to failure of proper repair:

1. The immune system is chronically activated, which will inhibit repair in the muscle even at locations where the debris is already cleared
2. The FAP cells are improperly activated and produce too much connective tissue (fibrosis) differentiate into fibroblasts and fat cells that produce fat tissue (adiposis)
3. The muscle stem cells are stuck in proliferation mode and due to the signals from the inflammatory cells and the FAPs they transdifferentiate into FAPs and also produce connective tissue and fat tissue.

 

Increased levels of HDACs in Duchenne

These pathological processes are worsened by increased levels of histone deacetylases (HDAC). dystrophin is part of a protein complex called DGC (dystrophin associated glycoprotein complex), that in addition to providing mechanical stability to muscle fibers, also recruits the nitric oxide synthase (NOS) enzyme to the membrane. This enzyme produces nitric oxide (NO). Without dystrophin, there is less NO formed, which amongst others leads to an increased level and activity of HDAC enzymes.

 

Functions of HDACs

HDACs are enzymes that remove acetyl groups from proteins. Their counterparts (histone acetylases, HATs) add acetyl groups to proteins. The acetylation and deacetylation of proteins, influences important processes in the muscle cells.

The most well-known proteins that are influenced by the addition and removal of acetyl groups are histones, the proteins around which DNA is wrapped. DNA contains our genetic material. Each cell contains 2 meters of DNA, which is a thin long molecule. To prevent the DNA from tangling and breaking, it is wrapped closely around histones. However, DNA also needs to be accessed to allow it to be read for gene expression and protein production. This process is influenced by acetylation – when the histones are acetylated the DNA unwraps and becomes accessible for the enzymes that transcribe genes. This happens only at the place of said gene – the rest of the DNA stays tightly wrapped.

 

The consequences of too much HDAC activity

Too much HDAC enzyme activity in Duchenne, means that too many acetyl groups are removed from the histones and that the DNA is too tightly wrapped, which prevents expression of genes and failure to produce specific proteins.
Specifically for muscle damage repair this means:

1. Immune system chronically activated: In immune cells, too much HDAC activity inhibits the production of anti-inflammatory proteins, which normally prevent the immune response to become chronic and which dampen the production of factors that are damaging to the tissue. So with too much HDAC activity, the immune response gets ‘out of control’.
2. Altered FAP activity: The FAP cells get stuck in connective tissue production mode and become fibroblasts and fat cells, instead of supporting the muscle stem cells to differentiate and repair muscle
3. Muscle stem cells can’t differentiate: The muscle stem cells cannot differentiate into new fibers without the help of the FAPs. Furthermore, the FAPs produce factors that will cause the muscle stem cells to transdifferentiate into FAPs, and they cannot repair muscle any more.

In addition to histones, other proteins are also regulated by the addition and removal of acetyl groups, which can further exacerbate muscle damage and pathology in Duchenne. For example, the key regulator of fibrosis, TGF-beta works by triggering the addition of a phosphate group to a protein complex (SMAD), which then leads to the formation of fibrosis. Normally, this addition is inhibited by acetylation of SMADs. However, with too much HDAC activity, these acetyl groups are actively removed, reducing the threshold for adding the phosphate groups and thus further increasing the levels of fibrosis formation.

 

HDAC inhibition in Duchenne

Given that multiple processes in muscle damage and repair are exacerbated by too much HDAC activity, it makes sense to anticipate that inhibiting HDAC activity will improve muscle repair.

HDAC activity inhibition is expected to allow:

1. The immune cells to go from the ‘pro-inflammatory’ mode, into a modulatory mode to dampen the immune response and to reduce the inhibition of the muscle repair system
2. The FAP cells to have a supportive role for muscle repair and stop producing fat and connective tissues
3. The muscle stem cells start to differentiate into muscle fibers rather than being stuck in proliferation mode

 

Summary results so far for Givinostat in Duchenne

Givinostat and other HDAC inhibitors have been tested in the mdx mouse model that misses mouse dystrophin. HDAC inhibition resulted in improved muscle strength and reduced inflammation, fibrosis and improved the amount of healthy muscle. Furthermore, it reduced the muscle damage induced after exercise.

Givinostat is the only HDAC inhibitor that has been tested in clinical trials in Duchenne patients. It was first tested in a small group of patients where muscle biopsies were taken before and after a year of treatment. This revealed a reduction in fibrosis and necrosis after treatment for all patients although the level of reduction varied.

Then, Givinostat was tested in a placebo-controlled trial, where both groups continued to receive glucocorticosteroids. After 18 months the treated group showed a significant reduction in the decline in the speed at which patients climbed 4 stairs (primary endpoint). Also for the secondary endpoints (e.g. north star ambulatory assessment, 6 minute walk test, muscle strength and time to rise), the treated patients did better than placebo. Finally, the givinostat treated patients had less fat infiltration in the leg muscles than placebo treated patients as measured with MRI analysis.

he most commonly reported side effect was diarrhea. It is known that givinostat treatment can cause a reduction in blood platelets. If this was too severe, it was managed by dose adjustments or temporary interruption of dosing.

 

Disclaimer

The reason for writing this document was to explain the working mechanism of givinostat to the patient community. I received no payment for the writing of this document. The content is my own, but it was sent to Italfarmaco for a fact check.

My employer (LUMC) has received honoraria from Italfarmaco for my speaker activities at a symposium and remuneration for ad hoc consulting. Furthermore, LUMC has done research with givinostat funded by Italfarmaco in a contract research setting. I have not personally received money from Italfarmaco for any of these activities.