Search Results (296)

The detection of circulating tumor cells (CTCs) using immunoaffinity-based methods often relies on epithelial-related markers, which may bias the selection of CTCs and limit the biological information obtained, depending on the targeted antigens. Herein, we compared the molecular profiles and clinical significance of CTCs based on the expression of epithelial-related markers (EPCAM, EGFR, and MET) in patients with head and neck squamous-cell carcinoma (HNSCC). CTCs were detected using density gradient separation and CD45-negative selection, followed by quantitative PCR for epithelial-related marker expression. Expression profiles of epithelial–mesenchymal transition (EMT)-related (VIM, CDH1, CDH2, SNAI1, ZEB1, ZEB2, and TWIST1) and immune-regulatory (CD274 and PDCD1LG2) genes were compared. Moreover, the association between marker expression and clinical factors was analyzed. Among the 60 patients with CTCs, 48 (80.0%), 20 (33.3%), and 31 (51.7%) were positive for EPCAM, EGFR, and MET, respectively. A significant correlation was observed between CTCs expressing EPCAM and EGFR. CTCs expressing distinct markers showed differing EMT-related and immune-regulatory gene expression. EPCAM+ CTCs were associated with advanced-stage disease, while EGFR+ CTCs were correlated with locoregional relapse and shorter progression-free survival (p = 0.007; hazard ratio = 3.254). Patients with EPCAM/EGFR double-positive CTCs had the poorest prognosis. These findings emphasize the importance of marker selection in liquid biopsy technologies and highlight the need for improved detection methods and the further investigation of CTC biology. Full article

Early diagnosis of diseases would significantly increase the survival rate of cancer patients. However, current screening methods are complex and costly, making them unsuitable for rapid health diagnosis in daily life. Here, we develop a portable platform based on a planar-gate graphene field-effect transistor functionalized with polydopamine self-assembled film (PDA-GFET), capable of identifying colon cancer through the detection of EpCAM protein, which is expressed on colon cancer-derived exosomes, in clinical samples within 10 min. The PDA self-assembled film on the graphene and gate surface enhances the biosensor’s functionalization area while suppressing non-specific adsorption, thereby achieving detection limits as low as 112 particles/mL. In addition, the PDA-GFET-based detection platform was used to identify EpCAM protein in real clinical samples from healthy individuals and colon cancer patients within 10 min, and the two showed significant differences (p < 0.001). Results indicate that the proposed PDA-GFET-based detection platform is expected to be a potential tool for the early diagnosis of colon cancer. Full article

Human milk is a complex biofluid containing a diverse array of cells crucial for infant health. Despite their importance, our understanding of these cells remains incomplete due to technical challenges. To fully comprehend human milk cells, high-resolution imaging technologies that can directly measure biological processes are required. We have developed a specialized imaging platform combining light and electron microscopy for human milk cell imaging. To identify different cell types, human milk cells were first stained with several specific cell markers (e.g., EpCAM and MUC1 for lactocytes, CD16 and CD66b for neutrophils, and HLA-DR and CD68 for macrophages) prior to light (confocal) microscopy. Following this, the same cells were processed with osmium staining, resin embedding, and sectioning for electron microscopy, allowing us to observe ultrastructural details. Our imaging workflow has enabled nanoscale visualization of human milk cells, resulting in a first-of-its-kind comprehensive database profiling the organelle-level ultrastructure of different cell types present in human milk. The cells in the human milk are highly heterogenous, featuring a large proportion of lactocytes and lipid droplets, binucleated lactocytes, neutrophil aggregation, neutrophil extracellular traps, dendritic cells/macrophages with bacteria, and immunophagocytosis. This study provides valuable cellular insights contributing to a deeper understanding of human milk biology. Full article

Circulating tumor cells (CTCs) are pivotal in cancer metastasis and serve as valuable biomarkers for diagnosis, prognosis, and treatment monitoring. Traditional CTC capture methods predominantly utilize the epithelial cell adhesion molecule (EpCAM) as a marker for isolation. However, the heterogeneity of these circulating cells and the epithelial-to-mesenchymal transition process (wherein epithelial cells acquire mesenchymal characteristics) limit the efficacy of EpCAM-based capture techniques. In this paper, we critically review the role of the EpCAM in CTC capture, explore the impact of epithelial-to-mesenchymal transition on EpCAM expression, and discuss alternative biomarkers and strategies to enhance CTC isolation. By evaluating the limitations of EpCAM-mediated capture and the challenges posed by epithelial-to-mesenchymal transition, we aim to provide insights into the development of more comprehensive liquid biopsy approaches for cancer management. Full article

