Microscopic images show significant differences in size and structure between brain organelles obtained from a patient with Pitt-Hopkins syndrome (right) and control (left). Credit: UC San Diego Health Sciences
A study by the University of California, San Diego (UCSD), uses laboratory-grown human brain tissue to diagnose neurological pathologies in Pete-Hopkins syndrome and to test gene therapy tools.
In a study published in the journal Nature Communications on May 2, 2022, 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 neural structure and function.
Several neurological and neuropsychiatric diseases, including autism spectrum disorder (ASD) and schizophrenia, are associated with a mutation in transcription factor 4 (TCF4), which is an essential gene for brain development. Transcription factors regulate the activation or deactivation of other genes, so their presence or absence may have a domino effect in the developing embryo. Yet little is known about what happens to the human brain when a TCF4 mutation occurs.
To address this question, the researchers focused on Pete-Hopkins syndrome, ASD, which is specifically caused by mutations in TCF4. Children with a genetic condition have profound cognitive and motor disorders and are usually nonverbal.
Pitt-Hopkins Syndrome (PTHS) is a rare genetic disorder characterized by developmental delay, epilepsy, distinctive facial features, and possible periodic hyperventilation followed by apnea. As more is discovered about Pete-Hopkins, the range of developmental disorders expands to include complications related to autism, anxiety, ADHD, and sensory disorders. It is linked to an abnormality on chromosome 18, specifically the inadequate expression of the TCF4 gene.
Existing mouse models of Pete-Hopkins syndrome do not accurately reflect the neural characteristics of patients, so the UCSD team instead developed a human study model for the disorder. Using stem cell technology, they transformed patients’ skin cells into stem cells, which later evolved into three-dimensional brain organoids, or “mini-brains.”
Initial observations on brain organoids revealed structural and functional differences between TCF4 mutant samples and their control.
“Even without a microscope, you can tell which brain organoid had a mutation,” said lead researcher Allison R. Muotrim, Ph.D., Professor at the UC San Diego School of Medicine, Director and Member of the UC San Diego Stem Cell Program. Sanford Consortium of Regenerative Medicine.
TCF4-mutated organoids were substantially smaller than normal organoids, and many cells were not actually neurons but neural precursors. These simple cells aim to multiply and then mature into specialized cells in the brain, but in mutated organelles some of this process was reversed.
A series of experiments showed that the TCF4 mutation caused the downstream disruption 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.
Because of this disorganization, neural precursors did not proliferate efficiently and, consequently, fewer cortical neurons were produced. Cells that matured into neurons were less excitable than normal and often stayed together in groups, instead of being sorted into finely tuned neural circuits.
This atypical cellular architecture has inhibited the flow of neural activity into the mutated brain organoid, which the authors say is likely to contribute to impaired cognitive and motor function.
“We were surprised to see such key development issues across all these different scales and wondered what we could do to address them,” said first author Fabio Papes, PhD, associate professor at the University of Campinas and visiting scientist at UC. The San Diego School of Medicine, which co-led the work with Muotri. Pipes has a relative with Pete-Hopkins Syndrome, which motivated him to study TCF4.
The group tested two different gene therapy strategies to restore functional genes in brain tissue. Both methods effectively increased TCF4 levels and thus corrected Pit-Hopkins syndrome phenotypes at the molecular, cellular, and electrophysiological scales.
“The fact that we can repair this one gene and the whole nervous system recovers itself, even at the functional level, is amazing,” Muotri said.
Muotri notes that these genetic interventions were performed in the prenatal stage of brain development, while children in the clinical setting were diagnosed and treated several years later. Thus, clinical trials must first confirm whether further intervention is safe and effective. The team is currently optimizing their recently licensed gene therapy tools for such an experiment, in which genetic vector spinal injections will hopefully restore TCF4 function in the brain.
“For these children and their loved ones, any improvement in motor-cognitive function and quality of life was worth the effort,” Muotri said.
“What really stands out in this paper is that these researchers go beyond the lab and work hard to bring these findings to the clinic,” said Audrey Davidov, president of the Pete Hopkins Research Foundation. “It’s much more than a stellar academic paper; “It’s a true measure of what good practical science can do to hopefully change people’s lives for the better.”
Reference: “Loss of transcription factor 4 function is associated with progenitor proliferation and cortical neuron deficiency” Fabio Papes, Antonio P. Camargo, Janaina S. de Souza, Vinicius MA Carvalho, Ryan A. Szeto, Erin LaMontagne, José R. 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 are: 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, José R. Techeira, Thiago s. Nakahara, Carolina n. Santo, Barbara MP at the University of Arajoo and Paulo Enf Velho Campinas.
This work was partially funded by the National Institutes of Health (Grant R01 MH123828), the Pete Hopkins Research Foundation, the S სანo Paulo Research Foundation (Grants 2020 / 11451-7, 2018 / 03613-7, 2018 / 04240-0) and a joint effort of the US Department of Energy Institute of Genome (DE-AC02-05CH11231).