Gene therapy proposal for cardiac arrhythmia, based on models made from patient cells: Tech: Nature World News

Researchers at the Boston Children's Hospital report having created the first human tissue model of hereditary cardiac arrhythmia, mimicking the abnormal heart rhythms of two patients in a cup, and then suppressing arrhythmia by gene therapy in a murine model.

Their work, published in two articles in the print edition of the newspaper of July 30 circulation, opens the possibility of developing single-dose gene therapy treatments for hereditary arrhythmias, and perhaps more common arrhythmias such as atrial fibrillation.

"Our hope is to give gene therapy in a single dose that would work indefinitely," said Vbadilios Bezzerides, MD, Ph.D., a cardiologist participating in the Inherited Cardiac Arrhythmias program, of the Boston Children's Hospital, which participated to both studies. "Our work provides proof of concept of a translatable gene therapy strategy for treating hereditary cardiac arrhythmia."

Both studies focused on catecholaminergic polymorphic ventricular tachycardia (TVPV), a leading cause of sudden death in children and young adults. The arrhythmia is usually triggered by exercise or emotional stress and it appears for the first time at the average age of 12, often in the form of a sudden loss of consciousness.

Current treatment includes drugs such as beta-blockers and flecainide, surgery to disconnect nerves from the left side of the heart, an implanted cardiac defibrillator (which can lead to life-threatening complications during the POS), and the fact that children exercise as much as possible.

"The treatment of the CPVT is currently rather inadequate: 25 to 30% of patients will suffer from recurring arrhythmias that put their lives at risk despite treatment," says Bezzerides.

Construct an arrhythmic tissue

A study, published online by circulation On July 17, tissue engineering models were used to study the functioning of CPVT at the cellular and molecular level. It was led by William T. Pu, MD, of the Boston Children's Hospital and Kevin Kit Parker, Ph.D., of the Boston Children's School and School of Engineering, Harvard Arts and Sciences (SEAS).

Working with the Hereditary Cardiac Arrhythmias Program, led by Dominic Abrams, MD, MBA, the researchers obtained blood samples from two Boston Children's Hospital patients with CPVT caused by distinct mutations of RYR2, the gene linked to most cases of CPVT. RYR2 encodes a channel that allows cells to release calcium – the first step in initiating cardiac contraction.

Scientists then reprogrammed patients' blood cells to become induced pluripotent stem cells (iPS), capable of making virtually any cell type. From these, they made cardiomyocytes (cardiac muscle cells) carrying the mutations of the CPVT and used them to build cardiac muscle tissue models.

"The cells were seeded on an artificial surface, so that they aligned themselves in a direction similar to that of the heart muscle," says Pu, director of basic cardiovascular and translational research at Boston Children & # 39; s . "The cells have very abnormal beats individually, but after badembly in the tissue, they beat together, which better models the disease, which is why tissue-level models are important."

Exercise test in a dish

With the help of a so-called optogenetic system, the team then applied blue light to one end of the tissue to activate the cells. This created a pulse that shifted along the cell sheet to produce a contraction. With the help of this system, they created a "test of exercise in a dish". To simulate exercise, they added isoproterenol (a substance similar to adrenaline, a stress hormone) and applied infrared light to speed up the heartbeat.

These tests revealed the underlying mechanisms of CPVT. When healthy heart tissue was tested for exercise, calcium moved into the tissues in uniform waves. However, in tissue models made from patients with CPCT, calcium waves moved at varying or no velocities in some parts of the tissue, resulting in abnormal circular motion called reentry – much like what happens in real life.

"When we surveyed cells faster, CPVT tissue experienced re-entrant arrhythmias, whereas normal tissue could treat it very well," Pu said.

To understand how stress makes CPVT patients vulnerable to life-threatening arrhythmias, Pu, Parker and colleagues identified adrenaline-activated signaling molecules, then drugs, and genome editing. CRISPR / Cas9 to selectively inhibit or modify them.

With this strategy, they discovered that in healthy heart tissue, an enzyme called CaM kinase (CaMKII) chemically modified RYR2, causing more calcium to be released from the heart muscle cells. In CPVT cells, this modification combines with the inherited RYR2 mutation to cause excess calcium in the cells, causing arrhythmias.

"Nature designed CaMKII as part of the response to combat or flying," says Pu. "When you're excited, you release more calcium so the heart beats faster, but when RYR2 is mutated, the channel leaks and the cell releases too much calcium, causing arrhythmia."

When the researchers blocked the CaMKII modification, they eliminated arrhythmias in the tissue model. They had the same effect when they blocked CaMKII itself with the AIP peptide, a potent and selective inhibitor of CaMKII.

"The coupling of iPS technology and organs on chips offers new possibilities for studies in precision medicine and for the benefit of patients," notes Parker. "Our vision is to use these technologies to screen patients with rare diseases for enrollment in clinical trials by replicating the patient's disease in vitro, we can test candidate treatments for the patient's disease and measure safety. and the effectiveness, so that the right patients are tested with the right medication. "

Inhibit CaMKII with gene therapy

Since the CaMKII enzyme acts on many tissues located next to the heart – and the brain needs it for memory formation – the team wanted to be able to specifically inhibit CaMKII in the heart. In a separate study, published online by circulation On June 3, a team led by Bezzerides and Pu tested a gene therapy approach in a murine TVPC model.

They developed a special virus that, injected with CPVT to mice, selectively returned to the heart and delivered a PIA. The tests showed that AIP was expressed in about 50% of cardiac cells, enough to suppress arrhythmias, but not significantly expressed in noncardiac tissues, including the brain.

Researchers now plan to refine their gene therapy strategy and test it on a large animal model, then on patients with CPTV, possibly in collaboration with other medical centers.

A general approach to heart disease?

Bezzerides and Pu believe that the treatment could be effective in patients with a TVPC caused by various mutations of the RYR2 gene (more than 160 mutations have been reported). And they believe that their overall strategy of inhibiting CaMKII in the heart could help treat more common causes of heart disease.

"CaMKII is not necessary for normal heart function, but it's active in many forms of heart disease," Pu says. "In murine models of many forms of heart disease, such as ischemic cardiomyopathy, atrial fibrillation or hypertrophic cardiomyopathy, the chronic activation of CaMKII is detrimental.It is possible that our approach to inhibition CaMKII through gene therapy can improve the outcomes of these other types of heart disease. "