Liquid biopsies refer to the isolation and analysis of tumor-derived biological material from body fluids, most commonly blood, in order to provide clinically valuable information for the management of cancer patients. Their non-invasive nature allows to overcome the limitations of tissue biopsy and complement the latter in guiding therapeutic decision-making. In the past years, several studies have demonstrated that circulating tumor DNA (ctDNA) detection can be used in the clinical setting to improve patient prognosis and monitor therapy response, especially in metastatic cancers. With the advent of significant technological advances in assay development, ctDNA can now be accurately and reliably identified in early-stage cancers despite its low levels in the bloodstream. In this review, we discuss the most important studies that highlight the potential clinical utility of ctDNA in early-stage breast cancer focusing on early diagnosis, detection of minimal residual disease and prediction of metastatic relapse. We also offer a concise description of the most sensitive techniques that are deemed appropriate for ctDNA detection in early-stage cancer and we examine their advantages and disadvantages, as they have been employed in various studies. Finally, we discuss future perspectives on how ctDNA could be better integrated into the everyday oncology practice.

Background: Gastric cancer (GC) is one of the most common and aggressive types of cancer. Immune checkpoint inhibitors (ICIs) have proven effective in treating various types of cancer. The use of ICIs in GC patients is currently an area of ongoing research. The tumor microenvironment (TME) also seems to play a crucial role in cancer progression. Tumor-associated macrophages (TAMs) are the most abundant population in the TME. TAMs are capable of displaying programmed cell death protein 1 (PD-1) on their surface and can form a ligand with programmed death ligand 1 (PD-L1), which is found on the surface of cancer cells. Therefore, it is expected that TAMs may significantly influence the immune response related to immune checkpoint inhibitors (ICIs). Aim of the study: Understanding the role of TAMs and PD-1/PD-L1 networking in GC. Methods: A systematic review of published data was performed using MEDLINE (PubMed), Embase, and Cochrane databases. We retrieved articles investigating the co-existence of TAMs and PD-1 in GC and the prognosis of patients expressing high levels of PD-1+ TAMs. Results: Ten articles with a total of 2277 patients were included in the systematic review. The examined data suggest that the expression of PD-L1 has a positive correlation with the infiltration of TAMs and that patients who express high levels of PD-1+ TAMs may have a worse prognosis than those who express low levels of PD-1+ TAMs. Conclusions: TAMs play a pivotal role in the regulation of PD-1/PD-L1 networking and the progression of GC cells. Nevertheless, additional studies are needed to better define the role of TAMs and PD-1/PD-L1 networking in GC.

Bladder cancer represents a major health issue. Transurethral resection is the first line treatment and an accurate assessment of tumor margins might warrant complete tumor removal. Genomic instability and proliferative potential are common hallmarks of cancer cells. We have previously demonstrated the utility of intraoperative flow cytometry (iFC), a next-generation margin evaluation methodology for assessment of DNA content, in the detection of several types of malignancy. In the current study we investigated the possible value of iFC in the characterization of bladder cancer during surgery. Samples from a population of 52 people with urothelial cancer were included in the study. The total time for iFC evaluation is 3–5 min per sample and included a two-step analysis, including DNA-index and Tumor-index calculation. First, DNA-index calculation revealed 24 hyperploid and one hypoploid tumor. Second, cell cycle analysis and Tumor-index calculation revealed that tumor samples are distinguished from normal cells based on their significantly higher proliferative potential. The standard for iFC evaluation was pathology assessment and revealed that our protocol exhibits an accuracy of 98% in defining the presence of cancer cells in a given sample. Our results support the further assessment of iFC value towards its use as a novel malignancy evaluation tool in transurethral resections.

Non-melanoma skin cancer (NMSC) is the most prevalent cancer in humans, with a high global incidence. We present a prospective clinical feasibility study on the use of intraoperative flow cytometry (iFC) for the instant diagnosis of NMSC and its complete surgical clearance. Flow cytometry, a laser-based technique, quantifies cell features, which has applications in cancer research. This study aim is to explore the potential applicability of iFC in detecting and characterizing NMSC and its surgical margins. In total, 30 patients who underwent diagnosis for NMSC were recruited. The method demonstrated high sensitivity (95.2%) and specificity (87.1%), with an accuracy of 91.1%, as confirmed with a receiver operating characteristic curve analysis. The results also indicated that most tumors were diploid, with two cases being hypoploid. The average G0/G1 fractions for normal and tumor tissue samples were 96.03 ± 0.30% and 88.03 ± 1.29%, respectively, with the tumor index escalating from 3.89 ± 0.30% to 11.95 ± 1.29% in cancerous cells. These findings underscore iFC’s capability for precise intraoperative NMSC characterization and margin evaluation, promising enhanced complete tumor excision rates. Given the technique’s successful application in various other malignancies, its implementation in NMSC diagnosis and treatment holds significant promise and warrants further research in clinical trials.

The accurate assessment of malignancy and the precise characterization of cancer type are pivotal in guiding clinical decisions and ensuring optimal patient outcomes. The challenging task of identifying the primary site of carcinoma, especially during a surgical procedure, is not always possible and necessitates the exploration of novel, innovative diagnostic techniques. In this report, we detail a unique case of carcinoma of unknown primary origin encountered during an orthopedic surgical procedure. We introduce a novel intraoperative flow cytometry (iFC) methodology, distinctly different from traditional flow cytometry, which is tailored for real-time assessment during surgeries. This iFC technique, applied in a bone metastasis case of unknown primary origin, enabled DNA content analysis and the quantification of cytokeratin for rapid malignancy characterization, presenting an avenue for immediate surgical guidance and decision making. The case was distinctively characterized using iFC, shedding light on the potential of this emerging technique. iFC has been gaining traction because of its ability to facilitate cancer cell assessment and margin evaluation. It has been successfully applied across a wide spectrum of neoplastic conditions. Our innovative, two-step approach using iFC encompasses (1) DNA content analysis, which serves as a reliable indicator for the detection of cancer cells, and (2) the quantification of cytokeratin, a pivotal marker, which aids in the characterization and classification of a malignancy. In this case, the malignancy was classified as a carcinoma. The findings obtained from iFC were subsequently validated through pathology assessment, confirming the accuracy and reliability of our approach. This noteworthy case strengthens the potential of iFC as a novel tool in malignancy assessment, which is not just limited to cell cycle analysis but instead extends beyond this application. The promising results obtained from this case study serve as a testament to the need for more extensive research in clinical studies.