MiRNAs in Extracellular Vesicles as Biomarkers in Plasma of Papillary Thyroid Cancer Patients: A Proof-of-Concept Study.

BACKGROUND: The incidence of various types of cancer, for example, papillary thyroid carcinoma (PTC), is on the rise. Since therapeutic success depends greatly on early diagnosis, reliable diagnostic biomarkers must be identified, and easy-to-apply tools for detecting them must urgently be standardized. Here, we contribute to solving this medical challenge by assessing miRNAs suspected of promoting carcinogenesis in extracellular vesicles (EVs) that can be routinely obtained via liquid biopsy. We profit from current progress in cancerology that provides innovations in liquid biopsy and EVs analysis, along with the identification of miRNAs and chaperone system (CS) components implicated in carcinogenesis. METHODS: We measured in EVs obtained from circulating blood plasma from PTC patients the levels of three miRNAs implicated in thyroid cancer, hsa-miR-1-3p, hsa-miR-206, and hsa-miR-221-3p, and most likely involved in the regulation of two members of the CS, Hsp60 and CCT. EVs were isolated from the plasma of patients with PTC and controls with benign goiter (BG) and from the culture medium of a PTC cell line (MDAT32) and were appropriately characterized. RESULTS: The levels of miRNAs determined by RT-qPCR were consistently higher in PTC patients and decreased down to control levels after thyroidectomy. Bioinformatics showed that the miRNAs target genes are associated with the molecular pathogenesis of PTC. CONCLUSIONS: Our exploratory study reaffirms the potential in clinics of the selected miRNAs in EVs as useful biomarkers of PTC easily accessible via liquid biopsy, which is minimally invasive and amenable to periodic repetition, an improvement compared to the established fine-needle aspirate biopsy.

Digital biomarkers: 3PM approach revolutionizing chronic disease management - EPMA 2024 position.

Non-communicable chronic diseases (NCDs) have become a major global health concern. They constitute the leading cause of disabilities, increased morbidity, mortality, and socio-economic disasters worldwide. Medical condition-specific digital biomarker (DB) panels have emerged as valuable tools to manage NCDs. DBs refer to the measurable and quantifiable physiological, behavioral, and environmental parameters collected for an individual through innovative digital health technologies, including wearables, smart devices, and medical sensors. By leveraging digital technologies, healthcare providers can gather real-time data and insights, enabling them to deliver more proactive and tailored interventions to individuals at risk and patients diagnosed with NCDs. Continuous monitoring of relevant health parameters through wearable devices or smartphone applications allows patients and clinicians to track the progression of NCDs in real time. With the introduction of digital biomarker monitoring (DBM), a new quality of primary and secondary healthcare is being offered with promising opportunities for health risk assessment and protection against health-to-disease transitions in vulnerable sub-populations. DBM enables healthcare providers to take the most cost-effective targeted preventive measures, to detect disease developments early, and to introduce personalized interventions. Consequently, they benefit the quality of life (QoL) of affected individuals, healthcare economy, and society at large. DBM is instrumental for the paradigm shift from reactive medical services to 3PM approach promoted by the European Association for Predictive, Preventive, and Personalized Medicine (EPMA) involving 3PM experts from 55 countries worldwide. This position manuscript consolidates multi-professional expertise in the area, demonstrating clinically relevant examples and providing the roadmap for implementing 3PM concepts facilitated through DBs.

Immune-related gene methylation prognostic instrument for stratification and targeted treatment of ovarian cancer patients toward advanced 3PM approach.

