Search Results (296)

Human papillomavirus (HPV)-driven oropharyngeal squamous cell carcinoma (OPSCC) has emerged as a biologically distinct entity, typically affecting younger, non-smoking patients and showing improved survival compared to HPV-negative tumors. Accurate HPV status determination is essential for correct staging, prognostic assessment, and treatment de-escalation. Despite advances, substantial variability persists among diagnostic methods and clinical workflows. A narrative review of PubMed, Scopus, and Web of Science databases was conducted up to July 2025. Studies addressing HPV detection techniques in OPSCC—including p16^INK4a^ immunohistochemistry (IHC), HPV DNA and RNA assays, liquid biopsy approaches, and computational surrogates—were critically analyzed regarding diagnostic accuracy, clinical applicability, and emerging innovations. Tissue-based assays remain the diagnostic reference standard. p16 IHC provides high sensitivity but limited specificity and should be confirmed with nucleic acid-based methods such as DNA PCR, in situ hybridization (ISH), or E6/E7 mRNA detection. Combined or “orthogonal” testing minimizes discordance and refines risk stratification. Liquid biopsy detection of circulating HPV DNA using droplet digital PCR or next-generation sequencing has shown high sensitivity and specificity in cohorts of patients with HPV-associated OPSCC, supporting its potential role as a complementary biomarker for treatment monitoring and surveillance. However, circulating HPV DNA alone does not unequivocally identify the anatomic source of HPV DNA and should be interpreted together with clinical, radiologic, and tissue-based findings. Oral rinse and saliva assays show moderate diagnostic performance, while artificial intelligence-based radiomic and histopathologic models are emerging as complementary tools. Reliable HPV attribution in OPSCC requires a multimodal diagnostic strategy integrating p16 IHC, molecular confirmation, and ctHPV-DNA monitoring. Methodological standardization and prospective validation are essential to implement precision-guided, cost-effective workflows in routine clinical practice. Full article

Minimal residual disease (MRD) refers to the persistence of low-level malignant cells or tumor-derived nucleic acids that remain after curative-intent therapy and are undetectable by conventional diagnostic methods. In oncology, MRD has emerged as a powerful biomarker with well-established prognostic value in hematologic malignancies and rapidly expanding relevance in solid tumors. Advances in sensitive detection technologies, including multiparameter flow cytometry, quantitative real-time polymerase chain reaction, next-generation sequencing, and digital polymerase chain reaction, have enabled the identification of residual disease at the molecular level, often preceding clinical or radiological relapse. Beyond its conventional role as a binary indicator of treatment response or cure, MRD is increasingly recognized as a dynamic longitudinal biomarker that supports personalized disease management. Within this evolving paradigm, patient-informed MRD strategies that incorporate tumor-specific molecular profiling and serial monitoring, particularly through circulating tumor DNA, offer the potential to guide treatment adaptation, including escalation, de-escalation, maintenance optimization, and surveillance strategies across both hematologic and solid malignancies. In this review, we summarize the biological basis of MRD, current and emerging detection methodologies, and clinical applications across cancer types, with a focus on patient-informed approaches. We also discuss key limitations, including assay standardization, biological variability in solid tumors, and the lack of clearly defined actionability thresholds. Finally, we highlight future directions for integrating MRD with multi-omics and AI-driven analytical frameworks to enable adaptive, risk-informed cancer management and advanced precision oncology. Full article

Hantaviruses are globally distributed, rodent-borne zoonotic pathogens. In Hungary, Dobrava-Belgrade virus (DOBV) and Puumala virus (PUUV) are circulating, causing hemorrhagic fever with renal syndrome and nephropathia epidemica, respectively. Due to the short viremic period, hantaviruses are primarily diagnosed by serological methods. Detection of viral nucleic acid by real-time or nested reverse transcription polymerase chain reaction (RT-PCR) is limited to samples collected in the early phase of disease. Between 2018 and 2025, 51 laboratory-confirmed hantavirus infections were identified in Hungary; 30 cases were assigned to DOBV, 20 to PUUV, and one remained undetermined. Most patients were male (82%), suggesting increased exposure-related risk. Viral RNA was detected in 21 cases, mainly from serum and whole blood samples, and sporadically from urine. In three DOBV cases, viral RNA was detectable exclusively in whole blood but not in paired serum samples. Phylogenetic analysis included four PUUV and six DOBV partial S segment sequences showing high similarity to other human- and rodent-derived samples from the region. Hantavirus infections remain infrequently diagnosed in Hungary. Our findings suggest that serum and whole blood may be useful specimen types for molecular detection, whereas urine had limited diagnostic value in our dataset. Full article

