Bridging the divide: addressing discrepancies between clinical guidelines, policy guidelines, and biomarker utilization.

OBJECTIVES: This paper aims to identify and address gaps in cancer treatment and diagnosis within European health services, focusing specifically on discrepancies between clinical guidelines and policy guidelines. It seeks to highlight how the underutilization of advanced diagnostic techniques recommended by medical societies contributes to missed opportunities for improving patient outcomes. METHODS: A comprehensive analysis was conducted across multiple European countries to assess the compliance and integration of clinical guidelines with the availability of advanced diagnostic technologies. Secondary data related to clinical and policy guidelines in cancer care were collected and analyzed. Key indicators of adoption and utilization of next-generation sequencing and liquid biopsy were examined to evaluate their impact on health service efficiency and patient care. RESULTS: The analysis revealed significant discrepancies between the recommendations of medical societies regarding advanced diagnostic techniques and their adoption in health policy decisions across Europe. Country-specific assessments indicated varying levels of alignment between clinical guidelines and the availability of advanced diagnostics. These findings underscored missed opportunities for optimizing patient care and health service efficiency through better alignment and integration of clinical guidelines with policy decisions. CONCLUSIONS: This study concludes that there is a critical need for health policy decision-makers to prioritize the adoption of clinical guidelines in resource allocation and health service organization. Greater attention to the recommendations of medical societies regarding advanced diagnostic techniques could significantly enhance diagnostic accuracy, treatment efficacy, and overall patient outcomes in cancer care. The paper advocates for policy reforms that acknowledge and leverage the potential benefits of advanced diagnostics in improving health service performance and patient-centered care across Europe.

The Significance of Circulating Tumor Cells in Pancreatic Cancer.

The notion that technically resectable pancreatic ductal adenocarcinoma presents as localized disease is now known to be inaccurate. Evidence supports that most patients have subclinical systemic dissemination at the time of diagnosis. It is now widely accepted that both a local and systemic component of disease coexist, each requiring treatment of improved survival and potential cure. The advent of multiagent chemotherapy regimens has resulted in a modest improvement in survival. Consequently, this article will emphasize the expanding potential and significance of circulating tumor cells in the prognostication and management of patients with pancreatic cancer.

Transformer-based AI technology improves early ovarian cancer diagnosis using cfDNA methylation markers.

Epithelial ovarian cancer (EOC) is the deadliest women's cancer and has a poor prognosis. Early detection is the key for improving survival (a 5-year survival rate in stage I/II is over 70% compared to that of 25% in stage III/IV) and can be achieved through methylation markers from circulating cell-free DNA (cfDNA) using a liquid biopsy. In this study, we first identify top 500 EOC markers differentiating EOC from healthy female controls from 3.3 million methylome-wide CpG sites and validated them in 1,800 independent cfDNA samples. We then utilize a pretrained AI transformer system called MethylBERT to develop an EOC diagnostic model which achieves 80% sensitivity and 95% specificity in early-stage EOC diagnosis. We next develop a simple digital droplet PCR (ddPCR) assay which archives good performance, facilitating early EOC detection.

The landscape of circulating tumor HPV DNA and TTMV-HPVDNA for surveillance of HPV-oropharyngeal carcinoma: systematic review and meta-analysis.

