Enhanced stability of microRNA expression facilitates classification of FFPE tumour samples exhibiting near total mRNA degradation

Abstract

Background:As degradation of formalin-fixed paraffin-embedded (FFPE) samples limits the ability to profile mRNA expression, we explored factors predicting the success of mRNA expression profiling of FFPE material and investigated an approach to overcome the limitation.Methods:Bladder (n=140, stored 3–8 years) and cervix (n=160, stored 8–23 years) carcinoma FFPE samples were hybridised to Affymetrix Exon 1.0ST arrays. Percentage detection above background (%DABG) measured technical success. Biological signal was assessed by distinguishing cervix squamous cell carcinoma (SCC) and adenocarcinoma (AC) using a gene signature. As miR-205 had been identified as a marker of SCC, precursor mir-205 was measured by Exon array and mature miR-205 by qRT–PCR. Genome-wide microRNA (miRNA) expression (Affymetrix miRNA v2.0 arrays) was compared in eight newer FFPE samples with biological signal and eight older samples without.Results:RNA quality controls (QCs) (e.g., RNA integrity (RIN) number) failed to predict profiling success, but sample age correlated with %DABG in bladder (R=−0.30, P<0.01) and cervix (R=−0.69, P<0.01). Biological signal was lost in older samples and neither a signature nor precursor mir-205 separated samples by histology. miR-205 qRT–PCR discriminated SCC from AC, validated by miRNA profiling (26-fold higher in SCC; P=1.10 × 10−5). Genome-wide miRNA (R=0.95) and small nucleolar RNA (R=0.97) expression correlated well in the eight newer vs older FFPE samples and better than mRNA expression (R=0.72).Conclusion:Sample age is the best predictor of successful mRNA profiling of FFPE material, and miRNA profiling overcomes the limitation of age and copes well with older samples.

miQ—A novel microRNA based diagnostic and prognostic tool for prostate cancer

“…miRNAs have been found to be very stable in FFPE tissue 18,19 and several studies have compared the miRNA expression in FFPE and fresh frozen sections and found it to have very high concordance. [20][21][22] In line with this we found most miRNAs to display similar expression patterns as seen in the discovery cohort constituting fresh frozen material.…”

miR-194 as a Predictor for Adenoma Recurrence in Patients with Advanced Colorectal Adenoma after Polypectomy

“…The detection of FFPE-based miRNAs has several distinct advantages: (i) miRNA expression is very stable in FFPE blocks (28), and has very high concordance in FFPE and fresh-frozen preserved samples (29); (ii) FFPE tissues are the most accessible biologic resources and are easily stored; (iii) the diagnosis of ACRA is established on hematoxylin and eosin-stained slides, which are only sourced from FFPE specimens.…”

Combined Label-Free Quantitative Proteomics and microRNA Expression Analysis of Breast Cancer Unravel Molecular Differences with Clinical Implications

“…In recent years, we and others have demonstrated that formalinfixed, paraffin-embedded (FFPE) tissue can be readily analyzable by MS-based technologies (6,7). On the other hand, previous studies demonstrate the possibility of performing experiments to analyze the expression of miRNAs in FFPE tissues (8). FFPE samples also constitute the basis for routine workup in pathology departments all over the world.…”