Background: Tumor-derived small extracellular vesicles (sEV), which we call TEX, carry a cargo of molecules that resembles the producer tumor cells. Circulating freely in body fluids, TEX potentially serve as a liquid tumor biopsy. TEX horizontally transfer their cargo to various recipient cells, imparting to them pro-tumor activity. Mechanisms of TEX-driven reprogramming might involve nucleic acids, especially double-stranded (ds)DNA. Methods: TEX isolated from supernatants of human tumor cells were identified as sEV, based on their size, endocytic origin and morphology. TEX treated with DNase/RNase cocktail were examined by transmission and cryo-electron microscopy and tested for biologic activity. DNA was extracted from enzyme-treated TEX, quantified by Qubit and analyzed for fragment sizes. The presence of genomic DNA in TEX was confirmed by PCR, and sequencing of the TP53 gene fragment for a mutational signature was performed. Results: Enzymatic and microscopic studies of TEX showed that nucleic acids are present in the biocorona on the outer surface. Their removal interfered with the biocorona integrity. A short TEX exposure to DNase/RNase altered their morphology without impairing vesicle functions; longer treatments induced TEX re-organization into smaller membrane-bound vesicles. The TEX lumen contained long fragments of protected genomic DNA with a mutational signature reflecting that of the tumor. Conclusions: Nucleic acids present on the TEX surface support the vesicular integrity. The TEX lumen contains membrane-protected large (ds)DNA fragments with the mutational signature of the parent tumor. The presence of surface and luminal nucleic acids in TEX, and especially their mutational signature, suggests that TEX may serve as highly promising cancer-specific biomarkers. Full article

Background/Objectives: Standardizing measurement of circulating cell-free DNA (cfDNA) in healthy individuals is critical for its application as a reference in biomarker research, yet methodological variability remains poorly documented. Methods: We systematically reviewed 35 studies (n = 1250 healthy subjects) assessing how pre-analytical handling, extraction kits, and quantification methods influence plasma cfDNA levels. We identified quantification approaches (qPCR vs. fluorometry) and use of custom extraction kits as the strongest drivers of variability. Results: In qPCR studies, including ≥ 40 subjects reduced variability, underscoring the importance of adequate sample size. Commercial kits produced more consistent yields than in-house protocols; in our dataset, many studies used Qiagen’s QIAamp Circulating Nucleic Acid Kit, which has historically served as a widely used reference platform. Blood collection in EDTA tubes had minimal impact when commercial kits were used. Conclusions: Based on these findings, we recommend EDTA tubes, a standardized commercial extraction kit, and qPCR quantification to minimize cfDNA measurement variability in healthy cohorts. Finally, we provide expected cfDNA ranges for healthy individuals based on methodological flow, which can guide future benchmarking efforts and biomarker studies, improving comparability and early-detection research. Full article

Despite the importance of wastewater environmental monitoring in disease prevention and response strategies, its use remains poorly documented in Senegal. In addition, there is more onsite sanitation than sewer networks in Dakar, and open drains channel for rainwater are also used as clandestine wastewater discharge into the sea. This study aimed to assess the presence of specific pathogens in wastewater, faecal sludge, and bathing water (the sea). Samples were taken at treatment plants, an open drain, and in the receiving environment (the sea) from June to December 2023. Total nucleic acid was subjected to multiplex qualitative qPCR using SeeGene Allplex™ kits targeting 34 gastrointestinal pathogens. Descriptive statistics, multiple correspondence analysis (MCA) and logistic regression were performed. Considering all matrices, across 51 analysed samples, the results revealed strong bacterial (96.08%, n = 49), parasitic (84.31%, n = 43), and viral (68.63%, n = 35) presence. These results showed high levels of Aeromonas spp. (96.08%), Blastocystis hominis (80.39%), Enterocytozoon (58.82%), and Norovirus GII (74.51%) among bacteria, protozoa, helminths, and viruses, respectively. Moreover, faecal sludge and pumping station samples show more identified pathogen than wastewater treatment plant and seawater samples. The MCA revealed that the dry season is spatially associated with a greater number of pathogens than the rainy season, but the latter showed a greater species diversity. Logistic regression showed that certain physicochemical parameters, including BOD5, turbidity, pH, and suspended solids, influence pathogen detection. However, qualitative detection and sampling period may constitute limitations. These results reveal that wastewater and bathing water can serve as sources of information on the circulation of pathogens of interest with epidemic potential. Therefore, this valuable epidemiological tool could serve as an adjunct to clinical surveillance in order to prevent future epidemics. Full article

