Folbigg: how scientists decide which mutations in your DNA are deadly

Genetics has been at the centre of the second inquiry into Kathleen Folbigg’s convictions, which is working to determine whether a plausible explanation exists as to the cause of deaths in her children.

Folbigg was convicted for infanticide in 2003, and has so far served 20 years of a three-decade sentence.

This inquiry was enabled by new research suggesting a gene variation, known as CALM2 G114R and possessed only by Folbigg and her daughters, could explain their girls’ sudden deaths.

Throughout the Folbigg inquiries, geneticists have repeatedly referenced the ACMG Guidelines.

These guidelines are central to the debates of whether G114R is deadly.

But what are these guidelines, and why do they matter to this inquiry so much?

Classifying mutants

Every human on Earth, right now, is walking around with variations in their genome – the complete set of DNA that provides instructions of how to make a complex, multicellular organism.

These are variations beyond the benchmark, or most common form of individual genes. Some are inherited, some arise in an individual for the first time.

Some gene variations are harmless, some cause disease (or help prevent it), others might even be beneficial.

To try and determine the likelihood of a variant gene causing disease, a system was developed by the American College of Medical Genetics (ACMG) to evaluate new mutations.

That system seeks to standardise the terminology around five classifications that tell experts the potential risk associated with gene variants. They are: benign, likely benign, a variant of uncertain significance, likely pathogenic and pathogenic.

A benign variant will have little or no impact on the person; a pathogenic variant would cause clinically significant health problems and possibly death.

These guidelines are the product of expert opinion, consensus and input from clinicians.

 For example, pathogenic changes to the CFTR gene are responsible for cystic fibrosis, around 300 disease-causing variations exist.

Since the rise of new technology enabled humans to decode their genome, and deliver quick and accurate identification of gene variants, geneticists have helped people understand the risks associated with their DNA.

Professor John Shine from the Garvan Institute of Medical Research (who was a signatory to the petition to the NSW attorney-general calling for Folbigg’s pardon) is an acclaimed molecular biologist.

He explains that, at its core, the use of ACMG classification allows genetics experts to work with people to educate them about the risks inherent in their biology.

That might, for instance, push prospective parents towards assisted reproduction, in order to avoid having a child with serious genetic disease.

“[Classification] has a major, major role in family planning,” Shine says.

“You see it so often in families which have a genetic disease in the family, who want to make sure that their child doesn’t have it.

“They’ll go through in vitro fertilisation and everything else to avoid it. You need to be able to accurately measure and tell them with absolute confidence that this is the pathogenic mutation and this embryo does or doesn’t have it.”

But the Folbigg Variant – G114R – isn’t meant to change

The second Folbigg inquiry was enabled by new research suggesting a variant to the calmodulin gene possessed only by Folbigg and her daughters could explain their girls’ sudden deaths.

Calmodulin proteins are coded for by three CALM genes. One of the important roles for calmodulin in vertebrates – animals with a backbone – is to regulate ion flows in heart cells that result in the heart beating. In vertebrates, all CALM genes are identical.

It’s the high level of gene conservation in the tiny CALM genes that has some scientists convinced any change to them make survival difficult.

Position 114 on the gene, which codes for an amino acid called glycine (referenced by the ‘G’ in G114R) is even better conserved. In vertebrates, plants and some fungal species like yeast, the CALM genes also produce glycine in this position.

Why proteins matter: the building blocks of who we are

Because of this, scientists argue that not only does nature avoid changes to CALM genes, it insists on preserving glycine at position 114.

Making glycine here, they say, is essential for an organism to functionally survive. As one expert says it “never changes”.

This is why the discovery of the G114R mutation is so important to the Folbigg inquiry. The number of people known to have a mutation at position 114 can be counted on one hand. One of them is Kathleen Folbigg.

Of the others, a four-year-old American child is known to have died due to a variation at her CALM3 gene at position 114. Her five-year-old brother had this mutation as well and was resuscitated after a cardiac arrest.

The other two are the deceased Sarah and Laura Folbigg.

The research conclusions

In this inquiry, experts have argued whether the Folbigg variant is likely pathogenic or a variant of unknown significance (VUS) on the ACMG scale.

The difference largely hinges on whether Kathleen Folbigg has a phenotype consistent with a cardiac problem.

The research team behind the G114R study which resulted in the new inquiry, which includes professors Carola Vinuesa, Peter Schwartz, Michael Toft Overgaard, Mette Nyegaard, Matthew Cook and Todor Arsov – all of whom have given evidence – consider the variant likely pathogenic.

They’ve pointed to a fainting episode experienced by Kathleen in adolescence as an indicator of the phenotype for catecholaminergic polymorphic ventricular tachycardia or CPVT, a known calmodulin condition that manifests because of adrenalin increase during exercise or stress.

Others have used Folbigg’s cardiological assessment, which, bar the one fainting episode, provides little conclusive indication she suffers from an arrythmia phenotype, in their evaluations of the gene.

Clinical geneticist, Professor Edwin Kirk from UNSW, has twice evaluated G114R and suggested while it’s “plausible” and “possible” it caused the girls’ deaths, he can’t yet say that it’s likely to have done so.  He considers G114R to be a variant of unknown significance.

And Dr Calum MacRae, in his evidence on Friday, said the disconnect between available clinical data and high quality functional testing led him to think the Folbigg variant likely presents no issues at all, in other words: benign.

On the other hand, even when considering uncertainty around whether Folbigg has a CPVT phenotype, Professor Arthur Wilde – a world leading cardiologist based in Amsterdam – told the inquiry he felt describing the variant as likely pathogenic is “a fair choice… for this particular variant”.

That’s because he found the quality of the functional testing conducted on the variant to be convincing.

Like him, Hugh Watkins, Professor of Cardiovascular Medicine at Oxford University, in another report to the inquiry, described the mutation as “likely pathogenic.“

So while the on-paper system to classify gene variants provides rigidity, scientists clearly exercise discretion. As Watkins said, there are “reasonable, thoughtful and authoritative experts are all looking at the same data but are drawing different inferences”.

“This is because there is substantial uncertainty, and so judgement, rather than interpretation of indisputable fact, is needed to reach a conclusion.”

In this inquiry that judgement will be made by the presiding officer, former Chief Justice of NSW, Tom Bathurst, KC.

Genetics, however, are not only questions being asked at this inquiry.

From Tuesday, the genetic researchers will be replaced by expert psychologists, neurologists and forensic pathologists on the stand.

That means, while the G114R research has been acknowledged by all experts so far to be of a high quality, it may be that another explanation arises that casts reasonable doubt over Kathleen Folbigg’s murder and manslaughter convictions.

More on the Folbigg inquiries

• Folbigg inquiry head must decide on evidence now, “can’t wait for science to fully develop”
• Researchers affirm science behind G114R study
• Why proteins matter
• Scientists grilled on advocacy and accuracy
• Calmodulin variants “not benign” say Danish experts
• Can the law keep up with science?
• Video explainer: What is Calmodulin