Scientists edit heart muscle gene in stem cells, may be able to predict risk

In our human genome, there are many elusive g...

Posted: Jun 18, 2018 1:39 PM
Updated: Jun 18, 2018 1:39 PM

In our human genome, there are many elusive genetic variants related to medical conditions, but the impact of these variants to actually cause a disease has not been conclusively determined -- or ruled out.

In other words, the impact certain variants could have on your health remains a guessing game. But a new study involving the gene-editing tool CRISPR could change that.

Scientists use CRISPR to determine if a heart condition-related genetic variant in a patient poses real risk

"This is one of the very first proof of principles to show that concept," researcher says

The study, published in the journal Circulation on Monday, demonstrates for the first time how pairing CRISPR with induced pluripotent stem cell technology could be used to determine the risk of a genetic variant for cardiovascular disease.

Induced pluripotent stem cells are adult cells, such as skin or blood cells, that have been genetically reprogrammed to be like embryonic stem cells. In general, stem cells have the potential to develop into many different cell types in the body.

"Right now the problem is that a lot of people are getting their DNA sequenced and to their surprise it comes back as variants, after variants, after variants -- essentially a whole bunch of variants of uncertain significance -- and they don't know what that means," said Dr. Joseph Wu, professor and director of the Stanford Cardiovascular Institute at the Stanford School of Medicine, who was lead author of the study.

"Patients often ask us what do these variants of uncertain significance mean. But in reality, we don't know most of the time ourselves. So we end up having to follow the patients for the next five, 10, 20, or 30 years to see if the patient manifests the disease or not," Wu said.

"Here, we now have a way to shorten that time because we can generate patients' induced pluripotent stem cells from blood."

How do those stem cells then help predict if a variant is harmful or not? They can be differentiated into heart cells.

If the heart cells look abnormal, that probably means the variant of uncertain significance is pathogenic, meaning it's capable of causing disease.

If the heart cells look normal, that probably means the variant of uncertain significance is actually benign.

"This is one of the very first proof of principles to show that concept," Wu said.

'An important step towards precision medicine'

For the new study, researchers sequenced the DNA of 54 healthy people with no clinical history of cardiac disease and took a close look at the genes associated with hypertrophic cardiomyopathy, a heart condition characterized by thickening of the heart muscle.

Hypertrophic cardiomyopathy is thought to be the most common inherited or genetic heart disease. The condition is a common cause of sudden cardiac arrest in young people, including young athletes.

The researchers found 592 genetic variants across the 54 people. While 78% of the variants were categorized as benign, there were 17 people who each carried a variant categorized as "likely pathogenic." For four of those people, their variant was hypertrophic cardiomyopathy-related.

One study participant, carrying a hypertrophic cardiomyopathic-related variant in the gene called MYL3, also had several family members carrying the gene. The researchers noticed that these family members were asymptomatic in nature, meaning they had no irregular heartbeat and no enlarged heart muscle tissue.

So the researchers then took that knowledge and used CRISPR to turn the patient's stem cells with this MYL3 genetic variant from being heterozygous, meaning they have one normal and one recessive form of the variant, to being homozygous, so that they have two recessive forms of the variant.

Specifically, the researchers took the one study participant's blood cells, turned them into induced pluripotent stem cells, and then used CRISPR to edit those cells in a petri dish. The researchers then differentiated the edited stem cells so they would become heart muscle cells, and performed a comprehensive analysis to evaluate the variant, determining exactly how harmful the variant was or whether it was benign.

In this case, the study participant's variant was predicted to be benign.

A risk with using CRISPR is that it could introduce some unintended changes, but no off-target mutations were detected in the gene-edited cells, the researchers reported in their study.

"Much work remains to further develop stepping stones between editing cells in a dish and genome editing therapeutics that can treat patients, but studies such as this one help identify variants that are promising targets for therapeutic editing," said David Liu, core institute member of the Broad Institute and professor of chemistry and chemical biology at Harvard University, who was not involved in the study.

This gene-editing approach was found to be feasible in this one patient, but more research is needed to determine whether similar results would emerge among more patients.

"While it's very elegant, the major limitation of this work is that it took years of expensive work by a team of very talented scientists to do this for just one patient," said Dr. Kiran Musunuru, an associate professor of cardiovascular medicine at the University of Pennsylvania's Perelman School of Medicine, who was not involved in the new study but has conducted separate research involving CRISPR.

"It's an important step towards precision medicine, but going forward we will need to scale this up and be able to do this for dozens, hundreds, or even thousands of patients at a time, in a matter of weeks and much more cheaply," he said.

Time and cost are also limitations of this approach, Wu said.

"Cost-wise, it takes us probably about $10,000 and time-wise about six months," he said. Those six months would involve making the induced pluripotent stem cells, using CRISPR to edit the cells and then analyzing the differentiated heart cells.

Wu added, "but keep in mind that six months is actually still much better than the current alternative that we have, which is to tell patients that we don't know what the variant means."

The alternative would be following a patient with a variant for years, with the worrisome chance of a disease possibly developing or not developing. In either scenario, the patient as well as family members could have anxiety and stress.

Is this the future of gene editing?

William Lagor, an assistant professor of molecular physiology and biophysics at Baylor College of Medicine in Houston, called the new study "impressive."

"This addresses a major unmet need in patient care by helping determine whether your specific mutation is something to worry about," said Lagor, who was not involved in the study but has conducted separate research on CRISPR.

Then once a mutation has been identified as disease-causing, "this is an ideal platform for testing potential new drugs or gene therapy approaches in a patient-specific manner. This is truly personalized medicine," he said.

"The first therapeutic application of this technology would be to correct rare genetic diseases of the heart itself, where the potential benefit far outweighs the risk to the patient. Some of this technology already exists today, and it is now a matter of demonstrating that this can be done safely and effectively," he said.

A separate study published last year in the journal Nature showed how CRISPR could be used in human embryos to correct a pathogenic gene mutation called MYBPC3, which is associated with inherited heart conditions, including left ventricular noncompaction, familial dilated cardiomyopathy, and familial hypertrophic cardiomyopathy.

Whereas, "in this study CRISPR was used to make edits in cells outside of the body, so that the scientists could better analyze the cells, which is straightforward even if it takes a lot of effort," said Musunuru, who led a separate study in February that involved using CRISPR to lower cholesterol levels in mice, reducing their risk of heart disease.

"However, present-day forms of CRISPR technology do not work well enough in the actual heart muscle in a living being to correct a mutation for a disease like cardiomyopathy," he said. "It's possible that some future generation of gene-editing technology might be able to do the job of treating disease in the heart muscle, years or more likely decades in the future."

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