Liquid Biopsies, Novel Approaches and Future Directions

Cancer is among the leading causes of death worldwide. Early diagnosis and prognosis are vital to improve patients’ outcomes. The gold standard of tumor characterization leading to tumor diagnosis and prognosis is tissue biopsy. Amongst the constraints of tissue biopsy collection is the sampling frequency and the incomplete representation of the entire tumor bulk. Liquid biopsy approaches, including the analysis of circulating tumor cells (CTCs), circulating tumor DNA (ctDNA), circulating miRNAs, and tumor-derived extracellular vesicles (EVs), as well as certain protein signatures that are released in the circulation from primary tumors and their metastatic sites, present a promising and more potent candidate for patient diagnosis and follow up monitoring. The minimally invasive nature of liquid biopsies, allowing frequent collection, can be used in the monitoring of therapy response in real time, allowing the development of novel approaches in the therapeutic management of cancer patients. In this review we will describe recent advances in the field of liquid biopsy markers focusing on their advantages and disadvantages

Determination of mutetions in the BRCA2 gene in greek patients with familial breast cancer

Family history is a well-recognized risk factor for the development of breast cancer. The isolation of BRCA1 and BRCA2 genes, the two major predisposing genes in familial and to early onset breast and ovarian cancer, has resulted to the identification of a large number of families with mutations in these two genes. Despite the large number of distinct mutations, detected in both genes, several mutations have been found to recur in unrelated families of diverse geographical origin. We have analyzed 27 Greek patients with familial breast cancer the majority of those having one first and one second degree relatives affected and 28 patients with sporadic breast cancer for BRCA2 germline mutations. The techniques used were single-strand conformation polymorphism analysis (SSCP) followed by sequencing. Furthermore the clinical presentation and prognosis of BRCA2 associated breast cancer cases was compared to 20 adequately matched for age and date of diagnosis (within one year) sporadic breast cancer patients. We identified three novel (3058delA, 6024delTA and 4147delG) in the ovarian cancer cluster region (OCCR) and one already known (2024del5) germline BRCA2 gene mutations in five different breast cancer families. The 4147delG mutation was detected in two unrelated patients. BRCA2 germline mutations were correlated with early-onset breast cancer RR=4.77 (95% CΙ: 0.666-34.463). Although patients with BRCA2 germline mutations did not have a distinct histological phenotype they had an improved overall survival (100% vs 65%). Our findings suggest that there is a cluster of novel mutations in exons 10 and 11 in Greek patients with familial breast cancer. These mutations appear to have a milder clinical phenotype when compared to the rest of the study group.Το οικογενειακό ιστορικό είναι ένας σημαντικός παράγοντας κινδύνου για την ανάπτυξη του καρκίνου του μαστού. Η απομόνωση των γονιδίων BRCA1 και BRCA2, των δύο κύριων προδιαθεσικών γονιδίων του οικογενούς, του πρώιμου καρκίνου του μαστού, καθώς και του καρκίνου των ωοθηκών, είχε σαν αποτέλεσμα την αναγνώριση μεγάλου αριθμού οικογενειών με μεταλλάξεις στα γονίδια αυτά. Αναλύσαμε 27 Έλληνες ασθενείς με οικογενή καρκίνο του μαστού. Η πλειοψηφία των ασθενών αυτών είχε τουλάχιστον έναν πρώτου και ένα δευτέρου βαθμού συγγενή με τη νόσο για μεταλλάξεις στο γονίδιο BRCA2. Μελετήσαμε ακόμα 28 ασθενείς με σποραδικό καρκίνο του μαστού. Οι τεχνικές που χρησιμοποιήθηκαν ήταν η ανάλυση της διαμόρφωσης της μονής έλικας του DNA (SSCP) και η απευθείας ανάλυση της αλληλουχίας των νουκλεοτιδίων (Sequencing). Επιπλέον η κλινική παρουσίαση και πρόγνωση των περιπτώσεων του καρκίνου του μαστού που συνδέονταν με μεταλλάξεις στο γονίδιο BRCA2, συγκρίθηκε με 20 επαρκώς αντίστοιχους για ηλικία και ημερομηνία διάγνωσης της νόσου ασθενείς (ομάδα ελέγχου), με σποραδικό καρκίνο του μαστού. Αναγνωρίσαμε τρεις καινούριες μεταλλάξεις (3058delA, 6024delTA και 4147delG) στην περιοχή που συνδέεται με καρκίνο των ωοθηκών (OCCR) και μία γνωστή (2024del5), στο γονίδιο BRCA2 σε πέντε διαφορετικές οικογένειες με καρκίνο του μαστού. Η μετάλλαξη 4147delG ανιχνεύθηκε σε δύο μη συγγενείς ασθενείς. Οι μεταλλάξεις στο γονίδιο BRCA2 συνδέονταν με πρώιμο καρκίνο του μαστού RR=4.77 (95% CΙ: 0.666-34.463). Οι ασθενείς με μεταλλάξεις στο γονίδιο BRCA2 δεν παρουσίασαν διαφορετικό ιστολογικό φαινότυπο ωστόσο είχαν βελτιωμένη συνολική επιβίωση (100% vs 65%). Τα ευρήματα μας συνιστούν ότι υπάρχει μία ομάδα νέων μεταλλάξεων στα εξόνια 10 και 11 σε Έλληνες ασθενείς με οικογενή καρκίνο του μαστού. Οι μεταλλάξεις αυτές παρουσίασαν ηπιότερο κλινικό φαινότυπο σε σύγκριση με τους υπόλοιπους ασθενείς που μελετήθηκαν

