420 Clinical Utility of Single Nucleotide Polymorphism (SNP) Microarrays in Pediatric Cancer and Non-Malignant Hematologic Disorders Xin-Yan Lu, Yi-Jue Zhao, Sivashankarappa Gurusiddappa, Ching C. Lau, Jason M. Shohet, Pulivarthi H. Rao, Karen R. Rabin, Sharon E. Plon Texas Children’s Cancer and Hematology Centers, Department of Pediatrics, Baylor College of Medicine, Houston, TX
Array-based technology has been showing great impact in clinical cancer cytogenetics. This study was to test the feasibility of single nucleotide polymorphism (SNP) microarrays in the clinical diagnosis of pediatric cancers. A total of 74 cases, including 39 acute leukemias (27 B-cell precursor acute lymphoblastic leukemia [BCPALL], 6 T-cell lymphoblastic leukemia, 5 acute myeloid leukemia, and 1 mix-phenotypic acute leukemia), 15 solid tumors, 5 myelodysplastic disorders, 6 non-malignant hematologic disorders, 5 lymphomas, 1 chronic myeloid leukemia cell line, as well as 3 normal remission marrows were tested using Illumina Cyto 12 and/ or 1M quad SNP arrays. Overall the SNP array data showed good concordance with cytogenetic/FISH data. SNP arrays revealed most cytogenetically unidentifiable abnormalities including copy neutral loss of heterozygosity (CN-LOH). In addition, SNP arrays provided prognostic information in 10 acute leukemia cases previously reported with normal cytogenetics and/or FISH and detected deletion of cancer genes (PAX5, MLL, PTEN and EVI1, etc.) at the exon level. Importantly, in 1 case with thrombocytopenia and a known constitutional RUNX1 gene deletion, SNP array revealed a low level (w20%) of mosaic CN-LOH for chromosome 21; and in 2 BCP-ALL cases reported as hyperdiploid by chromosome /FISH analyses, SNP array detected CN-LOH in all disomic chromosomes, indicating that the hyperdiploidy in fact resulted from doubling of a hypodiploid clone. Finally, SNP array detected 3 novel amplifications at 2p25.2 in a neuroblastoma case originally reported with MYCN amplification and amplicons at 2p25.1wp24.3 encompassing 16 genes in a high-grade glioma originally reported with double minute chromosomes. In conclusion, SNP arrays could not only confirm most karyotypic/FISH data but could also detect additional genomic aberrations, some with significant clinical implications. This pilot study shows that SNP array has great potential as a diagnostic tool in pediatric cancer cytogenetics and can be integrated into routine clinical cancer cytogenetic testing. Conflict of Interest: None.
Atlas of Cytogenomics in Oncology and Hematology: a Platform-Neutral Clinical Cancer Genomics Database Bixia Xiang a, Annette Leon b, Marilyn M. Li c, Anwar M. Iqbal d, Peining Li e, Shibo Li f, Peter R. Papenhausen g, Stuart Schwartz g, Xiao-Xiang Zhang g, Katherine B. Geiersbach h, Sarah South h, Guangyu Gu h, Jacqueline R. Batanian i, Xinyan Lu j, Daynna J. Wolff k, Iya Znoyko k, Rajyalakshmi Luthra l, Su S. Chen l, Keyur P. Patel l, Rachel L. Sargent l, Rizwan C. Naeem m, Lina Shao n, Renu Bajaj o, Stephen C. Peiper o, Zi-Xuan (Zoe) Wang o, Teresa Smolarek p, Lauren S. Jenkins q, Xu Li q, Feng Li q, Sainan Wei r, Jennelle C. Hodge s, Joyce L. Murata-Collins a, Zhiwei Che t, Ausaf Ahmad d, Ming Qi u, Stephen J. Forman a, Gail H. Vance v, Robert G. Best w a Cytogenetics Laboratory, City of Hope National Medical Center; b GenPath Diagnostics, BioReference Laboratories; c Department of Molecular and Human Genetics, Baylor College of Medicine;
Abstracts d
Department of Pathology and Laboratory Medicine, University of Rochester Medical Center; e Molecular Cytogenetics Laboratory, Department of Genetics, Yale School of Medicine; f Oklahoma University Health Sciences Center; g Laboratory Corporation of America; h Department of Pathology, ARUP Laboratories, University of Utah; i Department of Pediatrics and Pathology, Cardinal Glennon Children’s Hospital, Saint Louis University School of Medicine; j Texas Children’s Hospital, Baylor College of Medicine; k Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Molecular Diagnostic Laboratory, Department of Hematopathology; l The University of Texas M.D. Anderson Cancer Center; m Albert Einstein College of Medicine and Montefiore Medical Center; n Department of