Transcriptional (mRNA) Profiling in Clear Cell Renal Cell Carcinoma (CCRCC): Comparison of CCRCC to Non-Neoplastic Renal Tissue
Sheldon I Bastacky, Melissa L Bastacky, Somak Roy, Anil V Parwani, Rajiv Dhir, Maureen A Lyons-Weiler, Christin M Sciulli, Michael Krill-Burger, Milon Amin, William A LaFramboise. University of Pittsburgh Medical Center, Pittsburgh, PA
Background: CCRCC is the most common primary renal tumor, with a characteristic morphology and immunophenotype. Prior studies have found consistent chromosomal mutations, e.g. 3p deletions, and dysregulation of hypoxic inducible factor-1 (HIF-1) leading to transcription of HIF-1 inducible genes. The study aim was to characterize the mRNA expression profile of CCRCC using a high density mRNA array to identify a transcript signature and important cancer-related transcripts in CCRCC.
Design: RNA was purified from macrodissected specimens (nl=6, sporadic CCRCC=5) using the Qiagen miRNeasy kit (Valencia, CA) with spectrophotometric absorption: 260/280 >1.8 (NanoDrop, Wilmington, DE) and RIN value >8.0 (Bioanalyzer 2100, Agilent Technologies, Santa Clara, CA). mRNA (500ng) underwent cDNA synthesis and in vitro transcription using the Ambion WT Expression assay (Ambion Inc, Austin, TX) followed by fragmentation and hybridization on Human Exon 1.0 ST arrays (Affymetrix Corp.,Santa Clara, CA). Washing, staining and scanning of arrays was performed (Fluidics Station 450, Scanner 3000) after hybridization and signal intensity calculated by Microarray Suite version 5.0. Statistical comparisons (ANOVA) were performed (Partek Genomics Suite, St. Louis, MO) with false discovery rate: q value=0.05 and -2.0>fold-change>2.0.
Results: 415 CCRCC transcripts were significantly increased (n=191; 2 to 22x) or decreased (n=224; -2 to -92x)). 137 transcripts involved cancer-related genes. Altered tumor suppressor gene (n=14) and oncogene (n=10) transcripts were seen, with a partial list with greatest fold changes including: EGF variant(-17.5x), SCL481(-10x), MAL(-9.5x), GPC3(-7x), PTGS2(-6.5x), RARB(-2.3x) and CA-IX(10x), IGFBP3(6x), C3(4.6x), VEGF-A(4.6x), MDM2(2.5x). HIF-1α/1β, and VHL transcripts were not significantly altered. 21 transcripts were found in 13 different cancer signaling pathways (e.g. pancreas, bladder, ovary, lung (non-small cell and small cell), melanoma, breast, thyroid, other) with a mean+SD (range) of transcripts per pathway of 4.7+2.3 (1-9) (Ingenuity database).
Conclusions: CCRCC showed many mRNA transcript alterations, including increases and decreases of important oncogene and tumor suppressor transcripts. A subset of these transcript alterations is shared amongst a diverse group of other organ cancer signaling pathways, suggesting that CCRCC involves dysfunction of multiple growth regulatory pathways. The absence of significant HIF-1 transcript alterations is consistent with the understanding that HIF-1 is dysregulated at the protein level.
Category: Genitourinary (including renal tumors)
Tuesday, March 5, 2013 9:30 AM
Poster Session III # 90, Tuesday Morning