A five-year-old’s brains are behind groundbreaking research to cure debilitating genetic diseases in children. Denise Cullen explores the frontier of personalised medicine. This article was originally published in the Cosmos Print Magazine, September 2024.
Open the incubator door in a locked laboratory on the fourth floor of the University of Queensland’s Australian Institute of Bioengineering and Nanotechnology (AIBN) and you’ll find hundreds of tiny ‘brains’ belonging to a five-year-old girl called Tallulah Moon.
Nestled in rows of petri dishes, and ensconced in a solution containing salts, sugars, vitamins and growth factors, each one is barely the size of a grain of raw couscous.
Although they’re tiny, they sleep and dream, grow and change, develop new synaptic connections and prune old ones, and display the same alpha, beta and gamma waves as would any living human if you were to hook them up to an electroencephalogram (EEG).
Though researchers refer to them colloquially as ‘mini brains’, Senior Research Fellow with the AIBN, Hannah Leeson, admits this is a bit of a misnomer.
“These are cortical organoids, they don’t have a mid or hindbrain, and there’s no cerebellum,” she says.
More correctly, they are brain organoids – miniature, simplified parts of the cortical brain which have been grown in vitro from stem cells.
They can mimic the structure and function of a real brain, but on a much smaller scale.
For five years now, Leeson has been trying to unravel the mysteries they contain, by observing their electrical activity.
They were created using skin cells from Tallulah, then aged three, after she was diagnosed with a rare genetic disease called Hereditary Spastic Paraplegia Type 56 (SPG56).
The degenerative condition causes progressive weakness and spasticity (stiffness) of the lower limbs and arose due to a mutation in just one of Tallulah’s 30,000 genes – the CYP2U1 gene.
SPG56 is typically inherited in an autosomal recessive manner, meaning that Tallulah inherited two copies of the mutated gene, one from each of her parents, to manifest the disease. Her older brother is perfectly healthy.
The nightmare for Chris and Golden Whitrod, Tallulah’s parents, started in 2020, when she was 14 months old.
Within months, the bright-eyed toddler had reverted to crawling and, soon after, Tallulah could no longer talk or even hold up her head.
“When she started to choke on food and drink, we hurled ourselves into all-consuming state of terror and the hurried referrals for neurologists, blood tests, MRIs and nerve conduction studies could not come quickly enough,” says Golden, in a statement on the family’s Our Moon’s Mission website.
After months of medical investigations, Whole Genome Sequencing (WGS) mapped every gene in Tallulah’s body and identified the disease-causing mutation.
The devastating news was that there was no treatment and no cure.
In August 2021 the Moon family established a charitable foundation called Genetic Cures for Kids, with its first mission being to find a cure for SPG56.
Chris and Golden met with researchers at AIBN in 2021 to plot the route forward.
Animal models were unlikely to yield answers, says AIBN Senior Group Leader Ernst Wolvetang.
“The mouse model (of this disease) was happy as Larry, just walking round, doing his thing, even though it was missing the same gene as these kids who are ending up in wheelchairs and getting desperately sick,” he explains.
“For diseases affecting the brain, mouse models just don’t recapitulate the disease effectively.”
Researchers’ attention then turned to investigating how the mutation caused its effects and how gene therapy might ameliorate them.
The gene therapy approach
Leeson began by taking a sample of skin cells from Tallulah via a simple skin punch.
She then used the ground-breaking process called induced pluripotent stem cell (iPSC) reprogramming to revert these cells to a pluripotent state. In this state, they held the potential to develop into any cell type in the body, including the brain.
She took the same samples for Tallulah’s parents, who were used as controls. Leeson estimates she has created around 800 brain organoids each from Tallulah, Chris and Golden. At the time of writing, Leeson was in the process of creating hundreds more.
Then, by carefully observing and comparing the brain organoids through multiomic analyses (which combine data from genomics, transcriptomics, epigenetics, and proteomics) and by measuring the neuronal activity via multielectrode array, she sought to identify something that stood out about the disease.
“I was looking for differences that were specific to the patient, because if you see a difference somewhere, then you’ve got something you can measure when it comes to looking for a treatment,” Leeson explains.
It emerged that Tallulah’s mutated CYP2U1 gene metabolised glucose differently. Leeson also observed changes in fatty acid levels, which are crucial for healthy brain function.
“Normally, you get your sugar molecules and then that feeds into the citric acid cycle and that makes your adenosine triphosphate (ATP),” she explains. ATP is a crucial molecule in cellular metabolism which acts as the primary energy provider. “These cells don’t seem to follow that same mechanism.”
