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

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 of prognostic genomic aberrations in SLL in a clinical diagnostic setting that could be implemented in patient risk stratification.