Oral cancer, a subtype of head and neck cancer, poses significant global health challenges owing to its late diagnosis and high metastatic potential. The epithelial cell adhesion molecule (EpCAM), a transmembrane glycoprotein, has emerged as a critical player in cancer biology, particularly in oral cancer stem cells (CSCs). This review highlights the multifaceted roles of EPCAM in regulating oral cancer metastasis, tumorigenicity, and resistance to therapy. EpCAM influences key pathways, including Wnt/β-catenin and EGFR, modulating CSC self-renewal, epithelial-to-mesenchymal transition (EMT), and immune evasion. Moreover, EpCAM has been implicated in metabolic reprogramming, epigenetic regulation, and crosstalk with other signaling pathways. Advances in EpCAM-targeting strategies, such as monoclonal antibodies, chimeric antigen receptor (CAR) T/NK cell therapies, and aptamer-based systems hold promise for personalized cancer therapies. However, challenges remain in understanding the precise mechanism of EpCAM in CSC biology and its translation into clinical applications. This review highlights the need for further investigation into the role of EPCAM in oral CSCs and its potential as a therapeutic target to improve patient outcomes. Full article

Prostate cancer is a leading cause of cancer-related mortality in men, with radiotherapy (RT) playing a pivotal role in treatment. However, reliable biomarkers for assessing relapse risk following RT remain scarce. This study aimed to evaluate circulating epithelial tumor cells (CETC/CTC) as potential biomarkers for assessing relapse risk in prostate cancer patients undergoing RT. Peripheral blood samples were collected from 52 prostate cancer patients, and CETC/CTC were detected using the EpCAM surface marker. Patients received definitive, adjuvant, or salvage RT, and CETC/CTC counts were measured before, at mid-treatment, and at the end of RT. The association between changes in CETC/CTC counts and relapse risk was examined. CETC/CTC were detected in 96% of patients prior to RT. A significant reduction in CETC/CTC counts during RT, particularly in patients who had undergone surgery, was associated with a lower relapse risk. In contrast, an increase in CETC/CTC counts during or after RT was associated with a higher relapse risk (hazard ratio = 8.8; p = 0.002). Furthermore, 36% of patients receiving adjuvant RT and 14% of those receiving definitive RT relapsed, with a higher risk observed in patients showing increasing CETC/CTC counts during RT. Among patients receiving salvage RT, 18% relapsed, though changes in CETC/CTC counts were less significantly associated with relapse. Monitoring CETC/CTC levels during RT offers important prognostic insights into relapse risk in prostate cancer patients. While changes in CETC/CTC counts correlated with relapse, PSA levels measured during the study did not reliably reflect relapse risk in this cohort. CETC/CTC shows promise as a prognostic marker, though further studies are required to validate its clinical superiority over PSA. Full article

The presence of metastases in mediastinal lymph nodes (LNs) is essential for planning lung cancer treatment and assessing anticancer immune responses. The aim of the study was to assess LNs for the presence of neoplastic cells and evaluate lung cancer-selected antigen expression. LN aspirates were obtained during an EBUS/TBNA procedure. The cells were analyzed using a hematological analyzer and flow cytometry. It was possible to indicate the presence of cells characterized by high fluorescence connected with high metabolic activity using a hematological analyzer and to determine their non-hematopoietic origin using flow cytometry. Using these methods together, we detected very quickly a high proportion of cancer cells in LNs. We noticed that it was possible to determine a high expression of EpCAM, TTF-1, Ki67, cytokeratin, HER, and differences between non-small-cell (NSCLC) and small-cell lung cancer (SCLC) for the antigens MUC-1, CD56, HLA-DR, CD39, CD184, PD-L1, PD-L2 and CTLA-4 on tumor cells. We report, for the first time, that the detection of tumor cells in LNs with the expression of specific antigens is easy to evaluate using a hematological analyzer and flow cytometry in EBUS/TBNA samples. Such precise characteristics of non-hematopoietic cells in LNs may be of great diagnostic importance in the detection of micrometastases. Full article

