Genome-wide association study identifies two variants that may yield therapies
Researchers at Nationwide Children’s have found that two DNA variants that play a role in gene regulation are associated with prolonged ability to walk in boys with Duchenne muscular dystrophy (DMD).
Kevin Flanigan, MD, director of the Center for Gene Therapy, led what the research team believes is the first genome-wide association study to identify single-nucleotide polymorphisms (SNPs) influencing ambulation in boys with DMD. SNPs are the most common genomic variations in humans and represent a single change in a DNA building block, a nucleotide.
“Within Duchenne muscular dystrophy, there’s variability in severity. For example, some boys may stop walking at age 9 and other boys may stop walking at 12,” says Dr. Flanigan, who is also a professor of pediatrics and neurology at The Ohio State University College of Medicine. “We’re trying to understand the genetic causes of this variability.”
Veronica Vieland, PhD, director of the Battelle Center for Mathematical Medicine, led the statistical analysis of about 2.5 million SNPs distributed across the entire genome that showed two SNPs to be associated with a delay in loss of ambulation, suggesting the variants are protective.
The SNPs are found in stretches of DNA that do not encode genes, but instead appear to influence two specific genes at a distance. One SNP regulates the latent transforming growth factor beta-binding protein 4 (LTBP4) gene and the other regulates the thrombospondin-1 (THBS1) gene. Both of these genes encode proteins that control levels of the protein TGFβ, a cytokine secreted by many cells that has a role in regulating cell proliferation, differentiation, and inflammation, among other functions.
The two SNPS were associated with a decrease in TGFβ levels and signaling as well as longer ambulation. The finding, published in the Annals of Neurology, suggests potential pathways for therapies to modify DMD in children.
Flanigan led an earlier study of 253 boys with DMD that found a different variation within the LTBP4 gene itself influenced the age of ambulatory loss. Unlike the current results, the previous study looked specifically at the influence of LTBP4 after studies in mice had suggested that variation within that gene influenced the severity of muscle pathology.
“The current study, funded by the National Institute of Neurological Disorders and Stroke, took an unbiased approach instead of looking at one specific candidate gene,” Dr. Flanigan says.
The researchers studied the same 253 boys, identified among those who are followed in the United Dystrophinopathy Project, a longstanding patient database directed by Dr. Flanigan. This time the team used the latest tools to analyze a significant portion of SNPs across their entire genomes, and then employed statistical analyses to identify associations.
“This is considered to be a very small sample size for this kind of a study yet we found two SNPs at a genome-wide significance level,” says Dr. Vieland, who is also a professor of pediatrics and statistics at The Ohio State University.
As both genes are in the TGFβ pathway, the researchers postulate that the protective effects may be additive. For example, the researchers found that the patient who was oldest when he lost the ability to walk had protective variants, or alleles, for both genes on each of his chromosomes. Compared with patients who had none or one protective LTB4 allele, those who had the protective LTB4 allele on both chromosomes walked on average 1.2 years longer if they had no THBS1 protective allele, 3.5-years if they had one THBS1 allele and 6.8-years if they had two.
Drs. Flanigan and Vieland, along with their collaborator, Dr. Robert Weiss of the University of Utah, are now looking for modifiers that influence breathing, heart function and muscle performance in more than 650 DMD patients.
“We believe there are other modifiers that will prove important, by both identifying new pathways to target for therapies and by helping to expand our understanding of responses to the gene restorative therapies under development here at NCH and elsewhere,” Dr. Flanigan says.
In addition, the findings may inform clinical trials, Dr. Vieland says. “If genetic background is influencing phenotype, we need to know this going in. You may get most of the kids who you expect to do well in one arm of the study – getting the medication or the controls — and that could skew the outcome.”
Citation: Weiss RB, Vieland VJ, Dunn DM, Kaminoh Y, Flanigan KM, and the United Dystrophinopathy Project. Long-range genomic regulators of THBS1 and LTBP4 modify disease severity in Duchenne muscular dystrophy. Annals of Neurology. 2018 Aug;84(2):234-245.