Research Article: Whole-Genome Gene Expression Profiling of Formalin-Fixed, Paraffin-Embedded Tissue Samples

Date Published: December 3, 2009

Publisher: Public Library of Science

Author(s): Craig April, Brandy Klotzle, Thomas Royce, Eliza Wickham-Garcia, Tanya Boyaniwsky, John Izzo, Donald Cox, Wendell Jones, Renee Rubio, Kristina Holton, Ursula Matulonis, John Quackenbush, Jian-Bing Fan, Patrick Tan. http://doi.org/10.1371/journal.pone.0008162

Abstract: We have developed a gene expression assay (Whole-Genome DASL®), capable of generating whole-genome gene expression profiles from degraded samples such as formalin-fixed, paraffin-embedded (FFPE) specimens.

Partial Text: Formalin-fixed, paraffin-embedded (FFPE) tissues represent an invaluable resource for cancer research, as they are the most widely available material for which patient outcomes are known. There were over 300 million archived cancer tissue samples in the United States in 1999, with more samples accumulating at a rate of over 20 million per year [1]. The ability to perform gene expression profiling in these samples will enable both prospective and retrospective studies, and should greatly facilitate research in correlating expression profiles with clinical outcomes [2]. However, formalin fixation is known to render adenosine residues particularly prone to chemical modifications such as methylene dimerization and mono-methylolation [3] and generate degraded RNA fragments (up to 50% of which may not contain a poly-A tract) [4]. The degradation and chemical modification of RNA during tissue fixation and storage present challenges when applying conventional microarray technologies [3], [5]–[8].

Over the last two decades, significant strides have been made in RNA profiling in FFPE tissues, including efforts to standardize tissue handling and fixation procedures [24], [25] and improving RNA extraction methodologies for FFPE tissues [13], [26], [27]. Despite these advancements, very few technologies have emerged that are capable of robust whole transcriptome profiling in archived FFPE materials. Initial attempts at large-scale expression profiling in FFPE tissues yielded either poor reproducibility and sensitivity [28] or loss of gene signature information when compared with matched FF samples [29]. Most recently, several commercial and academic endeavors have focused on improving the efficiency of amplification and labeling of FFPE-derived RNAs and have met with varying degrees of success [5]–[8], [30]–[34]. While in some instances these technologies have demonstrated their ability to retrieve meaningful biological information from degraded samples [27], [35], concerns remain centered largely around the capability with which older archived tissues (>10 years) can be routinely profiled [29], [31] as well as reports of low signal intensities and poor (transcript) detection sensitivity in FFPE tissues [5], [7], [36].

Source:

http://doi.org/10.1371/journal.pone.0008162

 

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