BACKGROUND: Human papilloma virus (HPV) related cancers of the oropharynx are rapidly increasing in incidence and may soon represent the majority of all head and neck cancers. Improved monitoring and surveillance methods are thus an urgent need in public health. MAIN TEXT: The goal is to highlight the current potential and limitations of liquid biopsy through a meta analytic study on ctHPVDNA and TTMV-HPVDNA. It was performed a Literature search on articles published until December 2023 using three different databases: MEDLINE, Embase, and Cochrane Library. Studies that evaluated post-treatment ctHPVDNA and TTMV-HPVDNA in patients with HPV + OPSCC, studies reporting complete data on the diagnostic accuracy in recurrence, or in which the number of true positives, false positives, true negatives, and false negatives was extractable, and methods of detection of viral DNA clearly defined. The meta-analysis was conducted following the Meta-analysis Of Observational Studies in Epidemiology (MOOSE) reporting guidelines. The aim of this meta-analysis was to evaluate the sensitivity, specificity, and accuracy of ctHPVDNA and TTMV by ddPCR to define its efficacy in clinical setting for the follow up of HPV-OPSCC. CONCLUSION: The 12 studies included in the meta-analysis provided a total of 1311 patients for the analysis (398 valuated with ctHPVDNA and 913 with TTMV-HPVDNA). Pooled sensitivity and specificity were 86% (95% CI: 78%-91%) and 96% (95% CI: 91%-99%), respectively; negative and positive likelihood ratios were 0.072 (95% CI: 0.057-0.093) and 24.7 (95% CI: 6.5-93.2), respectively; pooled DOR was 371.66 (95% CI: 179.1-918). The area under the curve (AUC) was 0.81 (95% CI, 0.67-0.91). Liquid biopsy for the identification of cell free DNA might identify earlier recurrence in HPV + OPSCC patients. At the present time, liquid biopsy protocol needs to be standardized and liquid biopsy cannot yet be used in clinical setting. In the future, a multidimensional integrated approach which links multiple clinical, radiological, and laboratory data will contribute to obtain the best follow-up strategies for the follow-up of HPV-OPSCC.

Oligo cyc-DEP: On-chip cyclic immunofluorescence profiling of cell-derived nanoparticles.

We present a follow-on technique for the cyclic-immunofluorescence profiling of suspension particles isolated using dielectrophoresis. The original lab-on-chip technique ("cyc-DEP" [cyclic immunofluorescent imaging on dielectrophoretic chip]) was designed for the multiplex surveillance of circulating biomarkers. Nanoparticles were collected from low-volume liquid biopsies using microfluidic dielectrophoretic chip technology. Subsequent rounds of cyclic immunofluorescent labeling and quenching were imaged and quantified with a custom algorithm to detect multiple proteins. While cyc-DEP improved assay multiplicity, long runtimes threatened its clinical adoption. Here, we modify the original cyc-DEP platform to reduce assay runtimes. Nanoparticles were formulated from human prostate adenocarcinoma cells and collected using dielectrophoresis. Three proteins were labeled on-chip with a mixture of short oligonucleotide-conjugated antibodies. The sample was then incubated with complementary fluorophore-conjugated oligonucleotides, which were dehybridized using an ethylene carbonate buffer after each round of imaging. Oligonucleotide removal exhibited an average quenching efficiency of 98 ± 3% (n = 12 quenching events), matching the original cyc-DEP platform. The presented "oligo cyc-DEP" platform achieved clinically relevant sample-to-answer times, reducing the duration for three rounds of cyclic immunolabeling from approximately 20 to 6.5 h-a 67% decrease attributed to rapid fluorophore removal and the consolidated co-incubation of antibodies.

Challenges in the treatment of BRAF K601E-mutated lung carcinoma: a case report of rapid response and resistance to dabrafenib and trametinib.

We report a case of limited effectiveness of dabrafenib and trametinib in a 59-year-old man with poorly differentiated lung carcinoma and a rare BRAF K601E mutation. The patient, unresponsive to chemotherapy and immunotherapy, received these targeted agents as second-line treatment. Despite a notable initial response, tumor regression lasted only 52 days. A subsequent liquid biopsy revealed additional alterations (BRAF amplification, KIT amplification, TP53 S241F), indicating a complex resistance mechanism. This case underscores the challenges in treating BRAF K601E-mutant lung carcinoma, emphasizing the need for advanced molecular diagnostics, personalized approaches, and further research into more effective therapies for unique genetic profiles.

Recent progress in Surface-Enhanced Raman Spectroscopy detection of biomarkers in liquid biopsy for breast cancer.

Breast cancer is the most commonly diagnosed cancer in women globally and a leading cause of cancer-related mortality. However, current detection methods, such as X-rays, ultrasound, CT scans, MRI, and mammography, have their limitations. Recently, with the advancements in precision medicine and technologies like artificial intelligence, liquid biopsy, specifically utilizing Surface-Enhanced Raman Spectroscopy (SERS), has emerged as a promising approach to detect breast cancer. Liquid biopsy, as a minimally invasive technique, can provide a temporal reflection of breast cancer occurrence and progression, along with a spatial representation of overall tumor information. SERS has been extensively employed for biomarker detection, owing to its numerous advantages such as high sensitivity, minimal sample requirements, strong multi-detection ability, and controllable background interference. This paper presents a comprehensive review of the latest research on the application of SERS in the detection of breast cancer biomarkers, including exosomes, circulating tumor cells (CTCs), miRNA, proteins and others. The aim of this review is to provide valuable insights into the potential of SERS technology for early breast cancer diagnosis.

