Prognostic Significance of Gene Expression and DNA Methylation Markers in Circulating Tumor Cells and Paired Plasma Derived Exosomes in Metastatic Castration Resistant Prostate Cancer

Abstract

Liquid biopsy, based on the analysis of circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA), provides non-invasive real-time monitoring of tumor evolution and therapeutic efficacy. We performed for the first time a direct comparison study on gene expression and DNA methylation markers in CTCs and paired plasma-derived exosomes and evaluated their prognostic significance in metastatic castration resistant prostate cancer. This prospective liquid biopsy (LB) study was based on a group of 62 metastatic castration resistant prostate cancer (mCRPC) patients and 10 healthy donors (HD) as controls. Identical blood draws were used to: a) enumerate CTC and tumor-derived extracellular vesicles (tdEVs) using CellSearch (CS) and b) analyze CTCs and paired plasma-derived exosomes at the gene expression and DNA methylation level. CTCs were enumerated using CellSearch in 57/62 patients, with values ranging from 5 to 854 cells/7.5mLPB. Our results revealed for the first time a significantly higher positivity of gene expression markers (CK-8, CK-18, TWIST1, PSMA, AR-FL, AR-V7, AR-567 and PD-L1 mRNA) in EpCAM-positive CTCs compared to plasma-derived exosomes. GSTP1, RASSF1A and SCHLAFEN were methylated both in CTC and exosomes. In CTCs, Kaplan–Meier analysis revealed that CK-19(p = 0.009), PSMA(p = 0.001), TWIST1(p = 0.001) expression and GSTP1(p = 0.001) methylation were correlated with OS, while in exosomes GSTP1 (p = 0.007) and RASSF1A (p = 0.001) methylation was correlated with OS. Our direct comparison study of CTCs and exosomes at gene expression and DNA methylation level, revealed for the first time a significantly higher positivity in EpCAM-positive CTCs compared to plasma-derived exosomes. Future perspective of this study should be the evaluation of clinical utility of molecular biomarkers in CTCs and exosomes on independent multicentric cohorts with mCRPC patients.

Circulating methylated THBS1 DNAs as a novel marker for predicting peritoneal dissemination in gastric cancer

“…The examination of free tumor cells in PPLF in GC patients revealed peritoneal dissemination, predicted peritoneal recurrence and prognosis, and selected proper treatments 7,38 . Currently, cytology and molecular biology are the two most popular methods for the detection of peritoneal dissemination 7–25 …”

“…Promoter methylation of THBS1 gene has been studied in various malignancies, such as colorectal cancer, melanoma, and GC tissues 31–33 . Quantitative MSP technology has high sensitivity and specificity and can be used to detect the methylation levels of tumor‐related genes in liquid biopsy samples of tumor patients for tumor screening, early diagnosis, metastasis warning, recurrence monitoring, and prognostic assessment 18,39 . In this study, we did not find any abnormal methylation of the THBS1 gene in the serum or PPLF DNAs in tumor tissues.…”

“…Recently, methylation‐specific PCR (MSP) technology has been successfully used to analyze methylation markers in liquid biopsy specimens, including PPLF samples for peritoneal metastasis diagnosis 17–19 . Our previous studies suggest that DNA methylation in PPLF or serum may be a potential marker for detecting peritoneal dissemination in GC patients 20–25 …”

Cell-Secreted Vesicles: Novel Opportunities in Cancer Diagnosis, Monitoring and Treatment

“…They highlighted that the concentration of PSMA-positive EVs increased from low to high risk in prostate cancer patients [137]. Moreover, in exosomes, CK-8 expression and the presence of DNA methylation status were significantly correlated with a lower overall survival [138].…”

Dissecting the Hormonal Signaling Landscape in Castration-Resistant Prostate Cancer

“…In humans, the expression of AR-V7 was observed in tumor biopsies from CRPC patients and significantly correlated with tumor progression and short survival [ 88 , 89 , 90 , 91 , 92 ]. High levels of AR-V7 could also be detected in extracellular vesicles purified from the plasma of CRPC patients, as well as in circulating tumor cells (CTCs) from enzalutamide- or abiraterone-resistant patients, and were found to be associated with a poor prognosis [ 93 , 94 , 95 , 96 , 97 ].…”