A Reliable Method for the Selection of Exploitable Melanoma Archival Paraffin Embedded Tissues for Transcript Biomarker Profiling

Celeste Lebbe,Suzanne Menashi,Anne Janin,Nicole Basset-Seguin,Marie Pierre Podgorniak,Mickael Guedj,Samia Mourah,Rakesh K Srivastava

doi:10.1371/journal.pone.0029143

Abstract

The source tissue for biomarkers mRNA expression profiling of tumors has traditionally been fresh-frozen tissue. The adaptation of formalin-fixed, paraffin-embedded (FFPE) tissues for routine mRNA profiling would however be invaluable in view of their abundance and the clinical information related to them. However, their use in the clinic remains a challenge due to the poor quality of RNA extracted from such tissues. Here, we developed a method for the selection of melanoma archival paraffin-embedded tissues that can be reliably used for transcript biomarker profiling. For that, we used qRT-PCR to conduct a comparative study in matched pairs of frozen and FFPE melanoma tissues of the expression of 25 genes involved in angiogenesis/tumor invasion and 15 housekeeping genes. A classification method was developed that can select the samples with a good frozen/FFPE correlation and identify those that should be discarded on the basis of paraffin data for four reference genes only. We propose therefore a simple and inexpensive assay which improves reliability of mRNA profiling in FFPE samples by allowing the identification and analysis of “good” samples only. This assay which can be extended to other genes would however need validation at the clinical level and on independent tumor series.

Highlights

Malignant melanoma is one of the most rapidly spreading cancers in terms of worldwide incidence [1]
With the enormous amount of data retrievable stored in archived formalin-fixed paraffin-embedded tissue, it will prove invaluable if biomarkers transcript expression levels could be routinely and systematically analyzed in FFPE tissues, for retrospective studies and for the characterization of rare or small tumors
These studies, conducted mainly in breast cancer tissues have shown a strong correlation in transcript expression between paired FFPE and frozen tissues which was independent of tissue fixation time and storage in paraffin

Summary

Introduction

Malignant melanoma is one of the most rapidly spreading cancers in terms of worldwide incidence [1]. With the enormous amount of data retrievable stored in archived formalin-fixed paraffin-embedded tissue, it will prove invaluable if biomarkers transcript expression levels could be routinely and systematically analyzed in FFPE tissues, for retrospective studies and for the characterization of rare or small tumors Their routine use in the clinic has been hampered because of the poor quality of RNA extracted from them. A few emerging studies using qRT-PCR as well as microarrays suggested these FFPE samples can be used to validate biomarker signatures associated with clinical features, survival and therapeutic response [7,8,9,10,11,12] These studies, conducted mainly in breast cancer tissues have shown a strong correlation in transcript expression between paired FFPE and frozen tissues which was independent of tissue fixation time and storage in paraffin

Methods

Results

Conclusion