ORIGINAL ARTICLE High miR-96 levels in colorectal adenocarcinoma predict poor prognosis, particularly in patients without distant metastasis at the time of initial diagnosis Stamatia-Maria Rapti 1 & Christos K. Kontos 1 & Iordanis N. Papadopoulos 2 & Andreas Scorilas 1 Received: 27 November 2015 /Accepted: 18 March 2016 # International Society of Oncology and BioMarkers (ISOBM) 2016 Abstract MicroRNA-96 (miR-96) is an oncomiR that facili- tates the development of malignant tumors by promoting growth, proliferation, and survival of cancer cells. Previous studies using high-throughput techniques have shown that miR-96 is upregulated in colorectal cancer compared to adja- cent normal colorectal tissue. The aim of this study was the investigation of the potential clinical value of miR-96 as a molecular prognostic biomarker in colorectal adenocarcino- ma. For this purpose, total RNA was extracted from 108 pri- mary colorectal adenocarcinoma samples and 54 paired non- cancerous colorectal tissue specimens. After polyadenylation and reverse transcription, miR-96 molecules were determined using an in-house developed real-time quantitative PCR based on SYBR Green chemistry. Calculations were carried out with the comparative C T method, using SNORD48 as endogenous reference gene. Finally, extensive biostatistical analysis was performed and showed that miR-96 is significantly upregulat- ed in colorectal adenocarcinoma specimens compared to their non-cancerous counterparts (p <0.001) as well as in tumors having invaded regional lymph nodes (p = 0.009) and those of advanced TNM stage (p = 0.008). miR-96 expression is an unfavorable prognostic marker in colorectal adenocarcinoma, predicting poor disease-free and overall survival (p = 0.041 and 0.028, respectively), independently of classical clinico- pathological parameters. Most importantly, miR-96 expres- sion stratifies patients without distant metastasis (M0) at the time of diagnosis into two groups with substantially different prognosis (p = 0.040). In conclusion, high tissue levels of miR-96 are associated with advanced stages of colorectal ad- enocarcinoma and predict an increased risk for disease recur- rence and poor overall survival, especially in patients without distant metastasis at the time of diagnosis. Keywords MicroRNAs (miRNAs) . Molecular tumor markers . Prognostic biomarkers . Colorectal cancer . Real-time quantitative PCR (qPCR) Introduction Colorectal cancer (CRC) has a yearly incidence of about 1.26 million patients all over the world, thus comprising around 9.4 % of all cancer cases, according to the WHO 2008 report [1]. CRC, the third most commonly diagnosed cancer in both men and women, is more frequent in the Western world. In Europe and the USA, CRC constitutes the second most fre- quent cancer-related cause of death. In 2008, 608 thousands of patients died of this malignancy [2]. More than 95 % of CRCs are carcinomas and about 95 % of these are adenocarcinomas [3]. Although heritability contributes to CRC risk, the vast majority of 75 % of CRC patients have sporadic disease [4]. A number of acquired molecular events are associated with the transition from normal colonic epithelium to adenoma and, finally, to adenocarcinoma [5]. CRC is highly curable as the pathological tissue can be surgically removed, provided that the disease is detected early enough. However, CRC is often diagnosed at an advanced stage, when the tumor burden has already spread, thus giving Electronic supplementary material The online version of this article (doi:10.1007/s13277-016-5023-0) contains supplementary material, which is available to authorized users. * Andreas Scorilas ascorilas@biol.uoa.gr 1 Department of Biochemistry and Molecular Biology, University of Athens, Panepistimiopolis, Athens 15701, Greece 2 Fourth Surgery Department, University General Hospital BAttikon^, Haidari, Athens 12462, Greece Tumor Biol. DOI 10.1007/s13277-016-5023-0