Gene Therapy Reverses the Effects of Autism-Related Mutation in Human Brain Organoids

Microscopy images reveal significant differences in size and structure between brain organoids derived from a patient with Pitt-Hopkins syndrome (right) and a control (left). Credit: UC San Diego Health Sciences

The University of California, San Diego (UCSD) study uses laboratory-grown human brain tissue to identify neuronal abnormalities in Pitt-Hopkins syndrome and test gene therapy tools.

In a study published May 2, 2022 in the journal Communications of nature, scientists at the University of California San Diego School of Medicine used organoids from the human brain to discover how a genetic mutation associated with a severe form of autism alters neuronal development. The use of gene therapy tools to restore gene function successfully rescued neuronal structure and function.

Various neurological and neuropsychiatric disorders, such as Autism Spectrum Disorders (ASD) and Schizophrenia, have been linked to mutations in the transcription factor 4.TCF4), an essential gene in brain development. Transcription factors regulate when other genes are turned on or off, so their presence, or lack thereof, can have a domino effect on the developing embryo. However, little is known about what happens to the human brain when TCF4 is mutated.

To explore this question, the researchers focused on Pitt-Hopkins Syndrome, an ASD caused specifically by mutations in TCF4. Children with genetic disease have profound cognitive and motor disabilities and are usually nonverbal.

Pitt-Hopkins syndrome (PTHS) is a rare genetic disorder characterized by developmental delay, epilepsy, distinctive facial features, and possible intermittent hyperventilation followed by apnea. As more is learned about Pitt-Hopkins, the spectrum of developmental disorder widens to include difficulties with autism, anxiety, ADHD, and sensory disorders. It is related to an abnormality within chromosome 18, specifically an inappropriate expression of the TCF4 gene.

Existing mouse models of Pitt-Hopkins syndrome fail to accurately mimic the neural characteristics of patients, so the UCSD team created a human research model for the disorder. Using stem cell technology, they converted patients’ skin cells into stem cells, which later developed into three-dimensional brain organoids or “mini-brains.”

Initial observations of brain organoids revealed a large number of structural and functional differences between the TCF4-Mutated samples and their controls.

“Even without a microscope, you could tell which brain organoid had the mutation,” said study lead author Alysson R. Muotri, PhD, professor at UC San Diego School of Medicine, director of the UC San Diego Stem Cell Program and a member of the Sanford Consortium for Regenerative Medicine.

He TCF4-mutated organoids were substantially smaller than normal organoids, and many of the cells were not really neurons, but neuronal progenitors. These simple cells are destined to multiply and then mature into specialized brain cells, but in mutated organoids, part of that process had gone wrong.

A series of experiments revealed that the TCF4 the mutation caused a downstream deregulation of SOX genes and the Wnt pathway, two important molecular signals that guide embryonic cells to multiply, mature into neurons, and migrate to the correct location in the brain.

Due to this deregulation, neuronal progenitors did not multiply efficiently and therefore fewer cortical neurons were produced. Cells that matured into neurons were less excitable than normal and often remained clustered rather than organized into tuned neural circuits.

This atypical cell architecture disrupted the flow of neuronal activity to the mutated brain organoid, which the authors said would likely contribute to impaired cognitive and motor function below.

“We were surprised to see such important development issues at all these different scales, and it made us wonder what we could do to address them,” said lead author Fabio Papes, PhD, associate professor at the University of Campinas and visiting researcher at UC. San Diego School of Medicine, which jointly oversaw the work with Muotri. Papes has a family member with Pitt-Hopkins Syndrome, which motivated him to study TCF4.

The team tested two different gene therapy strategies to recover the functional gene in brain tissue. Both methods effectively increased TCF4 levels and, in doing so, the phenotypes of Pitt-Hopkins syndrome were corrected on a molecular, cellular, and electrophysiological scale.

“The fact that we can correct this gene and have the whole neural system restored, even on a functional level, is amazing,” Muotri said.

Muotri notes that these genetic interventions took place in a prenatal stage of brain development, while in a clinical setting, children received their diagnosis and treatment a few years later. Therefore, clinical trials must first confirm whether a subsequent intervention is still safe and effective. The team is currently optimizing its newly licensed gene therapy tools in preparation for such a trial, in which spinal injections of the genetic vector would restore TCF4 function to the brain.

“For these children and their loved ones, it would be worthwhile to try any improvement in motor cognitive function and quality of life,” Muotri said.

“What’s really exceptional about this work is that these researchers go beyond the lab and work hard to make those findings translate into the clinic,” said Audrey Davidow, president of the Pitt Hopkins Research Foundation. “This is much more than a stellar academic work; it is a true measure of what good science can achieve to change human lives for the better. “

Reference: “Loss of function of transcription factor 4 is associated with deficits in progenitor proliferation and cortical neuron content” by Fabio Papes, Antonio P. Camargo, Janaina S. de Souza, Vinicius MA Carvalho, Ryan A Szeto, Erin LaMontagne, José R. Teixeira, Simoni H. Avansini, Sandra M. Sánchez-Sánchez, Thiago S. Nakahara, Carolina N. Santo, Wei Wu, Hang Yao, Barbara MP Araújo, Paulo ENF Velho, Gabriel G. Haddad and Alysson R. Muotri, May 2, 2022, Communications of nature.
DOI: 10.1038 / s41467-022-29942-w

Co-authors include: Janaina S. de Souza, Ryan A. Szeto, Erin LaMontagne, Simoni H. Avansini, Sandra M. Sanchez-Sanchez, Wei Wu, Hang Yao, and Gabriel Haddad at UC San Diego; Antonio P. Camargo, Vinicius MA Carvalho, Jose R. Teixeira, Thiago S. Nakahara, Carolina N. Santo, Barbara MP Araujo and Paulo ENF Velho at the University of Campinas.

This work was funded in part by the National Institutes of Health (grant R01 MH123828), the Pitt Hopkins Research Foundation, the Sao Paulo Research Foundation (grants 2020 / 11451-7, 2018 / 03613-7, 2018 / 04240-0 ). ) and the U.S. Department of Energy Joint Genome Institute (DE-AC02-05CH11231).

Disclosures: Alysson R. Muotri is the co-founder of TISMOO, a company dedicated to the genetic analysis and organogenesis of the human brain and has a stake in the share capital.

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