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Scientists have used the CRISPR-Cas9 gene edition to alleviate some autism symptoms in mice with some form of fragile X syndrome, the most common cause of autism spectrum disorder.
Using gold nanoparticles to deliver the Cas9 enzyme into the brain, a technique developed at the University of California at Berkeley, and called CRISPR-Gold, researchers were able to edit the gene for 39, a neurotransmitter receptor and reduce repetitive behavior. Fragile X syndrome (FXS).
Since repetitive exaggerated behaviors are common features in autism spectrum disorders, the effective reduction of these behaviors in FXS mouse models demonstrates the potential application of this technique to other types of autism where the genetic cause is known, the researchers said.
"There are still no treatments or cures for autism, and many clinical trials of small molecule therapies targeting the proteins that cause autism have failed," Hye Young said. Lee, Adjunct Professor of Cellular and Integrative Physiology at the University of Texas. Center in San Antonio, said. "It is the first case where we have been able to edit a causative gene for autism in the brain and show the rescue of behavioral symptoms."
CRISPR-Gold has many advantages over other ways to put Cas9 in the body, such as using viruses, according to the researchers.
"The really compelling thing about this document is that Hye Young was able to show that if you injected CRISPR-Gold into the brain, you could eliminate pathogenic genes and see quite significant behavioral changes," Niren Murthy, CRISPR-Gold inventor and a UC Berkeley professor of bioengineering. "It's the first time anyone has shown it with a non-viral delivery."
Those with autism spectrum disorders have problems interacting with other people as well as exaggerated repetitive behaviors, such as tipping and flapping. Although ASD appears to have various causes, including multiple genetic mutations, monogenic disorders such as FXS are a simpler way to explore the causes and potential treatments.
While autism spectrum disorders affect more than one percent of all children, FXS is rare, occurring in one in every 4,000 and 6,000 girls.
The results were published on June 25 in the monthly newspaper, Biomedical engineering of nature.
The new study is the first demonstration that the Cas9 protein can be transported into the brain to knock out a gene and have therapeutic effects.
While other researchers have inserted genes for Cas9 into neurons via viruses such as the adeno-associated virus, problems arise because the gene continues to express the Cas9 enzyme, leading to random cutting. Other genes.
CRISPR-Gold transports the Cas9 complex itself, purified Cas9 protein and guide RNA, directly into the cells, where it cuts a few times and then disappears.
"If you inject CRISPR DNA using a virus, you can not control the amount of Cas9 protein and the guide RNA are expressed, so the injection via a virus has a potential problem" Lee said. "I think the CRISPR-Gold method is very cool because we can control the amount we want to inject and that probably minimizes the side effects of using CRISPR, for example out-of-target effects."
The technique opens the door to treating conditions ranging from opioid addiction and neuropathic pain to schizophrenia and epileptic seizures, said Murthy.
The amortization of an over-excited brain
In the experiment on mice with FXS, the researchers injected CRISPR-Gold carrying the Cas9 complex into the brain striatum, a region known for mediating habit formation, including that related to repetitive behaviors common to ASD.
Cas9 targeted an excitatory receptor, the metabotropic glutamate receptor 5 (mGluR5), which is involved in communication between neurons and is known to be dysregulated in FXS. By disabling the mGluR5 gene, researchers were able to mitigate the exaggerated signaling between cells and reduce repetitive behavior.
"Prior to this experiment, we did not know if the mGluR5 receptor in the striatum was specifically involved in exaggerated repetitive behavior, it is an important biological finding of our study," said Lee.
In mice with FXS, repetitive behavior included obsessive digging and periodic air jumps. Digging was reduced by about 30 percent, while the jump dropped by 70 percent. Approximately 50% of the mGluR5 genes in the striatum were edited and reduced the number of receptor proteins by almost half.
Pharmaceutical companies have tried to inject small molecule drugs into the bloodstream to block the same receptor, but if some reduction in repetitive behavior was noted, the mice did not respond to subsequent treatments, becoming apparently tolerant.
The promising CRISPR-Gold system was developed by Murthy, which focuses on the administration of drugs and the development of new antibiotics.
The technique uses gold nanoparticles coated with a forest of DNA chains that contain Cas9 molecules, a combination of a gene enzyme and a guide RNA that houses the mGluR5 gene. The package is encapsulated in a polymer that helps in getting into the proper cells.
Last year, Murthy and his colleagues demonstrated that CRISPR-Gold could carry Cas9 into muscle cells and replace a mutated gene with a normal gene to enhance strength in Duchenne muscular dystrophy mice.
The new paper proves that CRISPR-Gold can successfully get Cas9 into a variety of cells in the brain.
"We showed in this article that we were also able to edit non-neurons: microglia, which are part of the brain's immune system, and astrocytes, which support neurons," Murthy said. "We have edited them more efficiently than neurons, and these can play a very important role in many diseases."
In 2016, Murthy's lab created a start-up, GenEdit, to produce and test CRISPR-Gold therapies for a variety of genetic diseases.
The successful release of brain cells opens the door to the treatment of many disorders, according to GenEdit CEO and U.C. Kunwoo Lee of Berkeley.
"CRISPR-Gold can be used to treat a variety of genetic diseases, such as Huntington's disease," Lee said. "But it is not limited to monogenic diseases, it can also be used against polygenic diseases, once we understand the entire network of genes involved."
Important for this study, GenEdit found how to ship long-range CRISPR-Gold particles from Berkeley to San Antonio, and manufacture them reproducibly, eliminating a bottleneck that has limited the translation of many other nanotechnologies Murphy said.
Researchers are currently developing CRISPR-Gold particles that can be injected directly into the central nervous system through the spinal cord, thus avoiding opening the skull and injecting directly into the brain.
Lee is optimistic that it could be as effective as intracranial injection into the striatum to reduce repetitive behavior, and perhaps even solve some of the social interaction problems of ASDs.
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