t is not news to us in the field that researchers looking to determine causation of Neurodevelopmental Disorders have zeroed in on Molecular Proteins in the brains. To be specific, these disorders (namely Epilepsy, Intellectual Disability, Autism Spectrum Disorder, and Attention Deficit Hyperactivity Disorder) are being hailed as brain-based disorders due to the surging evidence in the last 2 years that indeed, some molecular proteins are atypical in both brain origin and development.
Let’s begin the survey in August 2013 where genetic studies were initiated in large scale. An international study on the genes involved in Epilepsy Disorder had uncovered 25 new mutations on 9 key genes behind a devastating form of epilepsy disorder during childhood. Among those were two genes never before associated with this form of epilepsy. One of these genes previously had been linked to autism and a rare neurological disorder, for which an effective therapy had previously been developed. With the findings of this research, the direction for developing genome-wide diagnostic screens for newborns to identify who is at risk for epilepsy improves potentially development of precise therapies for the condition.
“The limitations of what we currently can do for epilepsy patients are completely overwhelming,” said Daniel Lowenstein, MD, a UCSF neuroscientist and epilepsy expert. Along with Ruben Kuzniecky, MD from New York University, the pair was overseeing the Epilepsy Phenome/Genome Project (EPGP). “More than a third of our patients are not treatable with any medication, so the idea of finding specific drug targets, instead of a drug that just bathes the brain and may cause problems with normal brain function, is very appealing.”
“We knew there was something happening that was unique to these kids, but we had no idea what that was,” said Elliott Sherr, MD, PhD. He a pediatric neurologist at UCSF Benioff Children’s Hospital, and is the principal investigator of the Epi4K Epileptic Encephalopathy (EE) project. He was responsible for the development of this group of the target research patients within EPGP.
The team identified in in their research children with two classic forms of EE – infantile spasms and Lennox-Gastaut Syndrome – in which no other family member was affected. They excluded children who had identifiable causes of epilepsy, such as strokes at birth, which are a known risk for this group of disorders. Of the 4,000 patients whose genomes are being analyzed in the Epi4K, 264 children fit that description. The Epi4K sequencing team, led by David Goldstein, PhD from Duke University ran a genetic scan on the children and their parents. They compared their scans to thousands of people of similar heritage without epilepsy, used a cutting-edge new technique called exome sequencing. This method focuses on the exome, which is the 2 percent of our genetic code that represents active, protein-making genes. Those 25,000 genes are considered to be the code for what makes us unique, and is also responsible for disease mutations.
The genetic analysis revealed 439 new mutations in the children, with 181 of the children having at least one. Nine of the genes that hosted those mutations appeared in at least two children with EE and five of those had shown up in previous, smaller EE studies. Of the four other genes included, two may have been coincidental, the researchers found. But two new genes never before associated with EE – known scientifically as GABRB3 and ALG13 – each appeared with less than a one-in-40-billion statistical chance (p = 4.1×10-10) of being connected to EE by coincidence.
The findings implicated GABRB3, for the first time, as a single-gene cause of EE, and offered the strongest evidence to date for the gene’s role in any form of epilepsy, Sherr said. Knowing this about GABRB3, which is also involved with Angelman’s Syndrome, also offers the possibility that children with mutations only in this gene might benefit from the existing therapy for Angelman’s.
Another new gene, ALG13, is key to putting sugars on proteins, which points to a new way of thinking about the causes of and treatment for epilepsy.
‘The take-home is that a lot of these kids have genetic changes that are unique to them,” Sherr said. “Most of these genes have been implicated in these or other epilepsies – others were genes that have never been seen before – but many of the kids have one of these smoking guns.”
From GABRB3 and ALG13 genes in Epilepsy to misfiring neurons in the ADHD brain, the evidence continues to mount on how one size results do not fit all. In June 2104, Neuroscientists collaborating from the Mayo Clinic in Florida and rom Aarhus University in Denmark have shed light on why neurons in the brain’s reward system can be miswired, potentially contributing to disorders such as attention deficit hyperactivity disorder (ADHD).