Background: DNA methylation is an important mechanism in epigenetics, which can change the transcription ability of genes and is closely related to the pathogenesis of ovarian cancer (OC). We hypothesize that DNA methylation is significantly different in OCs compared to controls. Specific DNA methylation status can be used as a biomarker of OC, and targeted drugs targeting these methylation patterns and DNA methyltransferase may have better therapeutic effects. Studying the key DNA methylation sites of immune-related genes (IRGs) in OC patients and studying the effects of these methylation sites on the immune microenvironment may provide a new method for further exploring the pathogenesis of OC, realizing early detection and effective monitoring of OC, identifying effective biomarkers of DNA methylation subtypes and drug targets, improving the efficacy of targeted drugs or overcoming drug resistance, and better applying it to predictive diagnosis, prevention, and personalized medicine (PPPM; 3PM) of OC. Method: Hypermethylated subtypes (cluster 1) and hypomethylated subtypes (cluster 2) were established in OCs based on the abundance of different methylation sites in IRGs. The differences in immune score, immune checkpoints, immune cells, and overall survival were analyzed between different methylation subtypes in OC samples. The significant pathways, gene ontology (GO), and protein-protein interaction (PPI) network of the identified methylation sites in IRGs were enriched. In addition, the immune-related methylation signature was constructed with multiple regression analysis. A methylation site model based on IRGs was constructed and verified. Results: A total of 120 IRGs with 142 differentially methylated sites (DMSs) were identified. The DMSs were clustered into a high-level methylation group (cluster 1) and a low-level methylation group (cluster 2). The significant pathways and GO analysis showed many immune-related and cancer-associated enrichments. A methylation site signature based on IRGs was constructed, including RORC|cg25112191, S100A13|cg14467840, TNF|cg04425624, RLN2|cg03679581, and IL1RL2|cg22797169. The methylation sites of all five genes showed hypomethylation in OC, and there were statistically significant differences among RORC|cg25112191, S100A13|cg14467840, and TNF|cg04425624 (p < 0.05). This prognostic model based on low-level methylation and high-level methylation groups was significantly linked to the immune microenvironment as well as overall survival in OC. Conclusions: This study provided different methylation subtypes for OC patients according to the methylation sites of IRGs. In addition, it helps establish a relationship between methylation and the immune microenvironment, which showed specific differences in biological signaling pathways, genomic changes, and immune mechanisms within the two subgroups. These data provide ones to deeply understand the mechanism of immune-related methylation genes on the occurrence and development of OC. The methylation-site signature is also to establish new possibilities for OC therapy. These data are a precious resource for stratification and targeted treatment of OC patients toward an advanced 3PM approach. Supplementary Information: The online version contains supplementary material available at 10.1007/s13167-024-00359-3.

Copy Number Variations as Determinants of Colorectal Tumor Progression in Liquid Biopsies.

Over the years, increasing evidence has shown that copy number variations (CNVs) play an important role in the pathogenesis and prognosis of Colorectal Cancer (CRC). Colorectal adenomas are highly prevalent lesions, but only 5% of these adenomas ever progress to carcinoma. This review summarizes the different CNVs associated with adenoma-carcinoma CRC progression and with CRC staging. Characterization of CNVs in circulating free-RNA and in blood-derived exosomes augers well with the potential of using such assays for patient management and early detection of metastasis. To overcome the limitations related to tissue biopsies and tumor heterogeneity, using CNVs to characterize tumor-derived materials in biofluids provides less invasive sampling methods and a sample that collectively represents multiple tumor sites in heterogeneous samples. Liquid biopsies provide a source of circulating tumor DNA (ctDNA), circulating tumor cells (CTCs), tumor-derived exosomes (TDE), circulating free RNA, and non-coding RNA. This review provides an overview of the current diagnostic and predictive models from liquid biopsies.

PKC-mediated phosphorylation and activation of the MEK/ERK pathway as a mechanism of acquired trastuzumab resistance in HER2-positive breast cancer.

