|Tipo di tesi||Tesi di dottorato di ricerca|
|Titolo||Caratteristiche molecolari e stato di metilazione del cancro colorettale ad insorgenza precoce (≤ 40 anni di età)|
|Titolo in inglese||Molecular features and methylation status in early onset (≤40yr) colorectal cancer|
|Settore scientifico disciplinare||MED/06 - ONCOLOGIA MEDICA|
|Corso di studi||Scuola di D.R. in MEDICINA CLINICA E SPERIMENTALE (CLINICAL AND EXPERIMENTAL MEDICINE)|
|Data inizio appello||2014-02-18|
|Disponibilità||Accessibile via web (tutti i file della tesi sono accessibili)|
Introduzione: Pressochè sconosciuta fino a 3 o 4 decenni fa, la genetica del cancro ha portato a rilevanti avanzamenti della nostra conoscenza sullo sviluppo tumorale. Molte sindromi sono state identificate seguendo la legge dell’ereditarietà Mendeliana per cui la patologia era trasmessa da una generazione all’altra.
Background: Almost unknown until 3 to 4 decades ago, cancer genetics led to the most relevant advancements of our understanding in tumor development. Many syndromes have been identified in which the disease is transmitted from generation to generation following Mendelian inheritance. Colorectal cancer (CRC) is usually considered a disease of the elderly in both sexes and increases with advancing age. So the mean age of affected individuals is around 70 years. However a small fraction of patients develops colorectal cancer earlier. The reasons whereby some individuals develop CRC at an unusual age are poorly understood. Aim: Despite the clinical suspicion of a genetic origin (Lynch Syndrome and Familial Adenomatous Polyposis), the purpose of this study is to compare biomolecular features and methylation status of early onset CRC with those from tumors of elderly patients in order to elucidate possible pathogenetic mechanisms that onset in young individuals. Methods: In the study we included 33 patients with disease ≤40 years of onset and a control group composed of 41 patients with disease at ≥60 age, matched for the three characteristics of preponderance of male sex, C and D stages of Dukes and site of tumor especially in distal colon. All tumors were investigated for Microsatellite Instability (MSI) test and Mismatch Repair (MMR) immunohistochemistry analyses, identification of K-RAS and BRAF somatic mutations. Detection of MMR genes germline mutations was performed only in cases with positive screening tests. APC gene mutations were investigated in patients with a clinical suspect of Familial Polyposis, whereas MUTYH gene was detected in the 33 young cases. Finally, we explored the role of aberrant DNA methylation (hypermethylation and hypomethylation) of all 74 colorectal carcinomas. In particular hypermethylation was investigated by MS-MLPA method using three different kits from MRC-Holland (ME001, ME002, and ME011) and hypomethylation of long interspersed element 1 (LINE-1) was evaluated by bisulfite DNA treatment and pyrosequencing assay. Results: Early onset CRC showed an high incidence of hereditary forms (6 cases, 5 Lynch Syndrome and 1 Familial Adenomatous Polyposis, 18%) and K-RAS somatic mutations (33% versus 7%, p<0.005). BRAF somatic mutations were found only in four elderly CRC with loss of expression of MLH1 protein and MSI. The mean number of methylated genes in early onset CRC was 3.4 whereas in older CRC was 5.22, this difference was statistically significant (p< 0.005). High methylation frequency of ESR1, GATA5, WT1 genes in patients with onset ≤40 years and in control group >60 years. Moreover early onset CRC, excluding Lynch Syndrome cases, showed high hypomethylation frequency of LINE-1 sequences. Conclusion: We have found differences in the genetic make-up of carcinomas from young and elderly patients. Early onset tumors showed constitutional and somatic defective of Mismatch Repair System, more frequent K-RAS somatic mutations and a minor number of methylated genes. Hypermethylation of ESR1, GATA5, WT1 genes suggests possible markers in the earlier diagnosis of colorectal tumorigenesis.