In today’s world, with its continuing advancements in genetics, the identification of Lynch syndrome (LS) increasingly relies on sophisticated genetic testing techniques. Most guidelines recommend a tailored surveillance program, as well as personalized prophylactic and therapeutic approaches, according to the type of dMMR gene mutation. Carriers of path_MLH1 and path_MSH2 genes have a higher risk of developing colorectal cancer (CRC), despite intensive colonoscopic surveillance. Conversely, carriers of path_MSH6 and path_PMS2 genes have a lower risk of developing CRC, which may be due to their lower penetrance and later age of onset. Thus, carriers of path_MLH1 or path_MSH2 would theoretically derive greater benefits from total colectomy, compared to low-risk carriers (path_MSH6 and path_PMS2), in which colonoscopic surveillance might achieve an efficient prophylaxis. Furthermore, regarding the risk of endometrial/ovarian cancer development, there is a global agreement to offer both hysterectomy and bilateral salpingo-oophorectomy to path_MLH1, path_MSH2 and path_MSH6 carriers after the age of 40. In patients with CRC, preoperative knowledge of the diagnosis of LS is of tremendous importance, due to the high risk of metachronous CRC. However, this risk depends on the type of dMMR gene mutation. For carriers of the high-risk variants (MLH1, MSH2 and EPCAM) who have already developed colon cancer, it is strongly recommended a subtotal or total colectomy is performed, while partial colectomy followed by endoscopic surveillance is an appropriate management approach to treat colon cancer in carriers of the low-risk variants (MSH6 and PMS2). On the other hand, extended surgery for index rectal cancer (such as total proctocolectomy) is less effective than extended surgery for index colon cancer from the point of view of metachronous CRC risk reduction, and is associated with a decreased quality of life. Full article

Mebendazole (MBZ), a benzimidazole anthelmintic and cytoskeleton-disrupting compound, exhibits antitumor properties; however, its action on ovarian cancer (OC) is not clearly understood. This study evaluates the effect of MBZ on OC cell lines OVCAR3 and OAW42, focusing on cell proliferation, migration, invasion, and cancer stemness. The underlying mechanisms, including cytoskeletal disruption, epithelial–mesenchymal transition (EMT), and signaling pathways, were explored. MBZ inhibited OVCAR3 and OAW42 cell proliferation in a dose- and time-dependent manner. Additionally, MBZ significantly impedes migration, spheroid invasion, colony formation, and stemness. In addition, it reduced actin polymerization and down-regulated CSC markers (e.g., CD24, CD44, EpCAM). Moreover, MBZ suppressed MMP-9 activity and inhibited the EMT marker as judged by decreased N-Cadherin and Vimentin and increased E-Cadherin. Furthermore, MBZ induced G2/M cell cycle arrest by modulating Cyclin B1, CDC25C, and WEE1. Also, it triggered apoptosis by disrupting mitochondrial membrane potential. Mechanistic studies revealed a significant downregulation of Girdin, an Akt modulator, along with reduced p-Akt, p-IKKα/β, and p-NF-κB, indicating MBZ’s novel mechanism of action through the Girdin-mediated Akt/IKKα/β/NF-κB signaling axis. Thus, by targeting Girdin, MBZ presents a promising repurposed therapeutic strategy to inhibit cancer cell proliferation and metastasis in ovarian cancer. Full article

Background/Objectives: Photoimmunotherapy (PIT) is an innovative approach for the targeted therapy of cancer. In PIT, photosensitizer dyes are conjugated to tumor-specific antibodies for targeted delivery into cancer cells. Upon irradiation with visible light, the photosensitizer dye is activated and induces cancer-specific cell death. In the present article, we describe the PIT of prostate cancer (PC) as a therapeutic option for the targeted treatment of localized prostate cancer. Methods: We conjugated the silicon phthalocyanine dye WB692-CB2 to recombinant cysteine-modified anti-CD44 and anti-EpCAM antibodies via a maleimide linker and tested the antibody dye conjugates for PIT on PC cells and prostate cancer stem cell (PCSC)-like cells. Results: The anti-CD44 and anti-EpCAM antibody dye conjugates showed specific binding and high cytotoxicity against PC and PCSC-like cells following irradiation with red light. Combined treatment with both conjugates led to enhanced cytotoxic effects. Conclusions: PIT with our dye WB692-CB2 can serve as an effective focal therapy against prostate cancer, preserving the prostate gland and minimizing side effects. It can be employed during radical prostatectomy (RP) to treat residual tumor cells or lymph node metastases in areas where further surgical intervention is not feasible. Utilizing multiple conjugates against antigens expressed on differentiated PC and PCSC-like cells, such as CD44 and EpCAM, could be an effective method to eradicate residual cancer cells in heterogeneous tumors. This approach could reduce the risk of local recurrence after RP and thus increase the therapeutic outcome of PC patients. Full article

