CNVs Provide the Clue for a Cryptic t(5;11) in Acute Myeloid Leukemia

Abstract

s from the Cancer Cytogenomics Microarray Consortium/CAGdb meeting, Chicago, IL, August 5e7, 2013 Massive Parallel Sequencing of Leukemia Samples Using a Custom Designed Cancer Gene Mutation Panel Fengqi Chang, Liu Liu, Erica Fang, Guangcheng Zhang, Yanchun Li, Marilyn M. Li Baylor College of Medicine, Houston, TX, USA Leukemia is the result of accumulation of somatic mutations including point mutations, copy number variations, and chromosomal abnormalities. Since multiple mutations can be present in a given leukemia genome, a technology that can detect many mutations in different genes simultaneously is desirable. Next generation sequencing (NGS) technologies allow deep sequencing of numerous mutations and multiple samples at the same time. Among NGS technologies, targeted sequencing is appealing in clinical oncology due to its shorter turnaround time, lower costs, and simpler data analysis compared to whole genome or whole exome sequencing. We introduce here a new panel specific for leukemia using multiplex PCR followed by ion semiconductor deep sequencing technologies. The leukemia panel covers 48 genes and can detect over 3,200 leukemia-associated mutations, many of which are clinically actionable mutations. Twenty-five samples were used to validate the panel on different sample types, including blood, bone marrow, FFPE and cell free DNA. The uniformity of coverage was about 87.75% for all cases. The average depth of coverage for targeted region was more than 3000X and the mean 100X coverage of targeted mutations was 97.07%. The mutation panel detected mutations in NPM1, TP53, ABL1, JAK3, IDH1, IDH2, KRAS etc in these samples with as little as 20 ng DNA and the positive rate was about 56%. The most commonly mutated genes were: TP53 (7 cases) followed by CEBPA (3 cases), NPM1 (3 cases). Overall, the leukemia panel showed improved coverage uniformity, reading length and depth compared to certain commercially available kits. The turnaround time for the test can be as short as 3 to 5 days, which fits the needs of personalized oncology. Furthermore, samples can be multiplexed using barcoding, which leads to tremendous increase of sequencing efficiency and decrease of cost while maintaining the high quality of sequencing results. CNVs Provide the Clue for a Cryptic t(5;11) in Acute Myeloid Leukemia Nicole L. Chia , Howard R. Slater , Julia M. Potter a,b NU Medical School, Australian National University, Canberra ACT 0200, Australia; ACT Pathology, The Canberra Hospital, Canberra ACT, 2606 Australia; Victorian Clinical Genetics Services, Murdoch Children’s Research Institute, Royal Children’s Hospital, Melbourne, Vic 3052, Australia Molecular karyotyping has been introduced into diagnostic pathology services for first-tier investigation of constitutional anomalies. The clinical utility of this mode of diagnostic investigation in neoplasms is currently being explored. Evidence based interpretation of copy number change requires comparison with control cohorts. Cytogenetic investigation of neoplasms frequently requires identification of apparently balanced structural rearrangements that result in critical gene fusion. High resolution microarray investigations have shown that not all balanced rearrangements are in fact balanced at the sequence level. We describe a 25 year old male with acute myelomonocytic leukemia. A del(5)(q35) was identified as the sole abnormality by conventional cytogenetic analysis. SNP microarray analysis confirmed an acquired 6.0 Mb interstitial deletion at 5q35.1q35.3 and an adjacent 350kb deletion. In addition, two acquired adjacent deletions (2.1 Mb and 850kb) were observed at chromosome 11p15.5p15.4 and a 675kb gain at 10q11.22. Subsequent subtelomere FISH identified a cryptic t(5;11)(q35;p15) with putative NUP98/NSD1 fusion. Review of the “Aussie Normal” control cohort yielded no evidence of CNVs on chromosome 5 and 11 at the putative breakpoints. However, common CNVs were observed within the deleted regions for each chromosome. These are located within repetitive DNA (LINES and SINES). A low frequency CNV gain (220kb) in the Aussie Normal cohort, harboring the ANXA8L2 gene at 10q11.2 coincided with the CNV reported in the patient here. This CNV is located within a region of segmental duplication and may represent a rearrangement due to NAHR mechanisms. This case demonstrates the potential value for the application of molecular karyotyping of neoplasms. Comparison with control cohorts provides evidence upon which pathogenicity, predisposition, and putative mechanisms can be investigated.