In their study, scientists looked at dopaminergic neurons, which regulate pleasure, motivation, reward, and cognition, and have been implicated in development of ADHD. Together they unveiled a receptor system that is critical for correct wiring of the dopaminergic brain area during embryonic development. However they also discovered that after brain maturation, a cut in the same receptor, SorCS2, produces a two-chain receptor that induces cell death following damage to the peripheral nervous system.
It is the SorCS2 receptor that functions as a molecular switch between apparently opposing effects in proBDNF. ProBDNF is a neuronal growth factor that helps select cells that are most beneficial to the nervous system, while eliminating those that are less favorable in order to create a finely tuned neuronal network. The reserchers also found that some cells in mice deficient in SorCS2 are unresponsive to proBDNF and have dysfunctional contacts between dopaminergic neurons.
“This miswiring of dopaminergic neurons in mice results in hyperactivity and attention deficits. A number of studies have reported that ADHD patients commonly exhibit miswiring in this brain area, accompanied by altered dopaminergic function. We may now have an explanation as to why ADHD risk genes have been linked to regulation of neuronal growth,” says the study’s senior investigator, Anders Nykjaer, M.D., Ph.D., a neuroscientist at Mayo Clinic in Florida and at Aarhus University in Denmark.
On the other hand, a study published by Cell Press in the October 2014 issue of The American Journal of Human Genetics shows that Neurodevelopmental Disorders caused by distinct genetic mutations produce similar molecular effects in cells. This suggests a unique perspective in that a one-size-fits-all therapeutic approach could be effective for conditions, ranging from seizures to attention-deficit hyperactivity disorder.
“Neurodevelopmental disorders are rare, meaning trying to treat them is not efficient,” says senior study author Carl Ernst of McGill University. “Once we fully define the major common pathways involved, targeting these pathways for treatment becomes a viable option that can affect the largest number of people.”
Ernst and his team used human fetal brain cells to study the molecular effects of reducing the activity of genes that are mutated in two distinct autism-spectrum disorders. Changes in transcription factor 4 (TCF4) cause 18q21 deletion syndrome, which is characterized by intellectual disability and psychiatric problems. Mutations in euchromatic histone methyltransferase 1 (EHMT1) cause similar symptoms in a condition known as 9q34 deletion syndrome. “Our study suggests that one fundamental cause of disease is that neural stem cells choose to become full brain cells too early. This could affect how they incorporate into cellular networks, for example, leading to the clinical symptoms that we see in kids with these diseases,” Ernst says.
So far, we have learned about breakthroughs in genetic studies in Epilepsy, discoveries of misfiring of neurons in ADHD and in long lasting effects of mutations of certain brain cells leading to Intellectual Disability or psychiatric problems. Now let’s take a closer look at Austism Spectrum Disorder. Would we find some molecular or genetic aberration? Stanford University researchers in December 2014 mapped an entire molecular network of crucial protein interactions that contribute to autism.
While “much work remains to be done,” Dr. Charles Auffray of Université de Lyon who collaborated with the researchers, states this is “a bold attempt to leverage a number of rich sources of data and knowledge and to complement them with relevant additional measurements to unravel the molecular networks of ASD.”
Though further research is needed to fully understand autism’s origins, this study “contributes to the development of an openly shared methodological framework and tools for data analysis and integration that can be used to explore the complexity underlying many other rare or common diseases,” Auffray said.
In this current study of autism, the scientists did not just look at genes, they also looked at gene expression — the protein interactions — in patients with autism. After they had identified a “protein interaction module,” the researchers sequenced the genomes of 25 patients to confirm its involvement in autism. They then validated these findings with data from 500 additional patients. In the next step, the team examined gene expression within the module, partly by using the Allen Human Brain Atlas.