Lymphocyte antigen 6 complex, locus K (LY6K) is a putative oncogene in various human cancers, including colorectal cancer, where elevated expression is associated with poor prognosis. This study investigates the antitumor effects of LY6K in colon cancer stem cells (CCSCs) both in vitro and in vivo. EpCAM and CD44 surface markers were used to isolate CCSCs from HCT116 cells, and the expression of LY6K in CCSCs was analyzed by real-time PCR. RNA interference was used to silence LY6K to evaluate its potential role of LY6K on the proliferation, migration, invasion, and cell cycle of CCSCs. Functional assays, including MTS assays, flow cytometric analyses, Transwell migration and invasion assays, and a xenograft model, were used for analysis. The results revealed that LY6K was highly expressed in CCSCs. siRNA-mediated LY6K-silencing in CCSCs inhibited cell proliferation by inducing G1 phase cell cycle arrest and suppressed migration and invasion. In vivo, LY6K silencing effectively reduced tumor growth and extended survival in a mouse model. These findings suggest LY6K as a promising therapeutic target for eradicating CCSCs in colorectal cancer treatment. Full article

The detection and analysis of cancer cell exosomes with high sensitivity and precision are pivotal for the early diagnosis and treatment strategies of prostate cancer. To this end, a microfluidic chip, equipped with a cactus-like array substrate (CAS) based on surface-enhanced Raman spectroscopy (SERS) was designed and fabricated for the detection of exosome concentrations in Lymph Node Carcinoma of the Prostate (LNCaP). Double layers of polystyrene (PS) microspheres were self-assembled onto a polyethylene terephthalate (PET) film to form an ordered cactus-like nanoarray for detection and analysis. By combining EpCAM aptamer-labeled SERS nanoprobes and a CD63 aptamer-labeled CAS, a ‘sandwich’ structure was formed and applied to the microfluidic chips, further enhancing the Raman scattering signal of Raman reporter molecules. The results indicate that the integrated microfluidic sensor exhibits a good linear response within the detection concentration range of 10⁵ particles μL⁻¹ to 1 particle μL⁻¹. The detection limit of exosomes in cancer cells can reach 1 particle μL⁻¹. Therefore, we believed that the CAS integrated microfluidic sensor offers a superior solution for the early diagnosis and therapeutic intervention of prostate cancer. Full article

Using multigene panel testing for the diagnostic evaluation of patients with hereditary breast and ovarian cancer (HBOC) syndrome often identifies clinically actionable variants in genes with varying levels of penetrance. High-penetrance genes (BRCA1, BRCA2, CDH1, PALB2, PTEN, STK11, TP53) inform specific clinical surveillance and therapeutic decisions, while recommendations for moderate-penetrance genes (ATM, BARD1, BRIP1, CHEK2, MLH1, MSH2, MSH6, PMS2, EPCAM, NF1, RAD51C, RAD51D) are more limited. A detailed disease history, including pedigree data, helps formulate the most appropriate and personalised management strategies. In this study, we evaluated the clinical benefits of comprehensive hereditary cancer gene panel testing and a pre-sent questionnaire in Hungarian patients with suspected HBOC syndrome. We prospectively enrolled 513 patients referred for HBOC testing. Of these, 463 met the genetic testing criteria, while 50 did not but were tested due to potential therapeutic indications. Additionally, a retrospective cohort of 47 patients who met the testing criteria but had previously only been tested for BRCA1/2 was also analysed. Among the 463 patients in the prospective cohort, 96 (20.7%) harboured pathogenic/likely pathogenic (P/LP) variants—67 in high-penetrance genes and 29 in moderate-penetrance genes. This ratio was similar in the retrospective cohort (6/47; 12.7%). In patients who did not meet the testing criteria, no mutations in high-penetrance genes were found, and only 3 of 50 (6%) harboured P/LP variants in moderate-penetrance genes. Secondary findings (P/LP variants in non-HBOC-associated genes) were identified in two patients. In the prospective cohort, P/LP variants in BRCA1 and BRCA2 were the most prevalent (56/96; 58.3%), and the extended testing doubled the P/LP detection ratio. Among moderate-penetrance genes, five cases (three in the prospective and two in the retrospective cohorts) had P/LP variants in Lynch syndrome-associated genes. Further immunohistochemistry analysis of breast tumour tissues helped clarify the causative role of these variants. Comprehensive clinical and molecular genetic evaluation is beneficial for the diagnosis and management of patients with P/LP variants in hereditary tumour-predisposing genes and can serve as a basis for effective therapy selection, such as PARP inhibitors or immunotherapy. Full article

