Clinical Laboratory Implementation of the Detection of Genomic Aberrations in Formalin-Fixed Paraffin-Embedded Small Lymphocytic Lymphoma Specimens by Array-CGH

Abstract

Cancer Cytogenomics Microarray Consortium Third Annual Meeting, August 6e7, 2012 SESSION1:HEMATOLOGICMALIGNANCIES Clinical Laboratory Implementation of the Detection of Genomic Aberrations in Formalin-Fixed ParaffinEmbedded Small Lymphocytic Lymphoma Specimens by Array-CGH Charles Ma, Asha N. Guttapalli, Lizalynn M. Dias, Lynnette M. Cahill, Lan Wang, Jane Houldsworth Cancer Genetics Inc., Rutherford, NJ Due to the highly variable course of the disease, risk stratification is essential for the management of chronic lymphocytic leukemia (CLL)/small lymphocytic lymphoma (SLL) patients. For CLL, assessment of prognostic molecular markers including IGHV mutation status and specific genomic aberrations by either FISH and/or, more recently, array-CGH is routinely performed. The same molecular markers have been shown to offer similar prognostic value in SLL. However, the clinical diagnostic evaluation of these markers in SLL is hampered by the solid tissue type. Indeed, formalin-fixed, paraffin-embedded (FFPE) material is often the only available specimen for analysis. Using over 300 FFPE samples, we have established appropriate laboratory procedures and QC matrices for the clinical implementation of array-CGH for FFPE samples. In brief, DNA is extracted from 5 ten micron FFPE sections, and then a minimum of 1ug of DNA is heat fragmented, labeled enzymatically, and hybridized with a similarly fragmented commercial reference DNA to a custom-designed DNA oligonucleotide microarray representing genomic regions that are commonly altered in mature B-cell neoplasms. For ten SLL FFPE samples, adequate DNA was isolated and found to be of varied quality. Using ADM-2, the same genomic aberrations as commonly found in CLL (13q14 loss (MIR15A/16-1), 17p13 loss (TP53), and 11q22 loss (ATM)) were detected in the SLL specimens with high reproducibility and accuracy. DNA dilution studies indicated an analytical sensitivity of 60%70% while FISH in the specimens for detected aberrations revealed sensitivity closer to 30%-40%. All aberrations detected by array-CGH were independently validated by quantitative PCR for genes mapped within the respective regions. Thus, our data indicated that array-CGH is suitable for the detection 2210-7762/$ see front matter a 2012 Elsevier Inc. Elsevier Inc. http://dx.doi.org/10.1016/j.cancergen.2012.07.004 of prognostic genomic aberrations in SLL in a clinical diagnostic setting that could be implemented in patient risk stratification. Conflict of Interest: These authors are employees of Cancer Genetics, Inc., Rutherford, NJ. Array-Based Karyotyping Post Plasma Cell Enrichment for the Detection of Genomic Abnormalities in Multiple Myeloma Barbara K. Zehentner , Luise Hartmann , Krystal Johnson , Christine Stephenson , Douglas Chapman , Monica de Baca , Denise A. Wells , Michael R. Loken , Budi Tirtorahardjo , Shelly R. Gunn , Lony Lim b HematoLogics Inc., Seattle WA; Combimatrix, Irvine, CA Multiple myeloma (MM) is a hematopoietic neoplasm characterized by malignant plasma cells. The discovery of genomic abnormalities present in the monoclonal plasma cells has diagnostic, prognostic, as well as disease monitoring implications. However, technical and disease-related limitations hamper the detection of these abnormalities by metaphase cytogenetic analysis or FISH. In this study, we tested 28 bone marrow specimens with known plasma cell neoplasm for the presence of genomic abnormalities using microarray analysis post plasma cell enrichment and compared the results with other laboratory findings. Metaphase cytogenetic studies were performed on 15 of the 28 samples and identified disease-related genomic aberrations in only 3 of the 15 cases (20%) due to the low proliferative rate of plasma cells; 84.6% of specimens tested positive by MACS-FISH and 89.3% by array comparative genomic hybridization (aCGH). Furthermore, aCGH detected additional abnormalities in 24 cases as compared with FISH alone. We conclude that the combination of plasma cell enrichment and genomic copy number assessment using CGH microarray is a valuable approach for routine clinical use to achieve a more complete genetic characterization of multiple myeloma patients. Conflict of Interest: Employment, stock ownership.