Elevated serum soluble programmed death ligand 1 (sPD-L1) level correlate with clinical characteristics in breast cancer patients: A study at Hospital Universiti Sains Malaysia.

BACKGROUND: Elevated serum levels of soluble PD-L1 (sPD-L1) have been reported in many cancers; however, there is limited data of sPD-L1 in breast cancer, especially those representing Asian (Malay) women. The purpose of this study was to evaluate sPD-L1 serum levels and analyze its correlation with clinical characteristics in breast cancer patients at Hospital Universiti Sains Malaysia (HUSM). METHODS: Blood specimens were obtained from 92 malignant, 16 benign breast cancer patients and 23 healthy controls. The serum concentrations of sPD-L1 were assessed by enzyme-linked immunosorbent assay (ELISA). RESULTS: The median serum sPD-L1 concentration of malignant and benign breast cancer patients was significantly elevated compared to the healthy cohorts (12.50 ng/mL vs 13.97 ng/mL vs 8.75 ng/mL, p < 0.05). Optimal cut-off value of serum sPD-L1 for predicting disease progression was 8.84  ng/mL. Elevated serum sPD-L1 levels were significantly associated with menarche age, ethnicity, birth control usage, comorbidity and HER2 status (p < 0.05). Multivariate analysis showed the menarche age and birth control were the independent factors affecting sPD-L1 expression. CONCLUSION: Elevated serum levels of sPD-L1 were significantly associated with several clinical characteristics and warrant further investigation in evaluating patients pre-diagnosed with breast cancer.

Assessing the efficacy of an innovative diagnostic method for identifying 5 % variants in somatic ctDNA.

BACKGROUND: Liquid biopsy is considered a complementary and recently also an alternative method to surgical biopsy. It allows for the acquisition of valuable information regarding the potential presence of tumors, particularly through the analysis of circulating tumor DNA (ctDNA). CtDNA is a fraction of circulating free DNA (cfDNA) that can be extracted from various tissues, with blood being the most readily available. RESULTS: To maximize the yield of plasma separation, specific Streck tubes are recommended for blood collection. The MagPurix CFC DNA Extraction Kit can be used for cfDNA extraction, and the TWIST Library Preparation protocol can be optimized for further analysis. Next-generation sequencing (NGS) can be employed to compare somatic and germline lineages, enabling the identification of somatic variants with a Variant Allele Frequency (VAF) of 5 % or higher, which are absent in the germline lineage. CONCLUSION: This analysis helps in the assessment of recurrence, analysis, and monitoring of cancer tissue.

The systematic role of pancreatic cancer exosomes: distant communication, liquid biopsy and future therapy.

Pancreatic cancer remains one of the most lethal diseases worldwide. Cancer-derived exosomes, benefiting from the protective role of the lipid membrane, exhibit remarkable stability in the circulatory system. These exosomes, released by tumor microenvironment, contain various biomolecules such as proteins, RNAs, and lipids that plays a pivotal role in mediating distant communication between the local pancreatic tumor and other organs or tissues. They facilitate the transfer of oncogenic factors to distant sites, contributing to the compromised body immune system, distant metastasis, diabetes, cachexia, and promoting a microenvironment conducive to tumor growth and metastasis in pancreatic cancer patients. Beyond their intrinsic roles, circulating exosomes in peripheral blood can be detected to facilitate accurate liquid biopsy. This approach offers a novel and promising method for the diagnosis and management of pancreatic cancer. Consequently, circulating exosomes are not only crucial mediators of systemic cell-cell communication during pancreatic cancer progression but also hold great potential as precise tools for pancreatic cancer management and treatment. Exosome-based liquid biopsy and therapy represent promising advancements in the diagnosis and treatment of pancreatic cancer. Exosomes can serve as drug delivery vehicles, enhancing the targeting and efficacy of anticancer treatments, modulating the immune system, and facilitating gene editing to suppress tumor growth. Ongoing research focuses on biomarker identification, drug delivery systems, and clinical trials to validate the safety and efficacy of exosome-based therapies, offering new possibilities for early diagnosis and precision treatment in pancreatic cancer. Leveraging the therapeutic potential of exosomes, including their ability to deliver targeted drugs and modulate immune responses, opens new avenues for innovative treatment strategies.