Pathology, University of Michigan; o Department of Anatomy, Pathology and Cell Biology, Thomas Jefferson University Hospital; p Cincinnati Children’s Hospital Medical Center; q Cytogenetics Laboratory, Kaiser Permanente San Jose Medical Center; r Department of Pediatrics and Human Development, Michigan State University; s Department of Laboratory Medicine and Pathology, Mayo Clinic; t Department of Application Science, BioDiscovery, Inc; u Center for Genetic and Genomic Medicine, Zhejiang University School of Medicine, James Watson Institute of Genome Sciences, P.R. China; v Department of Medical and Molecular Genetics, Indiana University School of Medicine; w University of South Carolina School of Medicine
Clinical interpretation of complex somatic genomic data remains the largest obstacle for the widespread application of array technologies for personalized medicine approaches in hematology and oncology specimens. To address this challenge, a database, tentatively named the Atlas of Cytogenomics in Oncology and Hematology, has been designed with 2 major objectives: 1) cataloging copy number alterations (CNA) and loss of heterozygosity (LOH) incidence maps to build strong empirical evidence to facilitate interpretation of clinical cytogenomic findings, and 2) generation of new knowledge by using machine learning algorithms. The database already contains over 2000 cancer array cases from 22 clinical laboratories. Raw genomic data and associated clinical information are accepted from any array platform and currently includes Agilent, Affymetrix, Illumina, and NimbleGen. Cases span the common clinical indication categories including: chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), acute lymphocytic leukemia (ALL), myeloproliferative neoplasms (MPN), plasma cell myeloma (PCM), and some solid tumors. For each disease group, the database provides: 1) Incidence maps for CNA and LOH, 2) Hyperlinks to relevant cytogenetics resources, and 3) Cytogenomic literature collection, summary, and an interface for user interaction. This database is dynamic: acquiring continuous data contribution from clinical laboratories, being scalable to whole genome sequence (WGS) data, and accommodating increasingly complex cancer indications stratification. While the initial purpose is as a catalog of CNA, LOH, and genotype-phenotype correlations from clinical empirical data, the machine learning algorithms will identify new insights into complex genomic patterns toward the goal of developing personalized evidence-based therapeutic interventions. Conflict of Interest: None.
Ascertainment of Recurrent Translocations by Chromosomal Microarray Analysis Guangyu Gu a,b, Maria Sederberg b, Christian Paxton b, Leslie Rowe b, Katherine Geiersbach a,b, Sarah South a,b a Division of Medical Genetics, University of Utah, Salt Lake City, UT; b ARUP Laboratories, Salt Lake City, UT
Abstracts Detection and characterization of recurrent translocations play an important role in the diagnosis and treatment of hematological disorders. Chromosomal microarray analysis (CMA) is a powerful tool to detect copy number changes in hematological disorders. One of the limitations of CMA platforms currently in use is that truly balanced chromosome rearrangements cannot be detected. However, some chromosomal rearrangements have cryptic losses or gains at the breakpoints which may be detected by microarray. In addition, recurrent balanced translocations followed by gain or loss of one of the derivative chromosomes can be detected by CMA, and recurrent interstitial deletions resulting in gene fusions may also be detected by microarray. We reviewed over 50 cases where a high-density SNP-oligo microarray was used to detect chromosome aberrations in hematologic cases. The high density design combined with allele detection and good probe performance allowed for precise identification of breakpoints on the rearranged chromosomes, which was critical for detection and interpretation of the translocations. Many of the suspected recurrent translocations were also confirmed by FISH, which can potentially be used as markers in subsequent studies. Several cases will be discussed in detail in this study. Conflict of Interest: None.
