Conticelli, Floriana (2021) Molecular pathology in colorectal cancer: from early detection to treatment predictions. [Tesi di dottorato]

Preview

Text Conticelli_Floriana_34.pdf Download (1MB) | Preview

Collection description

Background: In the precision medicine era, the increasing request of clinical relevant biomarkers to improve the patients management lead to the need of most biological source. To address this issue, also if tissue represents the gold standard for the assessment of clinical relevant biomarkers mutational status, some alternative approaches, based on the analysis of circulating free DNA (cfDNA) extracted from “liquid biopsies”, are under evaluation. The aim of this thesis was to investigate the role of the molecular pathology in colorectal cancer from the “early detection” to treatment predictions. In particular, it has been investigated the role of the liquid biopsy as a screening tool for Colorectal Cancer (CRC) in comparison of Fetal Immunochemical Test (FIT) but also as a monitoring tool in patients with metastatic colorectal cancer to predict the possibility of relapse. Moreover, in order to the treatment approach for CRC patients, it was evaluated the feasibility to perform the Idylla™ Assay to select the wilde-type K-N-RAS patients to the treatment with the monoclonal antibodies that target and inhibit the EGFR gene. Methods: In relation to CRC patients, employing the analytical validated Real Time PCR-based ColoScape assay Kit, mutations in the APC, KRAS, BRAF and CTNNB1 genes were assessed on 52 prospectively collected whole-blood samples obtained from FIT+ patients enrolled in the CRC screening program of ASL NAPOLI 3 SUD, using colonoscopy as confirmation. It was, also, performed the NGS analysis of the cfDNA samples isolated from mCRC patients and their matching metastatic tissue using the SiRe® panel, which covers 568 mutations in six genes (EGFR, KRAS, NRAS, BRAF, cKIT and PDGFRα) to monitor the possibility of relapse. Moreover, it was assessed the K-NRAS mutational status by using a fully automated Real Time PCR, the Idylla™ System, as first-level screening method to quickly address the CRC patients to the best personalized treatment. Results: The application of Real Time PCR based ColoScape assay Kit as a screening tool for CRC patients, the assay's sensitivity for advanced adenomas was 53.8% and the specificity was 92.3%. The Positive Predictive Value was 70.0% and negative predictive value was 85.7%. Of note, four of the six positive cases missed by ColoScape had a less than suboptimal DNA input. Had they been ruled out as inadequate, sensitivity would have increased from 53.8% to 69%. Regarding the NGS analysis of the cfDNA samples isolated from mCRC patients the molecular analysis was successfully performed in matched tissue specimen for a preliminary set (n=15) of enrolled patients. In n=3 cases, gDNA was not available for molecular analysis. In details, tissue sample analysis showed at least a molecular alteration in n=8/12 (66,6%) cases. In order to evaluate the possibility to use the Idylla™ Assay to address the CRC patients to best target therapy n=450 mCRC patients and a total of 457 samples were tested. In particular, for 5 patients were analyzed also the metastatic tissue and for 2 patients we analyzed two synchronous cancers. A total of n= 200/457 (43,8%) samples gave a WT result, otherwise n=257/457 (56,2%) samples showed a mutation in the clinical relevant genes in patients with colorectal cancer. In particular n=213/257 (82,9%) were KRAS positive, n= 20/257 (7,8%) were NRAS positive and n=24/257 (9,3%) were positive in BRAF gene. Conclusions: In CRC patients setting, ColoScape is a promising tool for screening program aims to evaluate the triage of FIT+ patients and the cfDNA analysis, using the Sire® panel, has proven to be a valid monitoring tool to follow up the mCRC patients. Moreover, the Idylla ™ results combined with the treatment collected data from the oncologists could represent the best strategy to address wild type patients to a targeted therapy. The collected data obtained with this analysis could be useful to improve the treatment approach in the clinical practice setting.

Downloads

Actions (login required)

View Item