Acute myeloid leukemia (AML) is a biologically heterogeneous hematologic malignancy arising from the oncogenic transformation of hematopoietic stem and progenitor cells, resulting in clonal expansion and progressive subclonal diversification. Although considerable advances have deepened our understanding of AML pathogenesis, major challenges persist, particularly regarding relapses and therapeutic resistance. In recent years, exosomes—extracellular vesicles of 30–150 nm in diameter of endosomal origin—have emerged as critical mediators of intercellular communication within the AML tumor microenvironment. These vesicles transport a diverse cargo of proteins, metabolites, and nucleic acids, including mRNA, non-coding RNA species, and DNA, which is selectively packaged during their biogenesis. Circulating exosomes have garnered attention as promising liquid biomarkers for diagnosis, prognosis, and monitoring minimal residual disease, while also representing potential therapeutic targets or delivery platforms. Nonetheless, significant knowledge gaps remain regarding the mechanisms governing exosome biogenesis, cargo selection, and the functional impact on leukemia progression and immune modulation. This review focuses on the role of exosomes in acute myeloid leukemia, with an emphasis on the molecular mechanisms underlying their involvement in pathogenesis, tumor communication, and resistance to therapies, as well as their potential as diagnostic biomarkers. Full article

Flaviviruses such as dengue virus (DENV) and Zika virus (ZIKV) co-circulate widely and cause significant morbidity, yet effective broad-spectrum antivirals are limited. This study evaluated the antiviral efficacy, cytotoxicity, and host transcriptional responses to the nucleic acid–hydrolyzing antibody fragment 3D8 scFv in mono- and co-infection models. RNA sequencing of A549 cells treated with 3D8 scFv revealed a dose-dependent activation of the MAPK–HSP70 stress response, with minimal transcriptomic disruption at antiviral concentrations. Comparative transcriptomic analysis identified distinct host signatures for ZIKV and DENV2, and machine learning classifiers accurately distinguished infection states (AUC > 0.95). In Vero E6 cells, prophylactic treatment with 3D8 scFv significantly reduced viral RNA, protein expression, and infectious particle production for both viruses, including during co-infection. Optimized post-entry treatment also demonstrated antiviral activity. Cytotoxicity assays confirmed good tolerability at effective concentrations. These findings indicate that 3D8 scFv inhibits viral replication through early cleavage of viral nucleic acids while inducing a limited protective stress response, supporting its development as a broad-spectrum antiviral candidate. Full article

Liquid biopsy (LB) offers a minimally invasive approach to characterizing and monitoring glioblastoma (GB), a tumor marked by extensive heterogeneity, limited surgical accessibility and rapid molecular evolution. By analyzing circulating tumor-derived components such as circulating tumor DNA (ctDNA), extracellular vesicles (EVs), circulating RNA species and circulating tumor cells (CTC), LB provides dynamic molecular information that cannot be captured by neuroimaging or single-site tissue sampling. Cerebrospinal fluid (CSF) currently yields the highest sensitivity for detecting tumor-specific alterations, while plasma enables repeat monitoring despite lower biomarker abundance. EVs have gained particular prominence due to their ability to preserve DNA, RNA, and protein cargo that reflects key genomic changes, treatment resistance mechanisms, and immune evasion. Although advances are substantial, clinical implementation remains constrained by low analyte concentrations, methodological variability, limited standardization and the high cost of testing, which is rarely reimbursed by insurers. This review summarizes current evidence on circulating biomarkers in GB and highlights research priorities essential for integrating LB into future diagnostic and therapeutic workflows. Full article