Translation of Epigenetics in Cell-Free DNA Liquid Biopsy Technology and Precision Oncology.

Technological advancements in cell-free DNA (cfDNA) liquid biopsy have triggered exponential growth in numerous clinical applications. While cfDNA-based liquid biopsy has made significant strides in personalizing cancer treatment, the exploration and translation of epigenetics in liquid biopsy to clinical practice is still nascent. This comprehensive review seeks to provide a broad yet in-depth narrative of the present status of epigenetics in cfDNA liquid biopsy and its associated challenges. It highlights the potential of epigenetics in cfDNA liquid biopsy technologies with the hopes of enhancing its clinical translation. The momentum of cfDNA liquid biopsy technologies in recent years has propelled epigenetics to the forefront of molecular biology. We have only begun to reveal the true potential of epigenetics in both our understanding of disease and leveraging epigenetics in the diagnostic and therapeutic domains. Recent clinical applications of epigenetics-based cfDNA liquid biopsy revolve around DNA methylation in screening and early cancer detection, leading to the development of multi-cancer early detection tests and the capability to pinpoint tissues of origin. The clinical application of epigenetics in cfDNA liquid biopsy in minimal residual disease, monitoring, and surveillance are at their initial stages. A notable advancement in fragmentation patterns analysis has created a new avenue for epigenetic biomarkers. However, the widespread application of cfDNA liquid biopsy has many challenges, including biomarker sensitivity, specificity, logistics including infrastructure and personnel, data processing, handling, results interpretation, accessibility, and cost effectiveness. Exploring and translating epigenetics in cfDNA liquid biopsy technology can transform our understanding and perception of cancer prevention and management. cfDNA liquid biopsy has great potential in precision oncology to revolutionize conventional ways of early cancer detection, monitoring residual disease, treatment response, surveillance, and drug development. Adapting the implementation of liquid biopsy workflow to the local policy worldwide and developing point-of-care testing holds great potential to overcome global cancer disparity and improve cancer outcomes.

Serum Extracellular Vesicles Cargo Approach in Bitches with Mammary Tumors.

This study investigated serum extracellular vesicles (EVs) in bitches with mammary neoplasms, in order to understand their size, shape, and concentration, as well as their association with tumor malignancy. Thirty bitches were categorized into control (n = 10), mammary tumor grades I and II (GI, n = 13), and grade III (GII, n = 7). Serum was separated from blood collected during mastectomy, and EVs were isolated using size exclusion chromatography. The analysis revealed no significant differences in EV concentrations among groups, with similar concentrations for control, GI, and GII. Ninety-one proteins were identified in EV-enriched samples, with six showing varied abundance across groups. Notably, keratin 18 was highly abundant in GI, while sushi domain-containing protein, EvC ciliary subunit 2, and the joining chain of multimeric IgM and IgA were increased in GII. Additionally, protocadherin 17 and albumin were upregulated in both GI and GII. ROC curves identified potential biomarkers for differentiating tumor grades. Enrichment pathway analysis revealed AFP gene upregulation in the GI. Mass spectrometry proteomics data were deposited in Mendeley Data. The study provides valuable insights into serum EV characterization in bitches, suggesting keratin 18 and protocadherin 17 as potential biomarkers for canine mammary neoplasia, with implications for future diagnostic and therapeutic strategies.

A meta-analysis of liquid biopsy versus tumor histology for detecting EGFR mutations in non-small cell lung cancer.

