Microscopy images reveal significant differences in size and structure between brain organelles 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-developed human brain tissue to detect nerve abnormalities in Pitt-Hopkins syndrome and to try gene therapy tools.
In a study published May 2, 2022 in the journal Nature Communications, scientists at the University of California, San Diego School of Medicine used human brain organelles to find out how a genetic mutation associated with a severe form of autism disrupts neural development. The use of gene therapy tools to restore gene function has successfully saved nerve structure and function.
Several neurological and neuropsychiatric disorders, including autism spectrum disorders (ASD) and schizophrenia, have been linked to mutations in Transcription Factor 4 (TCF4), a key gene in brain development. Transcription factors regulate when other genes are activated or inactivated, so their presence or absence may have a domino effect on the developing fetus. Still little is known about what happens in the human brain when TCF4 is mutated.
To investigate this question, the researchers focused on Pitt-Hopkins syndrome, an ASD specifically caused by mutations in TCF4. Children with a genetic disorder have profound cognitive and motor disabilities and are usually non-verbal.
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 discovered about Pitt – Hopkins, the developmental spectrum of the disorder expands to include difficulties with autism, anxiety, ADHD, and sensory disturbances. It is associated with an abnormality on chromosome 18, namely an insufficient expression of the TCF4 gene.
Existing mouse models of Pitt-Hopkins syndrome fail to accurately mimic patients’ neural characteristics, so the UCSD team created a human research model of the disorder. Using stem cell technology, they transformed patients’ skin cells into stem cells, which then grew into three-dimensional brain organs or “mini-brains.”
Initial observations of brain organelles revealed a number of structural and functional differences between the TCF4 mutant samples and their controls.
“Even without a microscope, you could tell which organelles in the brain had the mutation,” said study lead author Alysson R. Muotri, PhD, professor at UC San Diego Medical School, director of the UC San Diego Stem Cell Program; member of the Sanford Consortium for Regenerative Medicine.
TCF4-mutant organoids were significantly smaller than normal organoids, and many of the cells were not actually neurons, but neural progenitors. These simple cells are meant to multiply and then mature into specialized brain cells, but in the mutant organelles, some of this process had gone wrong.
A series of experiments revealed that the TCF4 mutation led to deregulation of the SOX genes and the Wnt pathway, two important molecular signals that guide embryonic cells to proliferate, mature into neurons and migrate to the right place in the brain.
Due to this dysregulation, the neural progenitors did not proliferate efficiently and thus produced less cortical neurons. Cells that matured into neurons were less stimulated than normal and often remained concentrated instead of being arranged in finely tuned neural circuits.
This informal cellular architecture interrupted the flow of neural activity to the mutated organelle of the brain, which the authors said would likely contribute to impaired cognitive and motor function.
“We are surprised to see such significant developmental issues at all these different scales and it left us wondering what we could do to address them,” said Fabio Papes, PhD, an associate professor at the University of Campinas and a visiting scholar at UC. San Diego Medical School, which co-supervised the work with Muotri. Papes has a relative with Pitt-Hopkins Syndrome, who motivated him to study TCF4.
The team looked at two different gene therapy strategies for recovering the functional gene in brain tissue. Both methods effectively increased TCF4 levels and thus corrected Pitt-Hopkins syndrome phenotypes on a molecular, cellular and electrophysiological scale.
“The fact that we can repair this one gene and restore the entire nervous system, even at the functional level, is amazing,” Muotri said.
Muotri notes that these genetic interventions were performed at a prenatal stage of brain development, while in a clinical setting, children would be diagnosed and treated 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 recently licensed gene therapy tools in preparation for such a trial, in which the genetically engineered vertebral injections hopefully restore TCF4 function to the brain.
“For these children and their loved ones, any improvements in motor-cognitive function and quality of life would be worth it,” Muotri said.
“What’s really great about this work is that these researchers go beyond the lab and work hard to translate those findings into the clinic,” said Audrey Davidow, president of the Pitt Hopkins Research Foundation. “This is more than just an amazing academic project. it is a real measure of what well-practiced science can achieve in order to hopefully change human lives for the better. “
Reference: “Transcription factor 4 loss of function 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 ., 2 May 2022, Nature Communications.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, Sao Paulo Research Foundation (grants 2020 / 11451-7, 2018 / 03613-7, 2018/04240 ) and the US Department of Energy Joint Genome Institute (DE-AC02-05CH11231).