Despite extensive technological advances and an ever-growing body of literature, liquid biopsy has yet to achieve reliable early detection of pancreatic ductal adenocarcinoma (PDA). Numerous studies have investigated circulating tumor-derived components, including cell-free DNA (cfDNA), cell-free RNA (cfRNA), extracellular vesicles (EVs), and circulating tumor cells (CTCs), primarily using peripheral blood samples; however, their clinical utility for early-stage disease remains limited. The fundamental obstacles are biological rather than purely technical: early PDA and its precursor lesions, such as pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasms (IPMN), are characterized by minimal tumor burden, low levels of nucleic acid shedding, and substantial background signals from non-neoplastic tissues. Increasing analytical complexity through multilayered liquid biopsy approaches, including analyses from pancreas-associated fluid, has not consistently translated into improved diagnostic performance and, in some cases, has amplified issues related to specificity, reproducibility, and interpretability. Moreover, molecular alterations detected in body fluids may reflect clonal expansion without inevitable malignant progression, raising concerns regarding overdiagnosis and clinical decision-making. Pre-analytical variability, lack of standardization, and limited access to tumor-adjacent fluids further hinder clinical implementation. Liquid biopsy should therefore be regarded as a complementary modality rather than a substitute for histopathological diagnosis, with its precise clinical role in early detection still ill-defined. In this review, we critically examine why liquid biopsy has not yet succeeded in early PDA detection, highlighting the key biological, technical, and clinical barriers that must be addressed to move the field beyond exploratory research toward meaningful clinical application. Full article

Microfluidic chip technologies, also known as lab-on-a-chip systems, have profoundly transformed laboratory medicine by enabling the miniaturization, automation, and rapid processing of complex diagnostic assays using minimal sample volumes. Recent advances in chip design, fabrication methods—including 3D printing, modular and flexible substrates—and biosensor integration have significantly enhanced the performance, sensitivity, and clinical applicability of these devices. Integration of advanced biosensors allows for real-time detection of circulating tumor cells, nucleic acids, and exosomes, supporting innovative applications in cancer diagnostics, infectious disease detection, point-of-care testing (POCT), personalized medicine, and therapeutic monitoring. Notably, the convergence of microfluidics with artificial intelligence (AI) and machine learning has amplified device automation, reliability, and analytical power, resulting in “smart” diagnostic platforms capable of self-optimization, automated analysis, and clinical decision support. Emerging applications in fields such as neuroscience diagnostics and microbiome profiling further highlight the broad potential of microfluidic technology. Here, we present findings from a comprehensive review of recent innovations in microfluidic chip design and fabrication, advances in biosensor and AI integration, and their clinical applications in laboratory medicine. We also discuss current challenges in manufacturing, clinical validation, and system integration, as well as future directions for translating next-generation microfluidic technologies into routine clinical and public health practice. Full article

To address the technical bottlenecks commonly encountered with real-time quantitative PCR instruments, such as insufficient ramp rates and uneven chamber temperature distribution, this study proposes an innovative design scheme for a temperature control system that incorporates key hot air circulation and temperature field regulation technologies. By combining the PCR instruments’ working principles and structural characteristics, the failure mechanisms associated with the temperature control system are systematically analyzed, and a reliability-oriented thermodynamic analysis model is constructed to clarify the functional positioning of core components and to systematically test the airflow uniformity, temperature dynamics, and nucleic acid amplification efficiency. An integrated fixture for airflow rectifier and cruciform frames is designed, which enables precise quantitative characterization of the system temperature uniformity, ramp rates, and amplification efficiency on a multi-condition comparison platform. Through modeling analysis combined with experimental validation, the thermal performance differences among various heating chamber structures are compared, leading to a multidimensional optimization of the temperature control system. The test results demonstrate outstanding core performance metrics for the optimized system: the up ramp reaches 7.5 ± 0.1 °C/s, the down ramp reaches 13.5 ± 0.1 °C/s, and the steady-state temperature deviation is only ±0.1 °C. The total duration for 35 PCR cycles is recorded at 16.3 ± 0.6 min, with a nucleic acid amplification efficiency of 98.9 ± 0.2%. The core performance metrics comprehensively surpass those of mainstream global counterparts. The developed temperature control system is well-suited for practical applications such as rapid detection, providing critical technological support for the iterative upgrade of nucleic acid amplification techniques while laying a solid foundation for the engineering development of high-performance PCR instruments. Full article