It was in this stage that the researchers discovered the brain’s corpus callosum and oligodendrocyte cells made important contributions to ASD. Developmentally, the oligodendrocyte cells help form myelin, the insulating sheath of brain cells necessary for high velocity nerve conduction. And for patients with autism, for instance, these cells exhibited extensive gene mis‐expression in the corpus callosum, the bundle of nerve fibers connecting left and right brain hemispheres.
The findings from the Stanford University study were not only supported in 2014 by the Heidelberg University but also given more specificity in the mutations not only for those with ASD, but for neurodevelopmental disorders in general. These German Researchers posited that generally, these disorders are multi-faceted and can lead to intellectual disability, autism spectrum disorder and language impairment. Mutations in the Forkhead box FOXP1 gene have been linked to all these disorders, suggesting that it may play a central role in various cognitive and social processes.
Dysfunction of motor, social, sensory and cognitive aspects play a major role in autism spectrum disorder (ASD) and intellectual disability (ID). A high comorbidity is often observed between these disorders, suggesting that mutations in critical genes can cause a spectrum of neuropsychiatric phenotypes. The Forkhead box transcription factor FOXP1, for example, has been linked to various cognitive disorders. FOXP1-specific deletions, mutations and chromosomal breakpoints interrupting the gene have been reported in patients with Intellectual Disability, Autism Spectrum Disorder, speech and language deficits, and motor development delay.
They were interested to examine the behavioral phenotype of our Foxp1 KO mice, as FOXP1 mutations are associated with various behavioral deficits in humans, including social unattainability, hyperactivity, altered learning and memory, and specific obsessions.Results showed: Foxp1 KO mice have a reduced ability for short-term recognition memory and memory for spatial contexts, which have been described before in ASD patients and in mouse models of ASD. The effect on spatial memory may be explained by the CA1 hippocampal deficits we observed in Foxp1 KO as the hippocampus is important for spatial memory. The disruption of the striatal region in Foxp1 KO mice may also contribute to the deficits in learning and memory. It has been shown that striatal lesions and infusion of the striatum with a dopaminergic antagonist results in impaired performance in spatial learning tests, while object recognition is impaired by administration of glutamate antagonists to the striatum. Interestingly, the striatum has previously been associated with the pathology of ASD in both mice and humans.
Foxp1 KO mice also displayed a higher occurrence of repetitive behaviours, in accordance with previous findings in mouse models of autism. Repetitive motor behavior is associated with abnormal activation of dopaminergic cortical-basal ganglia circuitry and therefore might partially be explained by the morphological disruption we observed in the striatal region.
They also recorded a striking reduction of social interest in Foxp1 KO mice. Difficulties communicating and interacting with other people is a key feature of human ASD, and reduced social interaction as well as hyperactivity has been reported in mouse models of ASD before. A strong PPI deficit was observed in Foxp1 KO mice, indicating impaired abilities for sensorimotor integration. Reduced PPI has been previously reported in ASD patients. This effect on PPI in Foxp1 KO mice may be partly explained by the reduction in the striatal region as a cortico-limbic-striatopallidal circuit is involved in the circuit regulating PPI.
Excitatory and inhibitory imbalance is a hallmark brain feature of Autism Spectrum Disorder. Several studies have reported that ASD-related mutations selectively impact glutamatergic or GABAergic synapses without affecting the other, leading to an imbalance of excitatory and inhibitory inputs. WIth their research, they have ultimately shown that the amplitude of miniature excitatory postsynaptic currents but not miniature inhibitory postsynaptic currents is larger in Foxp1 KO CA1 hippocampal neurons. This suggests that Foxp1 KO neurons receive a disproportionate magnitude of excitatory to inhibitory input. In addition, excitability of CA1 pyramidal cells was reduced in Foxp1 KO mice.
With all this information, it is possible to hypothesize that treatment protocol will also change to a more direct, molecular level based on the genetic misfiring or aberration. In the next post, we will discuss the current therapeutic interventions available for these disorders.