Adequate tumor tissue for molecular studies was selected on H&E stained sections from frozen tissue tumor blocks. Areas of viable tumor highlighted on the slides were matched and macrodissected from the cryomolds for further nucleic acid extraction. Twenty-three samples with adequate quality RNA were studied on the U133A Affymetrix platform (22,000 transcripts), as previously described [9 (link)]. The transcriptional profile was compared to the expression of a well-characterized set of 34 soft tissue sarcomas (STS), spanning the most common histologic types, including: synovial sarcoma, clear cell sarcoma, gastrointestinal stromal tumor, myxoid liposarcoma, dedifferentiated liposarcoma, leiomyosarcoma, malignant fibrous histiocytoma, and fibrosarcoma [10 (link)]. These STS samples used in our original publication [10 (link)] were initially studied on the Affymetrix U95 chip platform, and then subsequently re-analyzed on the newer chip version, U133A, using the same RNA aliquot. CEL files and normalized value in MIAME format from the SFT and STT samples analyzed on the U133A chip are publically available, at below website: http://cbio.mskcc.org/Public/SFT Hierarchical clustering and differentially expressed genes were obtained using Partek® Genomics Suite™ software. The default normalization used was the Robust Multi-chip Average (RMA), Log probeset using base: 2 and ProbesetSummarization:Median Polish. The clustering was performed according to ‘Euclidean metric’ and ‘Average Linkage method’ for Rows and Columns, using a Hierarchical clustering method.
Additionally, we have imported and analyzed the cDNA array data set of 13 SFT samples from West and colleagues [11 (link)], which is publically available from the Stanford Microarray Database. Significance Analysis of Microarray (SAM) was performed on this data set as described by Tusher et al. [12 (link)]. In contrast with the Affymetrix oligonucleotide platform, the cDNA array technology is based on measuring expression between competing test and reference cDNAs. This data-set was used for comparison and validation of the differentially expressed genes obtained from our study, since it investigated a different group of patients, using a different platform and statistical software.
Additionally, we have imported and analyzed the cDNA array data set of 13 SFT samples from West and colleagues [11 (link)], which is publically available from the Stanford Microarray Database. Significance Analysis of Microarray (SAM) was performed on this data set as described by Tusher et al. [12 (link)]. In contrast with the Affymetrix oligonucleotide platform, the cDNA array technology is based on measuring expression between competing test and reference cDNAs. This data-set was used for comparison and validation of the differentially expressed genes obtained from our study, since it investigated a different group of patients, using a different platform and statistical software.
