In the last years new methods for molecular diagnosis gave us the possibility to detect circulating tumor cells (CTCs) in peripheral blood.The liquid biopsy is a modern uninvasive approach really promising. CTC molecular characterization offers the great potential to better understand the biology of metastasis and how and when they develope resistance to targeted therapies. This analysis could help us to follow our patients over time and, mostly important, identify them in an early-stage of reccurence, earlier than metastasis can be assessed by strumental evaluations. The main problem linked to their detection is that CTCs are very hard to find because they're rare, and the amount of available sample is limited (this is mainly due to extraction and conservation process). According to recent preclinical findings already reported in literature, the aim of our work is to dose levels of mutated CTCs in plasma of patients suffering from cancer of different site and, if it's possible, look for the overexpression of some inflammatory biomarker already used in the clinical practice.The primary objective is evaluate the existing agreement between mutational status of primary tumor and blood mutated CTCs levels; try to see if there's some relationship between their levels and the clinical outcome. The second endpoint is find a correlation with clinical response parameters such as TTP (time to progression) or RFS (relapse free survival).
This is an open multicenter study. From 1st May 2015 we enrolled patients (accrual is 50) suffering from melanoma, colorectal cancer and lung. Each patient was analysed for BRAF, KRAS, EGFR mutational status respectively, performed on primary or metastatic lesion according to international guidelines for clinical practice. We took a whole blood sample (5ml) from each patient at baseline, after each administration of chemotherapy and every month of follow up. Samples were used for the collection of the plasma, obtained by two following centrifugations (2500g x 10 min at 4Â°C). The recovered plasma was then divided into 1mL vials and stored at -80 Â°C up to the subsequent extraction of CTCs.The extraction of circulating DNA was carried out manually using a commercial kit, whilst the assessment of the DNA obtained was carried out by Real Time PCR.
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All authors have declared no conflicts of interest.