In early studies, gain-of-function mutations in PCSK9 were shown to be associated with an increase in hypercholesterolemia in two families, and researchers then identified common genetic markers in wider studies of more generations of relatives, including those with xanthomas, coronary artery disease and strokes. In 2006, researchers also discovered a loss-of-function mutation in the gene that resulted in reduced cholesterol.3 Further genetics and genome-wide association study analyses explore SNP associations with myocardial infarction risk, and by 2013, a novel monoclonal antibody to inhibit PCSK9 was found to dramatically reduce LDL cholesterol.
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Explore This IssueMarch 2018
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“Rare, familial diseases helped us learn how to target cholesterol. Allele function is key here, so knowing whether it was a gain-of-function mutation or a loss-of-function mutation was a critical aspect of this drug discovery process,” he said. “Another key feature of this story was that having an intermediary, causal phenotype. We are interested in preventing stroke and heart disease, but we know that to do that, we can treat LDL levels. In autoimmune diseases, we don’t always have that.”
Another successful genetics and genome-wide association study used for drug discovery involved age-related macular degeneration (AMD). A rare penetrant mutation in CFH, a gene that makes the protein complement factor H, is related to high AMD risk.
“We wanted to find out if, in this locus, we could identify a rare coding functional variant that had a strong effect, it might help us understand the biology of AMD more clearly,” said Dr. Raychaudhuri. Rare coding variants might be useful in drug discovery because they are less likely to be influenced by natural selection. “Common variants, which are the basis of most of our complex traits, are usually selected away by evolutionary forces, so a rare variant might be more useful to derive insights.” Finding rare coding variants with large effects in complex traits like AMD or rheumatic diseases is complex, so many efforts to sequence genes for these phenotypes do not necessarily lend themselves to rare variants, he said.
Researchers knew that individuals with one haplotype, H5, had a dramatically higher risk of getting AMD, so they studied this haplotype in a large cohort to look for a rare coding variant, and found a heterozygous arginine chain in the CFH position that had a strong association. After a larger, targeted sequencing study, researchers discovered a rare variant in the C3 gene. In the complement pathway, CFH binds C3. Using a strategy called burden testing, they found another gene, CFI, which had a dramatic burden of rare variance. “If you look at AMD cases versus controls, you see this rare mutation spread across individuals who have the disease. In controls, you rarely see this mutation. When you look at loss-of-function alleles, all individuals who had this seemed to be getting the disease,” he said. Through genotype data and sequencing, they eventually discovered a rare mutation that resulted in a R1210C substitution associated with AMD risk, he said.4