Protein expression, activation and stability are regulated through inter-connected signal transduction pathways resulting in specific cellular states. This study sought to differentiate between the complex mechanisms of intrinsic and acquired trastuzumab resistance, by quantifying changes in expression and activity of proteins (phospho-protein profile) in key signal transduction pathways, in breast cancer cellular models of trastuzumab resistance. To this effect, we utilized a multiplex, bead-based protein assay, DigiWest®, to measure around 100 proteins and protein modifications using specific antibodies. The main advantage of this methodology is the quantification of multiple analytes in one sample, utilising input volumes of a normal western blot. The intrinsically trastuzumab-resistant cell line JIMT-1 showed the largest number of concurrent resistance mechanisms, including PI3K/Akt and RAS/RAF/MEK/ERK activation, ß catenin stabilization by inhibitory phosphorylation of GSK3ß, cell cycle progression by Rb suppression, and CREB-mediated cell survival. MAPK (ERK) pathway activation was common to both intrinsic and acquired resistance cellular models. The overexpression of upstream RAS/RAF, however, was confined to JIMT 1; meanwhile, in a cellular model of acquired trastuzumab resistance generated in this study (T15), entry into the ERK pathway seemed to be mostly mediated by PKCα activation. This is a novel observation and merits further investigation that can lead to new therapeutic combinations in HER2-positive breast cancer with acquired therapeutic resistance.

A randomized, controlled trial on the effect of anesthesia on chronic pain after total knee arthroplasty.

Aim: The study sought to evaluate the influence of anesthesia on chronic pain after total knee arthroplasty (TKA). Methods: This was a single-center, randomized controlled study, with patients receiving a spinal anesthetic (SP) alone or a general anesthetic (GA) with femoral block, with follow-up at 3 and at 6 months. The primary outcome was the WOMAC® score at 6 months. Results: 199 patients were enrolled. Group SP had better function (WOMAC: GA: 16.9 vs SP: 14.4, p = 0.015) and less pain (WOMAC pain: GA: 3.04 vs SP: 2.69, p = 0.02) at 3 months, but not at 6 months. Overall, 11% of patients had chronic postsurgical pain (CPSP), with Group GA having a higher incidence of CPSP at 6 months. Neuropathic pain increased during the follow-up and was more common in patients with CPSP. Conclusion: An SP reduces pain and incidence of CPSP after TKA. Clinical Trial Registration: NCT04206046 (ClinicalTrials.gov).

Patients who have knee replacement surgery are at risk of remaining in significant pain even up to one year after surgery. Our study aims to look into the potential of reducing this risk by comparing two common forms of anesthesia. 199 patients were randomized to either have a spinal anesthetic (SP) or a general anesthetic with a femoral nerve block. Patients were then followed-up for 6 months, and then asked to report on a validated functional and pain scoring system (WOMAC^®). Patients who had had an SP had better function and less pain at 3 months after surgery, but not at 6 months. The incidence of chronic pain at six month seems to be less with an SP.

Differential Expression of the Sphingolipid Pathway Is Associated with Sensitivity to the PP2A Activator FTY720 in Colorectal Cancer Cell Lines.

Protein phosphatase 2A (PP2A) is a ubiquitously expressed intracellular serine/threonine phosphatase. Deregulation of PP2A is a common event associated with adenocarcinomas of the colon and rectum. We have previously shown that breast cancer cell lines are sensitive to the PP2A activator FTY720, and that sensitivity is predicted by high Aurora kinase A (AURKA) mRNA expression. In this study, we hypothesized that high relative AURKA expression could predict sensitivity to FTY720-induced apoptosis in colorectal cancer (CRC). The CRC cell lines NCI H716, COLO320DM, DLD-1, SW480, and HT-29 show a high relative AURKA expression as compared to LS411N, T84, HCT116, SW48, and LOVO. Following viability assays, LS411N, T84, HCT116, and SW480 were shown to be sensitive to FTY720, whereas DLD-1 and HT-29 were non-sensitive. Hence, AURKA mRNA expression does not predict sensitivity to FTY720 in CRC cell lines. Differentially expressed genes (DEGs) were obtained by comparing the sensitive CRC cell lines (LS411N and HCT116) against the non-sensitive (HT-29 and DLD-1). We found that 253 genes were significantly altered in expression, and upregulation of CERS4, PPP2R2C, GNAZ, PRKCG, BCL2, MAPK12, and MAPK11 suggests the involvement of the sphingolipid signaling pathway, known to be activated by phosphorylated-FTY720. In conclusion, although AURKA expression did not predict sensitivity to FTY720, it is evident that specific CRC cell lines are sensitive to 5 µM FTY720, potentially because of the differential expression of genes involved in the sphingolipid pathway.