OBJECTIVE: To assess the consistency of liquid biopsy and histologic analysis for detecting epidermal growth factor receptor (EGFR) gene mutations in patients with advanced non-small cell lung cancer (NSCLC). METHODS: The PubMed, Cochrane Library, and CNKI et al. databases were searched to collect studies comparing liquid biopsy and histopathologic specimens. The EGFR mutation status was extracted from the studies, and meta-analysis was carried out using Stata 12.0 software. RESULTS: We included 22 studies of 3359 NSCLC patients. In the meta-analysis, eight papers with a sample size of size <150 had an OR of 45, indicating that liquid biopsy had high sensitivity for detecting EGFR mutations. In addition, seven papers with a sample size ≥150, with an OR of 70, reported that liquid biopsy was highly susceptible to detecting EGFR mutations. The pooled diagnostic effect size of 6 for literature that included the T790M mutation was smaller than that of 69 for literature that did not include the T790M mutation, and I2 >50 %, showing that literature that did not include the T790M mutation was more heterogeneous. The combined diagnostic effect size of 34 in the exon 19 group was smaller than that in the group with no exon 19, with an I2>50 %. There was substantial heterogeneity in both the exon 19 group and the non-exon 19 group. The group with the L858R mutation had a greater diagnostic effect size of 28, lower I2, and less heterogeneity than the group without the L858R mutation. The exon 21 group had a larger pooled diagnostic effect size of 66, a smaller I2, and less heterogeneity than the group without exon 21. CONCLUSION: Liquid biopsy and histologic analysis have high concordance for detecting EGFR mutations in NSCLC. Liquid biopsy can provide an alternative technology for individualized treatment and monitoring of minimal residual disease (MRD) in advanced NSCLC patients with EGFR tyrosine kinase inhibitor-sensitive and drug resistance (T790M) mutations.

Half-curcumin-based chemiluminescence probes and their applications in detecting quasi-stable oxidized protein.

Numerous methods have been reported for detecting ROS/RNS in vitro and in vivo; however, detecting methods for the secondary products of the ROS/RNS reactions, particularly quasi-stable oxidized products, have been much less explored. In this report, we observed that half-curcumins could generate chemiluminescence. In contrast to other chemiluminescence scaffolds, the distinguishing feature of a half-curcumin is the formation of a carbanion intermediate of its acetylacetone moiety, opening unique avenues for applications. In this study, we designed a series of half-curcumins CRANAD-Xs and found that CRANAD-164 could be used to detect quasi-stable oxidized proteins (QSOP) in vivo and in patient serum samples. We illustrated that CRANAD-164 could be used to monitor the responses of taurine, an amino acid with newly reported anti-aging capacity, in an inflammatory mouse model. Remarkably, we further demonstrated that the QSOP levels were much higher in the disease serum samples, including Alzheimer's disease, compared to the samples from healthy controls. Moreover, our results revealed that the sera chemiluminescence intensities were higher in aged healthy controls compared to young healthy subjects, suggesting that CRANAD-164 can be used to monitor the increase of QSOP during aging.

Circulating tumor extracellular vesicles to monitor metastatic prostate cancer genomics and transcriptomic evolution.

Extracellular vesicles (EVs) secreted by tumors are abundant in plasma, but their potential for interrogating the molecular features of tumors through multi-omic profiling remains widely unexplored. Genomic and transcriptomic profiling of circulating EV-DNA and EV-RNA isolated from in vitro and in vivo models of metastatic prostate cancer (mPC) reveal a high contribution of tumor material to EV-loaded DNA/RNA, validating the findings in two cohorts of longitudinal plasma samples collected from patients during androgen receptor signaling inhibitor (ARSI) or taxane-based therapy. EV-DNA genomic features recapitulate matched-patient biopsies and circulating tumor DNA (ctDNA) and associate with clinical progression. We develop a novel approach to enable transcriptomic profiling of EV-RNA (RExCuE). We report how the transcriptome of circulating EVs is enriched for tumor-associated transcripts, captures certain patient and tumor features, and reflects on-therapy tumor adaptation changes. Altogether, we show that EV profiling enables longitudinal transcriptomic and genomic profiling of mPC in liquid biopsy.

Optimized AF4 combined with density cushion ultracentrifugation enables profiling of high-purity human blood extracellular vesicles.

