Bilge Aktas1, Hongxia Sun2, Hui Yao3, Weiwei Shi1, Rebekah Hubbard2, Ya Zhang2, Tingting Jiang1, Sophia N Ononye1, Vikram B Wali1, Lajos Pusztai1, W Fraser Symmans2, Christos Hatzis4. 1. Section of Medical Oncology, Yale Cancer Center, Yale School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA. 2. Department of Pathology, UT M.D. Anderson Cancer Center, P.O. Box 85, 1515 Holcombe Blvd, Houston, TX 77030, USA. 3. Department of Bioinformatics and Computational Biology, UT M.D. Anderson Cancer Center, P.O. Box 301402, Houston, TX 77230, USA. 4. Section of Medical Oncology, Yale Cancer Center, Yale School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA. Electronic address: christos.hatzis@yale.edu.
Abstract
BACKGROUND: Tissue handling can alter global gene expression potentially affecting the analytical performance of genomic signatures, but such effects have not been systematically evaluated. METHODS: Tissue samples from 11 previously untreated breast tumors were minced and aliquots were either snap frozen or placed in RNAlater immediately or after 20, 40, 60, 120 or 180 min at room temperature. RNA was profiled on Affymetrix HG-U133A arrays. We used probe-set-wise hierarchical models to evaluate the effect of preservation method on transcript expression and linear mixed effects models to assess the effect of cold ischemic delay on the expression of individual probe sets. Gene set enrichment analysis identified pathways overrepresented in the affected transcripts. We combined the levels of 41 most sensitive transcripts to develop an index of ischemic stress. RESULTS: Concordance in global gene expression between the baseline and 40 min delay was higher for samples preserved in RNAlater (average concordance correlation coefficient CCC = 0.92 compared to 0.88 for snap frozen). Overall, 481 transcripts (3%) were significantly affected by the preservation method, most of them involved in processes important in cancer. Prolonged cold ischemic delay of up to 3 h induced marginal global gene expression changes (average CCC = 0.90 between baseline and 3 h delay). However 41 transcripts were significantly affected by cold ischemic delay. Among the induced transcripts were stress response genes, apoptotic response genes; among the downregulated were genes involved in metabolism, protein processing and cell cycle regulation. An index combining the expression levels of these genes was proportional to the cold ischemic delay. CONCLUSIONS: Prolonged cold ischemia induces significant transcriptional changes in a small subset of transcripts in the tissue. Furthermore, the expression level of about 3% of the transcripts is affected by the preservation method. These sensitive transcripts should not be included in genomic signatures for more reliable analytical performance.
BACKGROUND: Tissue handling can alter global gene expression potentially affecting the analytical performance of genomic signatures, but such effects have not been systematically evaluated. METHODS: Tissue samples from 11 previously untreated breast tumors were minced and aliquots were either snap frozen or placed in RNAlater immediately or after 20, 40, 60, 120 or 180 min at room temperature. RNA was profiled on Affymetrix HG-U133A arrays. We used probe-set-wise hierarchical models to evaluate the effect of preservation method on transcript expression and linear mixed effects models to assess the effect of cold ischemic delay on the expression of individual probe sets. Gene set enrichment analysis identified pathways overrepresented in the affected transcripts. We combined the levels of 41 most sensitive transcripts to develop an index of ischemic stress. RESULTS: Concordance in global gene expression between the baseline and 40 min delay was higher for samples preserved in RNAlater (average concordance correlation coefficient CCC = 0.92 compared to 0.88 for snap frozen). Overall, 481 transcripts (3%) were significantly affected by the preservation method, most of them involved in processes important in cancer. Prolonged cold ischemic delay of up to 3 h induced marginal global gene expression changes (average CCC = 0.90 between baseline and 3 h delay). However 41 transcripts were significantly affected by cold ischemic delay. Among the induced transcripts were stress response genes, apoptotic response genes; among the downregulated were genes involved in metabolism, protein processing and cell cycle regulation. An index combining the expression levels of these genes was proportional to the cold ischemic delay. CONCLUSIONS: Prolonged cold ischemia induces significant transcriptional changes in a small subset of transcripts in the tissue. Furthermore, the expression level of about 3% of the transcripts is affected by the preservation method. These sensitive transcripts should not be included in genomic signatures for more reliable analytical performance.
Authors: Eldrid Borgan; Roy Navon; Hans Kristian Moen Vollan; Ellen Schlichting; Torill Sauer; Zohar Yakhini; Ole Christian Lingjærde; Therese Sørlie; Anne-Lise Børresen-Dale Journal: Mol Oncol Date: 2011-08-29 Impact factor: 6.603
Authors: Christos Sotiriou; Pratyaksha Wirapati; Sherene Loi; Adrian Harris; Steve Fox; Johanna Smeds; Hans Nordgren; Pierre Farmer; Viviane Praz; Benjamin Haibe-Kains; Christine Desmedt; Denis Larsimont; Fatima Cardoso; Hans Peterse; Dimitry Nuyten; Marc Buyse; Marc J Van de Vijver; Jonas Bergh; Martine Piccart; Mauro Delorenzi Journal: J Natl Cancer Inst Date: 2006-02-15 Impact factor: 13.506
Authors: Jubao Duan; Jianxin Shi; Xijin Ge; Lars Dölken; Winton Moy; Deli He; Sandra Shi; Alan R Sanders; Jeff Ross; Pablo V Gejman Journal: Sci Rep Date: 2013 Impact factor: 4.379
Authors: Ligia Craciun; Selim Alex Spinette; Marc Rassy; Roberto Salgado; Alexandre de Wind; Pieter Demetter; Laurine Verset; Maria Gomez-Galdon; Marie Chintinne; Nicolas Sirtaine; Nicolas de St Aubain; Ioanna Laios; Francoise Roy; Denis Larsimont Journal: Front Med (Lausanne) Date: 2019-10-17
Authors: Elena López-Knowles; Qiong Gao; Maggie Chon U Cheang; James Morden; Joel Parker; Lesley-Ann Martin; Isabel Pinhel; Fiona McNeill; Margaret Hills; Simone Detre; Maria Afentakis; Lila Zabaglo; Andrew Dodson; Anthony Skene; Chris Holcombe; John Robertson; Ian Smith; Judith M Bliss; Mitch Dowsett Journal: Breast Cancer Res Date: 2016-04-01 Impact factor: 6.466
Authors: Dominic A Pearce; Laura M Arthur; Arran K Turnbull; Lorna Renshaw; Vicky S Sabine; Jeremy S Thomas; John M S Bartlett; J Michael Dixon; Andrew H Sims Journal: Sci Rep Date: 2016-07-07 Impact factor: 4.379