Novel Tools for Prostate Cancer Prognosis, Diagnosis, and Follow-Up

Prostate-specific antigen (PSA) is the main diagnostic tool when it comes to prostate cancer but it possesses serious limitations. Therefore, there is an urgent need for more sensitive and specific biomarkers for prostate cancer prognosis and patient follow-up. Recent advances led to the discovery of many novel diagnostic/prognostic techniques and provided us with many worthwhile candidates. This paper briefly reviews the most promising biomarkers with respect to their implementation in screening, early detection, diagnostic confirmation, prognosis, and prediction of therapeutic response or monitoring disease and recurrence; and their use as possible therapeutic targets. This review also examines the possible future directions in the field of prostate cancer marker research

Cellular, Molecular and Proteomic Characteristics of Early Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) accounts for the majority of primary liver cancers. Early detection/diagnosis is vital for the prognosis of HCC, whereas diagnosis at late stages is associated with very low survival rate. Early diagnosis is based on 6-month surveillance of the patient and the use of at least two imaging modalities. The aim of this study was to investigate diagnostic markers for the detection of early HCC based on proteome analysis, microRNAs (miRNAs) and circulating tumor cells (CTCs) in the blood of patients with cirrhosis or early or advanced HCC. We studied 89 patients with HCC, of whom 33 had early HCC and 28 were cirrhotic. CTCs were detected by real-time quantitative reverse transcription PCR and immunofluorescence using the markers epithelial cell adhesion molecule (EPCAM), vimentin, alpha fetoprotein (aFP) and surface major vault protein (sMVP). Expression of the five most common HCC-involved miRNAs (miR-122, miR-200a, miR-200b, miR-221, miR-222) was examined in serum using quantitative real time PCR (qRT-PCR). Finally, patient serum was analyzed via whole proteome analysis (LC/MS). Of 53 patients with advanced HCC, 27 (51%) had detectable CTCs. Among these, 10/27 (37%) presented evidence of mesenchymal or intermediate stage cells (vimentin and/or sMVP positive). Moreover, 5/17 (29%) patients with early HCC and 2/28 (7%) cirrhotic patients had detectable CTCs. Patients with early or advanced HCC exhibited a significant increase in miR-200b when compared to cirrhotic patients. Our proteome analysis indicated that early HCC patients present a significant upregulation of APOA2, APOC3 proteins when compared to cirrhotic patients. When taken in combination, this covers the 100% of the patients with early HCC. miR-200b, APOA2 and APOC3 proteins are sensitive markers and can be potentially useful in combination for the early diagnosis of HCC