Evaluation of SureFISHtm DNA-FISH Probes for Validation of Array-CGH Aberrations Linda I. Barenboim, Jane Houldsworth Cancer Genetics, Inc., Rutherford, NJ
Currently in a diagnostic setting, genomic copy number changes detected by array-CGH must be validated by an independent technique, most commonly FISH. Given the multiple aberrations observed in cancer genomes, SureFISH repeat-free oligonucleotide probes (Agilent Technologies) offer an alternative to BACbased probes with respect to cost and time, though their performance has as yet to be assessed on a variety of tissue types. We investigated the performance of three SureFISH combination probes on peripheral blood (PB), bone marrow (BM), liquid biopsy cervical specimen (LBCS), and formalin fixed paraffin embedded (FFPE) breast tissue. Initially, hybridization conditions were optimized for each of the tissues based on a routine procedure used for BAC-based probes and the manufacturer’s recommendations. SureFISH ABL1 and BCR probes were hybridized to PB, BM (known to be positive for the ABL1/ BCR rearrangement), and FFPE where optimal signal quality and background were obtained using a 24-hour hybridization rather than 6-hour. Wash conditions were as recommended, with the exception of FFPE, where temperatures were decreased. Following the modifications, all hybridizations were considered acceptable for use in a clinical diagnostic setting despite the relatively smaller signal size compared with BAC-based probes. The second probe combination (chromosomes 13, 18, and 21) assessed was hybridized to PB, BM, and LBCS with similar findings. A custom probe for regions on chromosomes 3, 5, 7, and 20 was evaluated on PB and LBCS but optimization was required to balance probe concentrations with respect to signal strengths and fluorochrome. Hybridization signals could be accurately scored using an automated system. In conclusion, we have shown that SureFISH probes can be successfully used on a variety of specimen types that would be encountered in a clinical cytogenetics laboratory performing array-CGH, but requires some technical optimization.
421 Conflict of Interest: The authors are employees of Cancer Genetics, Inc., Rutherford, NJ.
The HEL Erythroleukemia Cell Line as a Test Case for Detailed Description of a Complex Karyotype by Combining SNP Array with Multiple FISH Approaches Ruth N. MacKinnon a,b, Adrian Zordan a, Meaghan Wall a, Lynda J. Campbell a,b a Victorian Cancer Cytogenetics Service, Melbourne, Australia; b Department of Medicine, St. Vincent’s Hospital, University of Melbourne
The HEL cell line is an erythroleukemia cell line that is used by many laboratories to study cell biology and differentiation. Several karyotypes have been published but these have many uncertainties. Using a combination of SNP array, array CGH, single locus FISH, multicolor FISH, and multicolor banding, we have produced a detailed karyotype which defines the breakpoints and structure of most of the abnormal chromosomes. We used SNP array data to identify the breakpoints of unbalanced translocations identified by M-FISH. Because the karyotype is very complex, there were still many ambiguities. Most of these could be clarified using M-BAND or locus-specific FISH probes selected using SNP array copy number data. Selected use of centromere probes identified the centromeres present in some abnormal chromosomes, including dicentric chromosomes. Using this combined approach we deduced that some complex rearrangements had occurred via the formation of dicentric chromosomes and subsequent breakage-fusion-bridge events. This included the production of a der(9) with concomitant amplification of JAK2, MLL, and the 20q11.2 region, which is amplified in some cases of del(20q) acute myeloid leukemia. B allele frequency data provided by SNP array enabled the aberrations involving different homologs of a chromosome to be distinguished in many instances. By combining SNP array data with various FISH techniques, karyotype abnormalities can be described more completely than by microarray alone. This approach takes advantage of advanced microarray karyotyping without losing information on translocation partners, dicentric chromosomes, and mechanisms of karyotype evolution that may be relevant to understanding cancer evolution in a research setting. Conflict of Interest: None.
SESSION 2: SOLID TUMORS Analysis of Esophageal Adenocarcinoma Using Combined aCGH e SNP (CCMC v2+EA) Microarray Platform Ausaf Ahmad a, Santhoshi Bandla b, Tony E. Godfrey b,c, M. Anwar Iqbal a a Department of Pathology and Laboratory Medicine, University of Rochester, Rochester, NY; b Department of Surgery, University of Rochester, Rochester, NY; c The James P. Wilmot Cancer Center, University of Rochester, Rochester, NY
Esophageal adenocarcinoma (EAC) is one of the most common cancers with very poor survival rates. We applied custom designed aCGH e SNP microarray platform (4180K, CCMC