Somatic copy number aberrations in metastatic patients: The promise of liquid biopsies.

Cancer metastasis is the leading cause of cancer-related mortality. The metastatic process involves measurable cellular changes that confer migratory potential, proliferative advantage and the ability to colonise a distinct microenvironment. Accumulation of aberrations and clonal evolution add complexity to patient management and the assessment of the therapeutic sensitivity profile of malignancies. Liquid biopsy presents a repeatable and minimally invasive assessment tool to detect early metastasis, characterise tumour phenotype and detect minimal residual disease. The promise of liquid biopsies is to inform patient management and therapeutic decisions in a timely manner. Clinical translation requires robust methodologies with high sensitivity and tumour specificity. This can be achieved through technological advances but also through novel biologically informed approaches that harness existing knowledge on tumorigenesis. Here we present a review of copy number variations as potential biomarkers for early detection of metastatic potential and outline a biomarker validation process in the context of liquid biopsies.

A loop involving NRF2, miR-29b-1-5p and AKT, regulates cell fate of MDA-MB-231 triple-negative breast cancer cells.

The present study shows that nuclear factor erythroid 2-related factor 2 (NRF2) and miR-29b-1-5p are two opposite forces which could regulate the fate of MDA-MB-231 cells, the most studied triple-negative breast cancer (TNBC) cell line. We show that NRF2 activation stimulates cell growth and markedly reduces reactive oxygen species (ROS) generation, whereas miR-29b-1-5p overexpression increases ROS generation and reduces cell proliferation. Moreover, NRF2 downregulates miR-29b-1-5p expression, whereas miR-29b-1-5p overexpression decreases p-AKT and p-NRF2. Furthermore, miR-29b-1-5p overexpression induces both inhibition of DNA N-methyltransferases (DNMT1, DNMT3A, and DNMT3B) expression and re-expression of HIN1, RASSF1A and CCND2. Conversely, NRF2 activation induces opposite effects. We also show that parthenolide, a naturally occurring small molecule, induces the expression of miR-29b-1-5p which could suppress NRF2 activation via AKT inhibition. Overall, this study uncovers a novel NRF2/miR-29b-1-5p/AKT regulatory loop that can regulate the fate (life/death) of MDA-MB-231 cells and suggests this loop as therapeutic target for TNBC.

The landscape of genomic copy number alterations in colorectal cancer and their consequences on gene expression levels and disease outcome.

Aneuploidy, the unbalanced state of the chromosome content, represents a hallmark of most solid tumors, including colorectal cancer. Such aneuploidies result in tumor specific genomic imbalances, which emerge in premalignant precursor lesions. Moreover, increasing levels of chromosomal instability have been observed in adenocarcinomas and are maintained in distant metastases. A number of studies have systematically integrated copy number alterations with gene expression changes in primary carcinomas, cell lines, and experimental models of aneuploidy. In fact, chromosomal aneuploidies target a number of genes conferring a selective advantage for the metabolism of the cancer cell. Copy number alterations not only have a positive correlation with expression changes of the majority of genes on the altered genomic segment, but also have effects on the transcriptional levels of genes genome-wide. Finally, copy number alterations have been associated with disease outcome; nevertheless, the translational applicability in clinical practice requires further studies. Here, we (i) review the spectrum of genetic alterations that lead to colorectal cancer, (ii) describe the most frequent copy number alterations at different stages of colorectal carcinogenesis, (iii) exemplify their positive correlation with gene expression levels, and (iv) discuss copy number alterations that are potentially involved in disease outcome of individual patients.