Use of Targeted Deep Sequencing To Analyze the Genomic Similarities of Concurrent Acute Myeloid Leukemia in the Peripheral Blood and Bone Marrow
Jeffery M Klco, David H Spencer, Chris Miller, Tamara Lamprecht, Robert S Fulton, Li Ding, Richard K Wilson, Timothy J Ley. Washington University School of Medicine, St Louis, MO; Washington University, St Louis, MO
Background: Acute myeloid leukemia (AML) is a clonally heterogeneous disease composed of a founding clone with hundreds of single nucleotide variants (SNVs) and frequently multiple subclones derived from the founding clone that contain additional variants. Recently it has been demonstrated that significant spatial heterogeneity exists within solid tumors. Whether AML demonstrates similar spatial heterogeneity (i.e. disease in the bone marrow versus peripheral blood), both at the level of specific subclones and individual SNVs, has not been evaluated, yet is critically important in the context of clinical testing.
Design: We identified 4 oligoclonal (3-4 clones) de novo bone marrow AML samples previously characterized by whole genome sequencing that also had significant peripheral blood involvement (69.2% blasts, mean). Using custom capture arrays containing all the known variants in these genomes (640.3 SNVs, mean) followed by deep sequencing on the Illumina platform, we analyzed the similarities in the variant allele frequencies (VAFs) in concurrent leukemia from the bone marrow and peripheral blood.
Results: We obtained deep coverage (569 x coverage, mean) of all the SNVs and exonic insertions/deletions identified in the 4 AML genomes, allowing us to digitally report VAFs and identify clusters of variants with similar VAFs, representing distinct subclones. When corrected for blast counts, the founding clones in each case were present at indistinguishable VAFs in the peripheral blood and bone marrow; the average difference in mean VAF between the founding clone in the peripheral blood and bone marrow was only 0.60% (range, 0.08-1.27%). Similar findings were seen for all the minor subclones. Analysis of individual cases demonstrated strong concordance for known AML mutations, including DNMT3A (BM: 52.5%; PB: 49.5%), NPM1 (BM: 44.6%; PB: 43.6%), IDH1 (BM: 34.4%; PB: 37.7%), and FLT3 (BM: 38.6%; PB: 31.4%).
Conclusions: In a small cohort of AML samples we found that clonal heterogeneity and VAFs of known pathologic mutations are similar in the bone marrow and peripheral blood at the time of diagnosis. These studies are currently being expanded to a larger cohort of AML samples and suggest that peripheral blood will be a suitable surrogate for clinical testing using next generation sequencing technologies for most AML cases.
Category: Special Category - Pan-genomic/Pan-proteomic approaches to Cancer
Tuesday, March 5, 2013 1:30 PM
Proffered Papers: Section H1, Tuesday Afternoon