A U.S.-led international whole-exome sequencing study has discovered that mutations in genes encoding for the growth of the neurites necessary for neural connectivity increase the risk for cerebral palsy (CP), the authors reported in the September 28, 2020, edition of Nature Genetics.

Besides environmental factors, genomic variants should therefore be considered when assessing an individual child's risk for CP, which in future may help identify likely responders to currently available therapies.

"Until now, the role of genetics in CP has been controversial, as neonatal hypoxia was widely considered the leading cause of CP," said corresponding author Michael Kruer, an associate professor of child health, cellular & molecular medicine, neurology, and genetics at the University of Arizona College of Medicine in Phoenix.

"Our study provides the first firm statistical and laboratory evidence that genetic mutations may lead to CP in a substantial proportion of cases," said Kruer, director of the Pediatric Movement Disorders Program of the Barrow Neurological Institute at Phoenix Children's Hospital.

An important neurodevelopmental disorder (NDD) affecting motor function, CP is characterized by movement disorders, including spasticity, dystonia and/or ataxia in early childhood.

However, there remains considerable debate about CP's etiology, particularly in individual cases, which has important medical implications, as current treatment options are limited.

"Currently available treatments for CP are based on suppressing symptoms and may have limited benefit," Kruer told BioWorld Science.

"However, our study suggests it may be possible to approach CP from a new molecular and cellular perspective and develop therapies based on the underlying neurobiology."

Several environmental factors, most notably hypoxia, are known major CP risk factors, but up to 40% of cases may lack a clear etiology.

Registry-based data suggest 20-40% of CP cases have an associated congenital anomaly, implicating genomic alterations, with an estimated heritability of around 40%.

To date, five studies have analyzed genomic copy number variations (CNVs) in CP, identifying predicted deleterious CNVs in 10-30% of cases.

Additionally, a large CP whole-exome sequencing study reported possibly deleterious variants in 14% of 98 parent-offspring trios with unselected forms of CP.

Collectively, these studies suggest important genetic risks in CP, but a lack of robust statistical methods and controls impeded their ability to discover risk genes, while functional validation of candidate genes was not performed.

Drilling down

These limitations were addressed in the new Nature Genetics study, in which whole-exome sequencing of 250 parent-offspring trios showed significant enrichment of damaging de novo mutations in CP and identified single genes that could account for the disorder.

This was an international collaboration of clinicians and scientists, many of whom are members of the International Cerebral Palsy Genomics Consortium (ICPGC), which is currently chaired by Kruer.

"The ICPGC is dedicated to understanding the molecular basis of CP through data sharing and collaboration," said Kruer, noting, "the families in our cohort were from diverse ethnic backgrounds, including individuals from North America, Europe, Australia and Asia."

The study identified two new genes, FBXO31 and RHOB, and showed that the RHOB mutation enhanced active-state Rho effector binding while the FBXO31 variant reduced cyclin D levels.

"In unrelated patients, these two genes harbored two identical de novo variants, which were shown via cell and protein-based studies to disrupt the normal function of these proteins, confirming the changes in these genes as bona fide CP-associated mutations," said study co-lead author Sheng Chih Jin, an assistant professor of genetics and pediatrics at Washington University School of Medicine.

As such, "these genes should be added to the growing list of genes that may cause the clinical phenotype," Jin told BioWorld Science.

"The mutations in each of these genes are anticipated to disrupt neuritogenesis in the developing human brain, thereby leading to CP."

Relationship to other NDDs

Candidate CP risk genes were shown to overlap with other NDD genes, said Kruer, noting, "CP is known to overlap clinically and epidemiologically with other NDDs and many CP patients may have concurrent learning problems, autism or epilepsy."

Therefore, "we looked for overlap between the CP candidate genes identified in our study with genes known to be associated with intellectual disability, epilepsy or autism," he said.

"Our findings indicate that genetic overlaps mirror clinical intersections, suggesting there may be shared underlying mechanisms that could lead to one NDD in one case and a distinct NDD in another.

"This further suggests that in some cases there may be a single fundamental genetic mutation that alters early brain development leading to intellectual disability, CP and epilepsy," said Kruer.

The researchers used network analyses to identify enrichment of the Rho GTPase, extracellular matrix, focal adhesion and cytoskeleton pathways, suggesting these pathways may contribute to CP etiology.

"We found many genes that associated with CP and noticed they have much in common; instead of being random, they fall in pathways that we know are important for how the brain develops and makes connections," said Sara Lewis, a postdoctoral researcher in Kruer's laboratory and co-lead study author.

CP risk genes in enriched pathways were further shown to regulate neuromotor function in Drosophila reverse genetic screens conducted by Lewis.

The researchers estimate that 14% of CP cases could be attributed to an excess of damaging de novo or recessive variants, with these findings collectively providing evidence for genetically mediated dysregulation of early neuronal connectivity in CP.

In particular, the mechanistic insights derived from the identification of core pathways via such CP genomic studies may help guide therapeutic development in a field that has not seen a new treatment introduced for decades.

"Understanding when, where and how brain development takes a wrong turn in CP should allow us to refocus therapeutic efforts in order to target important etiological mechanisms," Kruer said.

Looking ahead, he said, "we will expand our discovery-based sequencing efforts and collaborate with other groups to enhance statistical power and discover additional genes and pathways.

Meanwhile, "we are assessing the clinical implications of how CP is diagnosed in the context of new genetic information and how knowledge of genetic causes may change the care of CP patients.

"We are also using patient-derived induced pluripotent stem cell-based neurons and Drosophila models to understand crucial mechanisms and how genetic changes alter developmental processes in order to develop targeted treatments." (Jin, S.C. et al. Nat Genet 2020, Advanced publication).

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