Lung cancer in never-smokers (LCINS) represents a distinct clinical entity driven by dominant oncogenic alterations and characterized by a low tumor mutational burden. Although tyrosine kinase inhibitors (TKIs) achieve high initial response rates, their long-term efficacy is limited by suboptimal pharmacokinetics, restricted central nervous system (CNS) penetration, tumor microenvironment barriers, and acquired resistance. In this review, we critically assess the current state of nanotechnology-assisted drug delivery systems for LCINS, with a primary focus on how rationally designed nanocarriers can overcome biological barriers, enable molecular subtype-specific therapeutic strategies, and address mechanisms that limit clinical efficacy and durability of response. We conducted a structured literature search using PubMed and Web of Science (January 2022 to November 2025), focusing on primary studies reporting the preparation, physicochemical properties, and therapeutic performance of nanocarriers in in vitro and in vivo models, as well as available pharmacokinetic and clinical data. LCINS is characterized by inefficient vasculature, high extracellular matrix density, active efflux transporters, and immunosuppressive niches, and is frequently complicated by brain metastases. Nanocarrier-based platforms can enhance aqueous solubility, prolong systemic circulation, and improve tumor or CNS targeting. Co-delivery systems combining TKIs with nucleic acid-based therapeutics, together with stimuli-responsive platforms, offer the potential for simultaneous modulation of multiple oncogenic pathways and partial mitigation of resistance mechanisms. In summary, nanotechnology provides a promising strategy to improve both the efficacy and specificity of targeted therapies in LCINS. Successful clinical translation will depend on biologically aligned carrier–payload combinations, scalable and reproducible manufacturing processes, and biomarker-guided patient selection. Full article

Extracellular vesicles (EVs), particularly exosomes, are key mediators of intercellular communication, transporting biomolecules such as nucleic acids, lipids, and proteins that influence immune and metabolic pathways. In adipose tissue (AT), adipocyte-derived EVs (AdEVs) play a crucial role in maintaining metabolic homeostasis and have been implicated in obesity-related dysfunction. Among their bioactive cargo, microRNAs regulate post-transcriptional gene expression and participate in immunometabolic regulation. This study aimed to determine whether miR-34a expression in serum and circulating EVs varies according to body fat percentage, to explore its potential utility as a non-invasive biomarker of AT dysfunction. A total of 142 adults (mean age 36 ± 11 years) were classified by body fat percentage (≥25% in men, ≥35% in women). Exosomes were isolated (Invitrogen^®) and characterized by cryo-TEM, and miR-34a expression was quantified by qRT-PCR. miR-34a expression correlated negatively with Total Cholesterol, Triglycerides, LDLc/HDLc, TG/HDLc, BMI, C3, CRP, fasting insulin, HOMA-IR, HOMA-B, Body adiposity, Chemerin, CCL2, AdipoQT, and AdipoQ-H, but positively with HDLc and QUICKI. Notably, LDLc, sdLDLc, sdLDLc/LDLc, TC/HDLc, and fasting glucose showed opposite correlation patterns between serum and exosomes. Overall, serum miR-34a levels were higher than in exosomes, suggesting its potential as a biomarker of metabolic dysfunction and insulin resistance. Full article

Background/Objectives: HER2-low breast cancer has emerged as a clinically meaningful category that challenges the historical HER2-positive versus HER2-negative classification. Although not defined as a distinct biological subtype, HER2-low tumors exhibit unique clinicopathological features and differential sensitivity to novel antibody–drug conjugates. Accurate identification remains difficult due to limitations in immunohistochemistry performance, inter-observer variability, intratumoral heterogeneity, and dynamic shifts in HER2 expression over time. This review synthesizes current evidence on the biological and clinical characteristics of HER2-low breast cancer and evaluates emerging diagnostic innovations, with emphasis on liquid biopsy approaches and evolving technologies that may enhance diagnostic accuracy and monitoring. Methods: A narrative literature review was conducted, examining tissue-based HER2 testing, liquid biopsy modalities, including circulating tumor cells, circulating nucleic acids, extracellular vesicles, and soluble HER2 extracellular domains, and applications of artificial intelligence (AI) across histopathology and multimodal diagnostic systems. Results: Liquid biopsy technologies offer minimally invasive, real-time assessment of HER2 dynamics and may overcome fundamental limitations of tissue-based assays. However, these platforms require rigorous analytical validation and face regulatory and standardization challenges before widespread clinical adoption. Concurrently, AI-enhanced histopathology and multimodal diagnostic systems improve reproducibility, refine HER2 classification, and enable more accurate prediction of treatment response. Emerging biosensor- and AI-enabled monitoring frameworks further support continuous disease evaluation. Conclusions: HER2-low breast cancer sits at the intersection of evolving pathology and technological innovation. Integrating liquid biopsy platforms with AI-driven diagnostics has the potential to advance precision stratification and guide personalized therapeutic strategies for this expanding patient subgroup. Full article