WHOLE GENOME SEQUENCING OF MYELODYSPLASTIC SYNDROMES

Washington University, Saint Louis MO

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Abstract

DESCRIPTION (provided by applicant): PROJECT SUMMARY: This application responds to the NHLBI "Grand Opportunities" RC2 topic, "Large-scale DNA Sequencing and Molecular Profiling of Well-Phenotyped NHLBI Cohorts." We will use whole genome resequencing to identify genetic changes relevant for the pathogenesis of Myelodysplastic Syndromes (MDS). MDS is an increasingly prevalent clonal hematopoietic disorder that causes significant morbidity and mortality. Understanding of the genetic basis of MDS lags behind the closely related disease, acute myeloid leukemia (AML). Our group has pioneered the use of whole genome sequencing as a discovery tool in cancer genomics. We recently reported the complete DNA sequence of a cytogenetically normal AML genome and have now completed the analysis of a second AML case. Remarkably, we found eight novel non-synonymous somatic mutations in each case. One of these mutations is recurrent in 8.5% of 188 unselected adult AML cases and is associated with an adverse outcome. These results demonstrate the feasibility and utility of this strategy for mutation discovery. We will now apply these tools to MDS in the following specific aims: in Specific Aim 1, we will sequence the genomes of at least 10 paired tumor/normal samples from patients with intermediate risk de novo MDS. We will use "Next generation" sequencing to obtain 99% diploid coverage of the tumor and matched normal genomes. Copy number alterations (amplifications, deletions, and uniparental disomies) and RNA expression profiles will be analyzed on microarray platforms. In Specific Aim 2, we will perform bioinformatic analysis to identify all somatic mutations in at least 10 MDS genomes and define their frequencies in 144 other cases of MDS. Somatic mutations (present in the tumor, but not in the normal genomes) will be confirmed by deep resequencing of amplicon-based libraries. The prevalence of validated mutations will be assessed in pooled DNA obtained from a cohort of 144 MDS patients and recurrent mutations will be genotyped in individual samples. These experiments are ready to be initiated immediately, they can be completed in a two year time period, and with this significant short-term investment of resources, we can generate results that will greatly accelerate research in this field and have a lasting impact on the diagnosis, risk stratification, and treatment of patients with MDS. PUBLIC HEALTH RELEVANCE: The Myelodysplastic Syndromes are a poorly understood group of disorders that impair quality of life and shorten life expectancy. We will use state of the art sequencing technology to define the genetic basis of this disease. This represents a "Grand Opportunity" to rapidly increase knowledge that will have long-term benefits for the care of these patients.