Abstract
BackgroundNeuroendocrine lung cancer (NELC) represents 25% of all lung cancer cases and large patient collectives exist as formalin-fixed, paraffin-embedded (FFPE) tissue only. FFPE is controversially discussed as source for molecular biological analyses and reference genes for NELC are poorly establishes.Material and methodsForty-three representative FFPE-specimens were used for mRNA expression analysis using the digital nCounter technology (NanoString). Based on recent literature, a total of 91 mRNA targets were investigated as potential tumor markers or reference genes. The geNorm, NormFinder algorithms and coefficient of correlation were used to identify the most stable reference genes. Statistical analysis was performed by using the R programming environment (version 3.1.1)ResultsRNA integrity (RIN) ranged from 1.8 to 2.6 and concentrations from 34 to 2,109 ng/μl. However, the nCounter technology gave evaluable results for all samples tested. ACTB, CDKN1B, GAPDH, GRB2, RHOA and SDCBP were identified as constantly expressed genes with high stability (M-)values according to geNorm, NormFinder and coefficients of correlation.ConclusionFFPE-derived mRNA is suitable for molecular biological investigations via the nCounter technology, although it is highly degraded. ACTB, CDKN1B, GAPDH, GRB2, RHOA and SDCBP are potent reference genes in neuroendocrine tumors of the lung.
Highlights
Neuroendocrine lung cancer (NELC) represents 25% of all lung cancer cases and is divided into four subgroups (typical (TC) and atypical carcinoids (AC), large- (LCNEC) and small-cell cancer (SCLC)) [1].The majority of these clinical samples are stored as formalin-fixed, paraffin-embedded (FFPE) tissue
ACTB, CDKN1B, GAPDH, GRB2, RHOA and SDCBP were identified as constantly expressed genes with high stability (M-)values according to geNorm, NormFinder and coefficients of correlation
FFPE-derived mRNA is suitable for molecular biological investigations via the nCounter technology, it is highly degraded
Summary
Neuroendocrine lung cancer (NELC) represents 25% of all lung cancer cases and is divided into four subgroups (typical (TC) and atypical carcinoids (AC), large- (LCNEC) and small-cell (neuroendocrine) cancer (SCLC)) [1]. The majority of these clinical samples are stored as formalin-fixed, paraffin-embedded (FFPE) tissue. The technology enables the detection of short target regions (~100 nt) making it an ideal method for analyzing fragmented nucleic acids from FFPE tissue. Neuroendocrine lung cancer (NELC) represents 25% of all lung cancer cases and large patient collectives exist as formalin-fixed, paraffin-embedded (FFPE) tissue only. FFPE is controversially discussed as source for molecular biological analyses and reference genes for NELC are poorly establishes
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