Analysis of MicroRNAs in Pancreatic Fine-Needle Aspirates Can Classify Benign and Malignant Tissues

Abstract

BACKGROUND MicroRNAs (miRNAs) are RNA molecules that are involved in the regulation of many cellular processes, including those related to human cancers. The aim of this study was to determine, as a proof of principle, whether specific candidate miRNAs could be detected in fine-needle aspirate (FNA) biopsies of pancreatic ductal adenocarcinoma (PDAC) and could accurately differentiate malignant from benign pancreatic tissues. METHODS We used TaqMan® assays to quantify miRNA levels in FNA samples collected in RNARetain (n = 16) and compared the results with a training set consisting of frozen macrodissected pancreatic samples (n = 20). RESULTS Quantitative reverse-transcription PCR analysis confirmed that miRNA levels are affected in PDAC FNAs and correlate well with the changes observed in the training set of frozen pancreatic samples. Analysis of the amounts produced for a few specific miRNAs enabled identification of PDAC samples. The combination of miR-196a and miR-217 biomarkers further improved the ability to distinguish between healthy tissue, PDAC, and chronic pancreatitis in the training set (P = 8.2 × 10−10), as well as segregate PDAC FNA samples from other FNA samples (P = 1.1 × 10−5). Furthermore, we showed that miR-196a production is likely specific to PDAC cells and that its incidence paralleled the progression of PDAC. CONCLUSIONS To the best of our knowledge, this study is the first to evaluate the diagnostic potential of miRNAs in a clinical setting and has shown that miRNA analysis of pancreatic FNA biopsy samples can aid in the pathologic evaluation of suspicious cases and may provide a new strategy for improving the diagnosis of pancreatic diseases.

MicroRNA expression profiles associated with pancreatic adenocarcinoma and ampullary adenocarcinoma

“…Nine of the significantly differentially expressed miRNAs described by Bloomston et al 20 were also found in our study (miR-21, miR-100*, miR-143/miR-143*, miR-148a, miR-148a*, miR-205, miR-210, miR-222 and miR-375). Eleven of the significantly differentially expressed miRNAs described by Szafranska et al 21,26 were found in our study with very similar dm-values and fold change (miR-31, miR-130b, miR-143/miR-143*, miR-148a, miR-196b, miR-205, miR-210, miR-216, miR-217, miR-222/miR-222* and miR-375).…”

“…All miRNAs either up-or down-regulated compared with normal pancreas in other studies 20,21,26 were also significantly differentially expressed in our study before the strict Bonferonni correction for multiple comparisons, except miR-155 that was not present in our microRNA array (Table 2).…”

“…Some selected microRNAs (selection based on P-values, dm-values, fold change and novelty) and comparison with other studies are shown in Table 3. MiR-148a was also found to be significantly down-regulated by Bloomston et al, 20 Szafranska et al 21,26 also found that miR-148a, miR-196b, miR-196a and miR-205 were differentially expressed in pancreatic adenocarcinoma and chronic pancreatitis (Table 3).…”

“…Fourteen of the 19 microRNAs in this classifier are not included in earlier reports of microRNA profiles in pancreatic cancer tissue (miR-34c-5p, miR-122, miR-135b, P-values are Bonferroni corrected for multiple testing. P-values, dm and fold change for the microRNAs that are also found significantly differently expressed by Bloomston et al 20 and Szafranska et al 21,26 are listed. a The difference of means (dm) column corresponds to the log2 fold change and is the difference between class means (1st tissue À 2nd tissue mean C t -values).…”

“…[15][16][17][18][19] In pancreatic adenocarcinomas, the expression of miR-15b, miR-21, miR-95, miR-103, miR-107, miR-148a, miR-155, miR-196a, miR-200, miR-210, miR-217, miR-221, miR-222 and miR-375 are different from tissue of normal pancreas and chronic pancreatitis. 17,[20][21][22][23][24][25][26][27] The results reported by Bloomston et al 20 and Szafranska et al 26 give promises of significant clinical impact of microRNA expression profiles to separate tissue from pancreatic adenocarcinomas from normal pancreas and chronic pancreatitis. The study by Szafranska et al 26 included a combination of two microRNAs (the difference between miR-196a and miR-217) to separate pancreatic adenocarcinomas and chronic pancreatitis (ASURAGEN-test).…”