Researchers Create Gene Therapy for Arrhythmia Using a Stem Cell Model

A research team from the Boston Children’s Hospital recently created the first tissue model of an inherited heart arrhythmia and treated it with gene therapy in an animal model. This work not only provides more information regarding the condition but supports gene therapy as a one dose treatment for patients with the disease. These findings were published in two separate papers in the journal Circulation.

“Our hope is to give gene therapy in a single dose that would work indefinitely,” explained Vassilios Bezzerides, MD, PhD, a cardiologist in the Inherited Cardiac Arrhythmias Program at Boston Children’s Hospital who took part in both studies. “Our work provides proof-of-concept for a translatable gene therapy strategy to treat an inherited cardiac arrhythmia.”

Both studies were centered on catecholaminergic polymorphic ventricular tachycardia (CPVT), an arrhythmia that commonly causes sudden death in young adults and children. This inherited disease is often brought about through stress or exercise, with symptoms first manifesting around age 12. Currently, treatment options for CPVT patients are beta-blockers and other drugs, surgical disruption of the heart’s nerves, implanting a defibrillator, and having patients avoid exercise.

“Treatments for CPVT are currently pretty inadequate: 25 to 30 percent of patients will have recurrent life-threatening arrhythmias despite treatment,” said Bezzerides.

Using Stem Cells to Recreate CPVT

To better understand CPVT at a molecular level, researchers recreated the condition in a tissue model. This work was led by William T. Pu, MD of the Boston Children’s Hospital and Kevin Kit Parker, PhD, of Harvard’s School of Engineering, Arts, and Sciences (SEAS) and Boston Children’s. This portion of the CPVT research was published on July 17 in Circulation.

These researchers first obtained blood samples from two patients with CPVT that was caused by mutations in RYR2, the gene most closely linked to CPVT. This gene codes for a calcium channel protein that enables cells to release the ion and cause heart contraction.

Next, the team reprogrammed these blood cells to become induced pluripotent stem cells (iPSCs) which can be used to form all cell types. In this case, these iPSCs were used to create heart muscle cells that carry the RYR2 mutations. In doing so, the researchers created a model of the arrhythmia in artificial heart muscle tissue.

“The cells were seeded on an engineered surface so that they lined up in a specific direction similar to how heart muscle is organized,” said Pu, director of Basic and Translational Cardiovascular Research at Boston Children’s. “The cells have very abnormal beating individually, but after assembly into tissue, they beat together, better modeling the actual disease. That’s why tissue-level models are important.”

The team then applied blue light to the tissue to initiate contraction, creating an impulse that propagated through the tissue and activated the cells. This system was used to simulate an exercise test in the tissue, with the researchers adding isoproterenol (an adrenaline-like hormone) to it and applying infrared light to induce faster heartbeats.

Being that CPVT manifests during exercise, this test helped show the diseases molecular mechanisms. The researchers found that calcium moved through the cardiac tissue at variable speeds, whereas in healthy tissue, it moves in even waves. This abnormal calcium release resulted in the arrhythmic contractions seen in CPVT.

“When we paced the cells faster, the CPVT tissue sustained re-entrant arrhythmias, whereas normal tissue could handle it fine,” said Pu.

The team then identified molecules activated by adrenaline and used CRISPR-Cas9 and drugs to selectively modify them. In this manner, they identified the CaM kinase (CaMKII) that modifies RYR2 and makes healthy cells release more calcium. In the CPVT cells, however, this modification builds on the effect of the preexisting RYR2 mutation and leads to excessive calcium levels. Blocking CaMKII in the tissue model eliminated arrhythmias.

“Nature designed CaMKII as part of the fight-or-flight response,” said Pu. “When you get excited, you release more calcium so the heart can beat faster. But when RYR2 is mutated, the channel is leaky, so the cell releases way too much calcium, which causes arrhythmia.”

A Gene Therapy Approach to Treating Arrhythmia

Though inhibiting CaMKII in the heart is beneficial in treating CPVT, the enzyme is also required by the brain for necessary cognitive function. To inhibit CaMKII in the heart only, the researchers tested a gene therapy technique in a mouse model of the disease. This work was led by Bezzerides and Pu, and was published in Circulation on June 3.

They modified a virus to travel to the heart and deliver AIP, a peptide that selectively inhibits CaMKII. After this genetically altered virus was injected into the mice, Bezzerides, Pu and colleagues noted that roughly half of the heart cells expressed AIP. This quantity is high enough to treat the arrhythmia but not expressed significantly in other tissues like the brain.

Going forward, the team has plans to improve their gene therapy and apply it in larger animal models, then eventually move into clinical trials. The researchers also feel that this general approach of inhibiting CaMKII could be used to treat more common causes of heart disease as well.