Next-generation sequencing can sometimes find genetic mutations in challenging situations, but, Dr. Goldstein said, “we are getting answers only one-fourth to one-third of the time in cases where we expect a clear answer, presumed monogenic diseases. Why do we miss pathogenic variants?” His answer: “The variant is not called, possibly because the genomic region is difficult to sequence with current platforms; or the type of variant hard to call; or the variant is called but not recognized as pathogenic, possibly because of sample size.”
Dr. Goldstein offers a basic complex disease sequencing approach:
- Identify subjects, extreme phenotype or family based;
- Sequence (100+ individuals);
- Align to reference and call variants;
- Compare to hundreds of sequential controls;
- Follow up genotyping in larger cohorts; and
- Select experiment using thousands of samples.
In experiments on schizophrenia and epilepsy, however, Dr. Goldstein found that this sequencing approach using thousands of samples did not yield good evidence. He called this problem “locus heterogeneity blight” and asked, “What kind of sample size do you need as a function of the genes that carry mutations that influence the disease you are studying?” He noted that, “if locus heterogeneity is high, the sample size must be very high to get significant evidence.”
Looking to the future, Dr. Goldstein listed these steps for sequencing:
- Large sample sizes;
- Low to medium throughput;
- Functional evaluations; and
- Rephenotyping of patients after discovery of variants.
He also advocates establishing methods to evaluate biological functions of the variants implicated.
Drug Response
As part of the commitment to improving patient care as a goal of genetic study, Dr. Goldstein offered several reminders. Clinical trials data represent a critical opportunity to understand variable responses to treatment, he noted, and larger trials with quantitative measures of response provide important opportunities (often the case in rheumatology). Looking to the future, as risk factors for common diseases are identified, trial populations can and should be stratified.
Ann Kepler is a medical journalist based in Chicago.
