[2062] Level of Agreement between Paired Paraffin and Frozen Renal Tumor Samples on SNP Microarray

Selene Koo, Agata Minor, Poluru Reddy, Tatjana Antic, Loren Joseph, Carrie Fitzpatrick, Maria Tretiakova. University of Chicago, Chicago, IL

Background: Copy number alterations occur in many solid tumors including renal cell carcinomas (RCC), which can be classified due to distinctive chromosomal abnormalities. Single nucleotide polymorphism (SNP) microarrays may be useful for RCC characterization, as they provide whole-genome copy number data at higher resolution and can detect copy number-neutral loss of heterozygosity (LOH). While there have been advances in utilizing SNP microarrays in formalin-fixed paraffin-embedded (FFPE) tissue, few comparisons between results from FFPE tissue and fresh-frozen (FF) tissue have been made, and none of those studies investigated paired RCC samples.
Design: Paired, morphologically matched FF and FFPE tumor tissues were obtained from five RCC specimens, including clear cell, papillary, and chromophobe subtypes. Genomic DNA was quantitated using Picogreen, assessed for integrity using gel electrophoresis, and processed according to the standard protocol for Affymetrix Genome-Wide Human SNP Array 6.0. Acquired data were analyzed for copy number changes (CNC) and LOH using the Affymetrix Chromosome Analysis Suite.
Results: Following previously published optimization steps for FFPE samples, including increased amount of starting DNA, number of PCR cycles and DNase fragmentation time, CNC and LOH were compared for FFPE and matched FF samples. Results are summarized in the table.

 Identified in both FFPE and FFIdentified in FFPE, not FFIdentified in FF, not FFPE% agreement
Normal copy number850298%
Overall CNC1341048%
Whole/partial chromosome gain/loss94745%
Mosaic gain/loss40357%

While agreement between FFPE and FF tissue is excellent (98% agreement) for chromosomes where the copy number is unaltered, agreement is significantly decreased with CNC (48%). Specifically, SNP genotype data from FFPE samples were difficult to interpret, while SNP data from FF samples were readily interpretable and were used in multiple cases to confirm or reject apparent CNC. The most difficult-to-interpret CNC were whole chromosome gains and losses. Similarly, LOH could not be detected in the FFPE samples.
Conclusions: Although SNP microarrays on FFPE samples can provide useful genetic information, the overall agreement for CNC between FFPE and FF RCC samples is less than 50%. This low degree of agreement can be problematic for cytogenetic analysis and diagnosis for these tumors if only paraffin tissue is available. In an effort to improve data quality, we plan to expand the scope of this study in more cases using higher-resolution SNP microarrays and an FFPE-specific reference set for data analysis.
Category: Techniques

Wednesday, March 6, 2013 9:30 AM

Poster Session V # 277, Wednesday Morning


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