Monoclonal antibodies (mAbs) are widely used in cancer therapy but often show limited efficacy for solid tumors. Enhancing anti-tumor activity by fusing cytokines to tumor-targeting mAbs, which specifically activate immune cells within the tumor microenvironment, represents a promising strategy. However, the optimal design and therapeutic efficacy of antibody–cytokine fusion formats remain unclear. The epithelial cell adhesion molecule (EpCAM), frequently overexpressed in a variety of carcinomas, serves as the target for immunotherapies. In this study, we identified a fully human mAb targeting EpCAM, designated as m801, from a previously constructed phage-displayed fully human antibody library. By fusing m801 with an IL-2 variant (IL-2v) in two configurations, m801.2 (2 anti-EpCAM Fab + 1 IL-2v) and m801.3 (1 anti-EpCAM Fab + 1 IL-2v), we identified m801.2 as the lead candidate due to its superior biophysical properties, including high thermal stability, homogeneity, and low aggregation. Furthermore, m801.2 showed strong binding affinity to EpCAM, with KD values of 0.6 nM, and an EpCAM-expressing tumor cell line, comparable to the original IgG m801. Additionally, m801.2 exhibited IL-2 receptor β subunit (IL-2Rβ)-biased binding activity, with a KD of 27.3 nM, resulting in superior effective T cell activation. In an SW480 xenograft mice model, m801.2 significantly inhibited tumor growth and demonstrated high tolerability. These findings suggest a valuable framework for the future design of immunocytokine therapies. Full article

Single-vesicle molecular profiling of cancer-associated extracellular vesicles (EVs) is increasingly being recognized as a powerful tool for cancer detection and monitoring. Mask and target dual imaging is a facile method to quantify the fraction of the molecularly targeted population of EVs in biofluids at the single-vesicle level. However, accurate and efficient dual imaging vesicle analysis has been challenging due to the interference of false signals on the mask images and the need to analyze a large number of images in clinical samples. In this work, we report a fully automatic dual imaging analysis method based on machine learning and use it with dual imaging single-vesicle technology (DISVT) to detect breast cancer at different stages. The convolutional neural network Resnet34 was used along with transfer learning to produce a suitable machine learning model that could accurately identify areas of interest in experimental data. A combination of experimental and synthetic data were used to train the model. Using DISVT and our machine learning-assisted image analysis platform, we determined the fractions of EpCAM-positive EVs and CD24-positive EVs over captured plasma EVs with CD81 marker in the blood plasma of pilot HER2-positive breast cancer patients and compared to those from healthy donors. The amount of both EpCAM-positive and CD24-positive EVs was found negligible for both healthy donors and Stage I patients. The amount of EpCAM-positive EVs (also CD81-positive) increased from 18% to 29% as the cancer progressed from Stage II to III. No significant increase was found with further progression to Stage IV. A similar trend was found for the CD24-positive EVs. Statistical analysis showed that both EpCAM and CD24 markers can detect HER2-positive breast cancer at Stages II, III, or IV. They can also differentiate individual cancer stages except those between Stage III and Stage IV. Due to the simplicity, high sensitivity, and high efficiency, the DISVT with the AI-assisted dual imaging analysis can be widely used for both basic research and clinical applications to quantitatively characterize molecularly targeted EV subtypes in biofluids. Full article