Growing evidence suggests that analysis of circulating tumor cells (CTC) or circulating tumor DNA (ctDNA) can provide a comprehensive real-time picture of the tumor-associated changes in an individual cancer patient and might address both spatial and temporal heterogeneity of cancer. The prognostic and predictive ability of the combination of both biomarkers may yield complementary information and thereby improve diagnostic sensitivity. CTC are shed by a primary tumor into the blood stream of cancer patients, and their number is correlated to prognosis. To date, the CellSearch is the only CTC test cleared by FDA for monitoring of metastatic cancer patients. The system detects CTC from whole blood of cancer patients through an immunomagnetic selection using a suspension of magnetic nanoparticles conjugated to a mouse monoclonal antibody recognizing the EpCAM present on surface of epithelial origin cells. Nevertheless, in the following years several studies have demonstrated that a mixture of EpCAM-positive and EpCAM-negative tumor cells circulate in the blood of cancer patients. Therefore, an unsolved question is the frequency, the molecular characteristics and the clinical relevance of those CTC which do not express EpCAM and are thus currently not detected by the CellSearch.
The use of ctDNA as a clinical response marker for cancer patients has already moved into the clinical routine and mutational analysis from plasma has been proven to complement tissue-based testing in many cancer types.
Aim of this proposal is to compare the genetic information provided by EpCAM-positive, EpCAM-negative and plasma ctDNA from a single blood draw in order to understand the predictive value of each liquid biopsy source in longitudinal monitoring of targeted therapies.
Recent technical advances for molecular diagnosis have made it possible to obtain molecular information on tumors from peripheral blood through liquid biopsy . This approach relies mainly on 2 different sources of circulating genetic information, which are to date the only to have received regulatory approval for clinical use: ctDNA and CTCs. Despite the progress that has taken place in this field, several issues are still unsolved ,being the insufficient evidence of clinical utility for the majority of ctDNA and CTCs assays the most important question to be addressed. In 2004, the U.S. Food and Drug Administration (FDA) approved an epithelial cell adhesion molecule (EpCAM)-dependent technique for use as a prognostic biomarker in breast cancer , which was then expanded for use in prostate and colorectal cancer patients . Since its approval, this EpCAM-dependent device has become the gold standard for CTC isolation . Nevertheless, its utility to guide therapeutic choice has still not been demonstrated, thus strongly limiting its clinical use. The first CTC-guided clinical trial ( SWOG500 ), for instance, clearly demonstrated that a change of treatment based on CTC status did not improve patients outcome.This might reflect the still unsolved methodological issues related to the inadequacy of Cellsearch to address CTC heterogeneity .In fact CellSearch relies on a restricted pool of markers for CTC identification, leading to possible false-negative results . With such an EpCAM-dependent enrichment technique, the marker-negative CTCs are undetectable, creating uncertainty in the accuracy of a patients CTC status. Analysis with Cellsearch is then limited to cancers with CTCs that predominantly maintain epithelial characteristics.The inadequacy of EpCAM as universal marker for CTCs detection seems thus unquestionable and alternative methods able to recognize a broader spectrum of phenotypes are definitely needed to avoid loss of CTCs characterized by EpCAM downregulation and phenotypic plasticity. Recently Terstappen et al.for the first time validated a method of microscopic examination the EpCAM-low CTC collected in the blood discarded by the CellSearch system after immunomagnetic enrichment of EpCAM-high cells. To date the clinical utility of these discarded CTC has not been investigated. A further item to be still addressed is that very limited comparative data are available to date between analysis of ctDNA and CTCs, due to limitations for parallel detection and quantification of different analytes from small volumes of blood. Here we propose a single tube approach from a limited volume of blood enabling the simultaneous capture and analysis of heterogeneous subpopulations of CTCs (EpCAM-high CTC, EpCAM-low CTC) as well as ctDNA isolation as a comprehensive liquid biopsy analysis to monitor response to targeted therapies.
References
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