Researchers headed by teams at the Chinese Academy of Sciences, and Massachusetts Institute of Technology (MIT), have used CRISPR-Cas9 gene editing to engineer macaque monkeys with germline-transmissible mutations in Shank3, a gene that is linked with a form of autism and other human neurodevelopmental disorders. The engineered animals demonstrated altered brain connectivity patterns, and similar behavioral, motor, and social abnormalities that are seen in people with mutations in the same gene.
The researchers say the primate model is more relevant than existing rodent models for studying human neurodevelopmental disorders, and could potentially help scientists identify therapeutic strategies and drug candidates. “Our goal is to generate a model to help us better understand the neural biological mechanism of autism, and ultimately to discover treatment options that will be much more translatable to humans,” said MIT’s Guoping Feng, PhD, who is the James W. and Patricia Poitras professor of neuroscience, a member of MIT’s McGovern Institute for Brain Research, and one of the senior and corresponding authors of the scientists’ published paper in Nature, which is titled, “Atypical behavior and connectivity in SHANK3-mutant macaques.” Feng is also an institute member of the Broad Institute of MIT and Harvard and a senior scientist in the Broad’s Stanley Center for Psychiatric Research.
Shank3 is a protein found at the synapses between neurons, and mutations in its gene account for about 1% of spontaneous forms of autism spectrum disorder (ASD), the authors wrote. Mutations in the Shank3 gene are also a major cause of neurodevelopmental abnormalities in a rare disease known as Phelan-McDermid syndrome, which causes intellectual disability, impaired speech and sleep, and repetitive behaviors. Most patients with Phelan-McDermid syndrome are also diagnosed with ASD. “Patients with a Shank3 gene mutation often exhibit a variety of comorbid traits, which include global developmental delay, severe sleep disturbances, lack of speech or severe language delay, and characteristic features of autism spectrum disorder (such as social impairments and stereotypes),” the authors explained.
Prior studies by Feng’s team have shown that mice with Shank3 mutations exhibit some of the same traits associated with autism, including avoidance of social interaction, and repetitive behaviors, but mice aren’t ideal models for studying human neurodevelopmental disorders. Unlike humans and primates, mice don’t have a highly developed prefrontal cortex, which is the area of the brain associated with making decisions, sustaining focused attention, and interpreting social cues. These are all neurological processes affected by brain disorders. And while studying mouse models of autism and other neurodevelopmental disorders has resulted in the development of clinical-stage drug candidates, none of these has yet been successful, the team pointed out. “… it is increasingly apparent that the validity of Shank3-mutant rodent models for human patients is limited.”
“We urgently need new treatment options for autism spectrum disorder, and treatments developed in mice have so far been disappointing,” noted co-author Robert Desimone, PhD, the director of MIT’s McGovern Institute for Brain Research, and the Doris and Don Berkey professor of neuroscience. “While the mouse research remains very important, we believe that primate genetic models will help us to develop better medicines and possibly even gene therapies for some severe forms of autism.”
The researchers used CRISPR-Cas9 gene editing to engineer macaque monkeys (the founder animals) with mutations in the Shank3 gene, and subsequently confirmed through assisted reproductive techniques that using sperm with Shank3 mutations to fertilize a normal macaque oocyte could also give rise to offspring with one inherited mutant Shank3 gene. Magnetic Resonance Imaging (MRI) studies revealed changes to neural connectivity in the founder macaques, which indicated abnormal brain circuitry. There was evidence of reduced functional connectivity in the striatum and in the thalamus, an area of the brain that relays sensory and motor signals and is also involved in regulating sleep. In contrast, connectivity was strengthened in other regions of the mutant-Shank3 primates, including the sensory cortex.
Founder mutant Shank3 macaques also demonstrated behavioral patterns, including sleep disturbances and compulsive, repetitive behaviors, and reduced social interaction, that were similar to those seen in humans with the mutant gene. “These results suggest the Shank3-mutant monkeys display reduced levels of exploration, social interaction, and vocalization, which seem to parallel some aspects of the phenotypes that are found in humans with Phelan–McDermid syndrome or autism spectrum disorder,” the researchers stated. “Shank3 mutants exhibit notable sleep disturbances and activity differences, which may assist in the discovery of characteristic biomarkers for Phelan–McDermid syndrome, autism spectrum disorder, and other neurodevelopmental disorders in humans.”
The five founder Shank3-mutant macaques didn’t all display the same severity of behavioral and motor impairments and learning problems, possibly because the underlying genetic background of each animal was different, the authors pointed out. It will be important to confirm the reported findings in larger numbers of animals. Nevertheless, the scientists hope that within a year it may be possible to start testing potential treatments for autism-related symptoms. It might also be possible to identify biomarkers—such as the brain connectivity patterns observed in the macaque MRI scans—that might help to evaluate the effectiveness of drug treatments.
Similar approaches could also be used to study other neurological disorders such as Rett Syndrome and Fragile X Syndrome, which are also caused by well-characterized gene mutations. Fragile X is the most common inherited form of intellectual disability, and affects about 1 in 4,000 males and 1 in 8,000 females. Rett Syndrome, which is more rare and almost exclusively affects girls, results in severe impairments in language and motor skills and can also cause seizures and breathing problems.
“Given the limitations of mouse models, patients really need this kind of advance to bring them hope,” Feng said. “We don’t know whether this will succeed in developing treatments, but we will see in the next few years how this can help us to translate some of the findings from the lab to the clinic.”