Periodic Reporting for period 1 - circRNA4DMD (Circular RNA Therapeutics for Duchenne Muscular Dystrophy)
Okres sprawozdawczy: 2023-01-01 do 2025-06-30
Duchenne Muscular Dystrophy (DMD) is a fatal genetic disease caused by the absence of dystrophin protein. Dystrophin is required to preserve muscle tissue integrity, it acts as a scaffold for many different proteins, structural support, and contraction-induced shock absorption. DMD affects around 1/5000 boys born. The symptoms include progressive muscle weakness, starting from 2-3 years old. By the age of 15, patients commonly lose ambulation. The current life expectancy is limited to 30 years because weakened cardiac and diaphragm muscles fail to maintain proper heart and lung functioning.
At present, most patients receive corticosteroids as the main treatment. Corticosteroids may improve the symptoms but cannot cure the disease. Multiple attempts have been made at cure development, including the recently approved gene therapy that uses a virus-like to deliver a miniaturized gene version of the dystrophin to at least partially support muscle function. Unfortunately, this treatment has major limitations partially rooted in the delivery method used for the dystrophin gene. Some patients are not eligible for this treatment due to anti-viral antibodies, others only feel marginal benefit since the minigene is not capable of restoring the function to the full extent.
Based on the recent advances in the field of lipid nanoparticles (LNPs), largely initiated by the success of the LNP-based COVID-19 vaccine, we aim to develop a new method for delivering genes to muscles. Although mRNA delivery has already shown promise in treating other diseases in the academic setting, the major obstacle in going from bench to bedside is the short-term mRNA persistence in the cells.
To overcome this, we propose to deliver circular RNA encoding functional dystrophin instead. Circular RNA has shown promise in overcoming the limitations of gene therapy, as it can bypass the constraints of traditional linear RNA and potentially lead to more durable and robust dystrophin expression in muscle cells. By designing lipid nanoparticles that will home to the muscle cells and encapsulate the circular RNA construct within them, we hope to efficiently deliver the therapeutic RNA to the target tissues and achieve sustained restoration of dystrophin function.
At present, most patients receive corticosteroids as the main treatment. Corticosteroids may improve the symptoms but cannot cure the disease. Multiple attempts have been made at cure development, including the recently approved gene therapy that uses a virus-like to deliver a miniaturized gene version of the dystrophin to at least partially support muscle function. Unfortunately, this treatment has major limitations partially rooted in the delivery method used for the dystrophin gene. Some patients are not eligible for this treatment due to anti-viral antibodies, others only feel marginal benefit since the minigene is not capable of restoring the function to the full extent.
Based on the recent advances in the field of lipid nanoparticles (LNPs), largely initiated by the success of the LNP-based COVID-19 vaccine, we aim to develop a new method for delivering genes to muscles. Although mRNA delivery has already shown promise in treating other diseases in the academic setting, the major obstacle in going from bench to bedside is the short-term mRNA persistence in the cells.
To overcome this, we propose to deliver circular RNA encoding functional dystrophin instead. Circular RNA has shown promise in overcoming the limitations of gene therapy, as it can bypass the constraints of traditional linear RNA and potentially lead to more durable and robust dystrophin expression in muscle cells. By designing lipid nanoparticles that will home to the muscle cells and encapsulate the circular RNA construct within them, we hope to efficiently deliver the therapeutic RNA to the target tissues and achieve sustained restoration of dystrophin function.
Throughout the project, we have laid the foundation for the therapeutic development of non-viral gene therapy for muscle diseases.
• We have explored the in-house library of novel ionizable lipids to find the most suitable lipid for muscle tissue.
• We elucidated the potential composition of the lipid nanoparticle that will passively target the muscle tissue.
• Potential moieties for active targeting were selected and tested in cell culture and a suitable Duchenne Muscular Dystrophy (DMD) mouse model.
• Steps were taken to test the hypothesis that circular RNA would enable more robust and persistent therapeutic gene expression compared to linear mRNA.
• Several versions of the dystrophin gene were synthesized in a circular RNA form, aiming to achieve the synthesis of the full coding sequence.
• We have explored the in-house library of novel ionizable lipids to find the most suitable lipid for muscle tissue.
• We elucidated the potential composition of the lipid nanoparticle that will passively target the muscle tissue.
• Potential moieties for active targeting were selected and tested in cell culture and a suitable Duchenne Muscular Dystrophy (DMD) mouse model.
• Steps were taken to test the hypothesis that circular RNA would enable more robust and persistent therapeutic gene expression compared to linear mRNA.
• Several versions of the dystrophin gene were synthesized in a circular RNA form, aiming to achieve the synthesis of the full coding sequence.
The research is taking steps to develop non-viral gene delivery methods beyond the state-of-the-art by employing novel methods of targeting muscle tissue. In particular, a lipid nanoparticle (LNPs) composition was developed and decorated by active targeting moieties. Targeted LNPs have been shown to achieve superior muscle penetration. On the payload side, multiple long circular RNA versions of the dystrophin gene were synthesized and tested successfully, reaching lengths that were longer than the miniaturized gene version of the dystrophin that had not been reported before