Extracellular vesicles (EVs) have emerged as a promising tool for clinical liquid biopsy. However, the identification of EVs derived from blood samples is hindered by the presence of abundant plasma proteins, which impairs the downstream biochemical analysis of EV-associated proteins and nucleic acids. Here, we employed optimized asymmetric flow field-flow fractionation (AF4) combined with density cushion ultracentrifugation (UC) to obtain high-purity and intact EVs with very low lipoprotein contamination from human plasma and serum. Further proteomic analysis revealed more than 1000 EV-associated proteins, a large proportion of which has not been previously reported. Specifically, we found that cell-line-derived EV markers are incompatible with the identification of plasma-EVs and proposed that the proteins MYCT1, TSPAN14, MPIG6B and MYADM, as well as the traditional EV markers CD63 and CD147, are plasma-EV markers. Benefiting from the high-purity of EVs, we conducted comprehensive miRNA profiling of plasma EVs and nanosized particles (NPs), as well as compared plasma- and serum-derived EVs, which provides a valuable resource for the EV research community. Overall, our findings provide a comprehensive assessment of human blood EVs as a basis for clinical biopsy applications.

Exosome nanovesicles: biomarkers and new strategies for treatment of human diseases.

Exosomes are nanoscale vesicles of cellular origin. One of the main characteristics of exosomes is their ability to carry a wide range of biomolecules from their parental cells, which are important mediators of intercellular communication and play an important role in physiological and pathological processes. Exosomes have the advantages of biocompatibility, low immunogenicity, and wide biodistribution. As researchers' understanding of exosomes has increased, various strategies have been proposed for their use in diagnosing and treating diseases. Here, we provide an overview of the biogenesis and composition of exosomes, describe the relationship between exosomes and disease progression, and focus on the use of exosomes as biomarkers for early screening, disease monitoring, and guiding therapy in refractory diseases such as tumors and neurodegenerative diseases. We also summarize the current applications of exosomes, especially engineered exosomes, for efficient drug delivery, targeted therapies, gene therapies, and immune vaccines. Finally, the current challenges and potential research directions for the clinical application of exosomes are also discussed. In conclusion, exosomes, as an emerging molecule that can be used in the diagnosis and treatment of diseases, combined with multidisciplinary innovative solutions, will play an important role in clinical applications.

Detection and Characterization of Methylated Circulating Tumor DNA in Gastric Cancer.

Gastric cancer is the fifth most common disease in the world and the fourth most common cause of death. It is diagnosed through esophagogastroduodenoscopy with biopsy; however, there are limitations in finding lesions in the early stages. Recently, research has been actively conducted to use liquid biopsy to diagnose various cancers, including gastric cancer. Various substances derived from cancer are reflected in the blood. By analyzing these substances, it was expected that not only the presence or absence of cancer but also the type of cancer can be diagnosed. However, the amount of these substances is extremely small, and even these have various variables depending on the characteristics of the individual or the characteristics of the cancer. To overcome these, we collected methylated DNA fragments using MeDIP and compared them with normal plasma to characterize gastric cancer tissue or patients' plasma. We attempted to diagnose gastric cancer using the characteristics of cancer reflected in the blood through the cancer tissue and patients' plasma. As a result, we confirmed that the consistency of common methylated fragments between tissue and plasma was approximately 41.2% and we found the possibility of diagnosing and characterizing cancer using the characteristics of the fragments through SFR and 5'end-motif analysis.

Liquid Biopsy in Whole Blood for Identification of Gene Expression Patterns (mRNA and miRNA) Associated with Recurrence of Glioblastoma WHO CNS Grade 4.

GBM WHO CNS Grade 4 represents a major challenge for oncology due to its aggressive behavior. Conventional imaging has restrictions in detecting tumor recurrence. This prospective study aims to identify gene-based biomarkers in whole blood instead of isolating exosomes for the early detection of tumor recurrence. Blood samples (n = 33) were collected from seven GBM patients at time points before and after surgery as well as upon tumor recurrence. Four tumor tissue samples were assessed in parallel. Next-generation sequencing (NGS), including mRNA-seq and small RNA-seq, was used to analyze gene expression profiles in blood samples and tumor tissues. A novel filtering pipeline was invented to narrow down potential candidate genes. In total, between 6-93 mRNA and 1-19 small RNA candidates could be identified among the seven patients. The overlap of genes between the patients was minimal, indicating significant inter-individual variance among GBM patients. In summary, this prospective study supports the applicability of gene expression measurements in whole blood for the detection of tumor recurrence. It might provide an alternative to the challenging workflow of liquid biopsy after laborious exosome isolation from whole blood.