It took two years of investigation to slot these pieces of the jigsaw puzzle into place – but many unknowns remain.
“We haven’t figured out exactly how CYP2U1 controls these processes, or why this causes spastic paraplegia. But now we have something to work with – we have things that are different in patients, compared to controls.”
Mohita Singh (left) and Hannah Leeson (right). Credit: Supplied.
How common are rare diseases?
SPG56 is just one of around 7,000 rare diseases – life-threatening or chronically debilitating conditions that affect a small percentage of the population.
In Australia, a disease is considered rare if it affects less than 5 in 10,000 people, according to the federal Department of Health and Aged Care.
Besides SPG56, there are other forms of hereditary spastic paraplegia. Combined, they’re still so rare that there are only 0.1–9.6 instances in every 100,000 people around the world and the majority are children.
As with other rare diseases, correct diagnosis can take time because these conditions are complex, doctors don’t see them frequently enough, and they require genome sequencing before a mutation can be identified.
Meanwhile, there’s minimal appetite among big pharmaceutical companies to sink funds into research. Their investment won’t be recouped because of the small number of people who will benefit from the product developed.
And even if Big Pharma was willing, Wolvetang points out practical problems associated with researching rare diseases.
“Clinical trials are not even possible because there’s only five kids in Australia that have this disease,” he says. (Tallulah is, in fact, the only known child in Australia with SPG56.)
“A clinical trial with controls and the double-blind placebo crossover design? You can forget it.”
Collectively, however, it’s estimated that 8% of Australians live with a rare disease, making them surprisingly prevalent.
Given that four in five rare diseases are genetic, it’s anticipated that rapidly developing gene therapies will provide hope not just to people with SPG56 but many other rare diseases.
Gene therapy highlights the modern march towards personalised medicine, which focuses on individual differences in genetics, environment and lifestyle to tailor diagnostics, treatment and preventive strategies.
As such, it inverts the traditional ‘one size fits all’ approach to medical research and treatment, perhaps best exemplified by the race to mass vaccinate the planet against COVID-19.
Developing a Trojan horse
With the differences in metabolism and electrical activity between Tallulah’s brain organoids and those of her parents identified, the next step is delivering the functioning gene to replace the faulty one.
“The problem is that it’s very difficult to test this in humans, because you only get one shot at it,” says Wolvetang.
The functioning gene is delivered via a viral vector – a common adeno-associated virus (AAV) – which has been gutted and replaced with desirable genes.
“It’s like a Trojan horse,” says Wolvetang. This is because the AAV is used to ‘sneak’ the correct copy of the gene into the cells.
The vector is placed in the solution surrounding the brain organoid – the laboratory equivalent to delivering drugs via cerebrospinal fluid.
In 2023 Leeson and a student assistant, Mohita Singh, conducted a series of assays, to ensure that there were no toxic impacts upon the brain organoids.
They’ll soon find out how effective introducing the functioning gene will be.
“We know it’s not going to be toxic, but will replacing this gene actually fix the problem?” Leeson asks.
“Are we delivering enough (of the gene therapy)? Are we getting it into enough cells?”
Another outstanding question is whether the deleterious effects of SPG56 can be rewound, or whether the gene therapy needs to be introduced before a critical period of development.
“Tallulah’s five, and I don’t have five-year-old brain organoids, so I’m not really going to be able to answer questions like that,” says Leeson.
“There is a factor of the unknown, when we eventually make it to the stage of putting (the gene therapy) into Tallulah, which we anticipate will be next year.”
Intensive physiotherapy is credited with preserving some of Tallulah’s muscle function against the ravages of SPG56, but even still, she currently uses a wheelchair, and it’s unknown whether gene therapy will change her condition significantly.
Leeson hopes it will, and she speaks about Tallulah with obvious affection.
“She zooms around in her wheelchair and thinks it’s funny to sneak up behind people and bang you in the ankles, like playing tag,” Leeson says.
Entering the unknown
With hopes riding high, there are no guarantees of success.
Even though gene therapies have already been used to treat a range of different diseases, Tallulah’s treatment represents the first to be tested in brain organoids – and that creates uncertainty.
“We don’t know if a brain organoid is going to respond the same way a patient does,” says Leeson.
Weighed against these considerations is the tantalising possibility of seeing the therapy change lives.
“There’s so much we don’t know,” says Leeson, “but this work could change people’s lives – and that’s both exciting and important.”