MicroRNAs are small RNAs that regulate gene expression at the post-transcriptional level. miRNAs exert an important role in the regulation of biological processes, and abnormal expression of miRNAs are associated with different cancers. Considering the strong association between genetic alterations and neoplastic diseases, it is not surprising that there is a special focus on the correlation between miRNAs and cancer. Several experimental studies on colorectal cancer, the most common cancer site and furthermore the second most common cause of death due to cancer, have highlighted the critical role of miRNA in regulation of wellknown oncogenic and tumour suppressor signalling pathways. Furthermore, several investigations have described the ability of microRNA expression patterns to predict prognosis in colon cancer and support diagnosis of poorly differentiated tumours. Currently, two different approaches are applied to investigate the association between miRNA and colorectal cancer (CRC). On the one hand, miRNAs seem to regulate many known oncogenic and tumour suppressor pathways involved in the pathogenesis of CRC. This is of particular interest in colorectal neoplasms as many proteins involved in key signalling pathways in this tumour, like p53, RAS and epithelial–mesenchymal transition (EMT) transcription factors as well as members of the PI-3-K and the Wnt/β-catenin pathway seem to be affected by miRNA regulation. Their dissection in functional studies is critical for a better understanding of cancer biology, eventually aiming for the identification of novel pharmaceutical targets. On the other hand, expression profiles of hundreds of different miRNAs have been shown to bear a much higher potential as biomarkers than their mRNA counterparts. This allows a prediction of prognosis and a distinction of certain disease entities including colorectal cancer sub-types. In order to investigate miRNA differential expression in human colorectal cancer, we analyzed by microarray and real-time PCR the expression of 939 mature miRNAs in total RNA extracted from 39 rectal fresh frozen biopsies, with the aim to found a “signature” that correlates with the response to treatments. Total RNA was extracted from biopsies of patients affected by a locally advanced rectal cancer, treated with neo-adjuant chemo-radiotherapy, follwing by surgery. RNA concentration was tested by Nanodrop ND 1000 spectrophotometer and by Agilent Bioanalyzer Assay to measure the total RNA integrity (RIN) and concentration. Also small RNA was tested by Small RNA Kit to characterize the fraction of miRNAs. The expression level of mature miRNAs found their biopsy specimen, obtained prior to the combination therapy, were quantified using microarray analysis. The miRNA expression pattern was classified based on the final pathological response to the combination therapy. We found that the expression level of 14 miRNAs were significantly different in patients with TRG 1 if compared with patients with other TRG, with a sensitivity of 78% and a specificity of 80%. The miRNA expression pattern is associated with therapeutic outcome and may be a specific “signature”. This information could be utilized as a novel biomarker to predict drug response. La candidata ha analizzato l'espressione di una serie di microRNA in campioni di tessuto derivati da neoplasie del retto localmente avanzate, trattate con radio-chemioterapia concomitante a scopo adiuvante. Un cluster di 14 microRNA erano sovraespressi nei pazienti che ottenevano una risposta completa patologica (Tumor Regression Grade 1) dopo il trattamento neoadiuvante (sensibilità 78%; specificità 80%)
TESI DI DOTTORATO: New biomolecular prognostic and predictive factors of response to treatments in colo-rectal cancer. CANDIDATO: G. Della Vittoria Scarpati. RELATORE: S.Pepe
PEPE, Stefano
2010-01-01
Abstract
MicroRNAs are small RNAs that regulate gene expression at the post-transcriptional level. miRNAs exert an important role in the regulation of biological processes, and abnormal expression of miRNAs are associated with different cancers. Considering the strong association between genetic alterations and neoplastic diseases, it is not surprising that there is a special focus on the correlation between miRNAs and cancer. Several experimental studies on colorectal cancer, the most common cancer site and furthermore the second most common cause of death due to cancer, have highlighted the critical role of miRNA in regulation of wellknown oncogenic and tumour suppressor signalling pathways. Furthermore, several investigations have described the ability of microRNA expression patterns to predict prognosis in colon cancer and support diagnosis of poorly differentiated tumours. Currently, two different approaches are applied to investigate the association between miRNA and colorectal cancer (CRC). On the one hand, miRNAs seem to regulate many known oncogenic and tumour suppressor pathways involved in the pathogenesis of CRC. This is of particular interest in colorectal neoplasms as many proteins involved in key signalling pathways in this tumour, like p53, RAS and epithelial–mesenchymal transition (EMT) transcription factors as well as members of the PI-3-K and the Wnt/β-catenin pathway seem to be affected by miRNA regulation. Their dissection in functional studies is critical for a better understanding of cancer biology, eventually aiming for the identification of novel pharmaceutical targets. On the other hand, expression profiles of hundreds of different miRNAs have been shown to bear a much higher potential as biomarkers than their mRNA counterparts. This allows a prediction of prognosis and a distinction of certain disease entities including colorectal cancer sub-types. In order to investigate miRNA differential expression in human colorectal cancer, we analyzed by microarray and real-time PCR the expression of 939 mature miRNAs in total RNA extracted from 39 rectal fresh frozen biopsies, with the aim to found a “signature” that correlates with the response to treatments. Total RNA was extracted from biopsies of patients affected by a locally advanced rectal cancer, treated with neo-adjuant chemo-radiotherapy, follwing by surgery. RNA concentration was tested by Nanodrop ND 1000 spectrophotometer and by Agilent Bioanalyzer Assay to measure the total RNA integrity (RIN) and concentration. Also small RNA was tested by Small RNA Kit to characterize the fraction of miRNAs. The expression level of mature miRNAs found their biopsy specimen, obtained prior to the combination therapy, were quantified using microarray analysis. The miRNA expression pattern was classified based on the final pathological response to the combination therapy. We found that the expression level of 14 miRNAs were significantly different in patients with TRG 1 if compared with patients with other TRG, with a sensitivity of 78% and a specificity of 80%. The miRNA expression pattern is associated with therapeutic outcome and may be a specific “signature”. This information could be utilized as a novel biomarker to predict drug response. La candidata ha analizzato l'espressione di una serie di microRNA in campioni di tessuto derivati da neoplasie del retto localmente avanzate, trattate con radio-chemioterapia concomitante a scopo adiuvante. Un cluster di 14 microRNA erano sovraespressi nei pazienti che ottenevano una risposta completa patologica (Tumor Regression Grade 1) dopo il trattamento neoadiuvante (sensibilità 78%; specificità 80%)I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
