Sysmex (Germany)

Torque teno virus (TTV) is proposed as a biomarker of immune function, potentially aiding in personalized immunosuppressive dosing after solid organ transplantation. TTV is thought to replicate in leukocytes, particularly immature granulocytes. We primarily aimed to assess the association between immature granulocyte count and TTV load, while also exploring associations between TTV load and other leukocyte populations. A total of 701 SOTR at 3 months post-transplantation were included from the TransplantLines Study for cross-sectional analyses. TTV load was measured by PCR assay (TTV R-GENE®, bioMérieux) and leukocytes were counted by a Sysmex XN hematology analyzer. Linear regression analyses were performed to assess associations. The mean age of the cohort was 57.0 ± 13.4 years, and 35.8% were female. In multivariable regression analyses, after adjusting for factors known to potentially influence TTV load, immature granulocyte count was positively associated with TTV load (st.β = 0.45, P < 0.001), while total granulocyte count was negatively associated (st.β=-0.39, P < 0.001). These results provide evidence to support the theory that TTV replicates in immature granulocytes. The negative association with total granulocyte count suggests possible antiviral effects or altered granulocyte lifespan. These relationships may help clinical interpretation of TTV as a marker of immune function.

Abstract Background and Aims Chronic kidney disease (CKD) is one of the fastest growing global causes of death, characterized by a gradual loss of renal function over time leading to kidney failure. Early detection of progression and treatment are key for CKD management. The rate of estimated glomerular filtration rate (eGFR) decline can vary substantially among CKD patients, ranging from stable levels to relatively rapid reduction in kidney function. Cross-sectional observational studies demonstrate an association between chronic inflammation, evidenced by elevated levels of circulating inflammatory cytokines, and lower eGFR. Emerging experimental evidence suggests that inflammatory cytokines may directly contribute to glomerular and tubular injury, promote fibrosis, and exacerbate vascular damage, collectively compromising nephron integrity function. Therefore, it is hypothesized that the inflammatory status plays a critical role in the progression of kidney damage. In this study, we examined the relationship between plasma and urinary cytokine levels and the long-term prognosis of progression in CKD. Method This study included 165 CKD patients from the original PROGRESER cohort. Plasma and urine cytokine levels were measured at baseline, 18 and 36 months using the Sysmex fully automated immunoassay analysers (HISCL-5000 and HISCL-800). Three-year follow-up clinical parameters were retrieved from the original PROGRESER case report forms, and additional 10-year follow-up data were collected from participating centers. CKD patients were categorized into three groups—non-diabetic (non-DM, N = 100), type 1 diabetes (T1D, N = 5), and type 2 diabetes (T2D, N = 60)—and compared to healthy controls (N = 30). Disease progression was calculated by dividing changes in clinical outcomes by the number of observation days. Cytokines with significant correlations to clinical parameters (eGFR, urine albumin-to-creatinine ratio -UACR-, HbA1c) were pre-selected through univariate and multivariate analyses. Multiple logistic regression analysis between KDIGO and other risk categories identified three cytokines for cluster analysis. Descriptive data were reported as medians with interquartile ranges (IQR) due to non-normal distributions. Statistical tests, including Fisher’s exact test, Steel-Dwass test, and Mann–Whitney U test, were conducted using R software (version 4.0.3). Two-tailed p-values &amp;lt; 0.05 were considered statistically significant. Results The median age was 70 years for diabetic CKD patients and 66 years for non-DM CKD patients, UACR 59.7 mg/g and 31.3 mg/g, respectively. All participants had CKD G3–G5. Plasma levels of IL-5, IL-6, IL-8, IL-17, IL-18, PROTEIN-A*, CXCL9, PROTEIN-B, BAFF, and GDF-15 were significantly elevated, whereas IL-10, CCL5, and TWEAK were reduced in non-diabetic and diabetic CKD compared to healthy controls (P &amp;lt; .05). Urinary PROTEIN-A was also significantly elevated (P &amp;lt; .05). Cluster analysis based on IL-8, PROTEIN-B, and GDF-15 identified clusters with distinct inflammatory profiles. Clusters 3 and 4, characterized by higher inflammation, had a greater proportion of patients classified as Very High Risk by KDIGO criteria and a higher prevalence of diabetic kidney disease compared to Cluster 5, which exhibited lower inflammation. Cluster 3 also demonstrated a larger increase in UACR and a higher number of all-cause deaths (P &amp;lt; 0.0001), primarily due to cardiovascular causes. Conclusion Inflammatory cytokine clusters are strongly associated with CKD progression and adverse outcomes. Patients in high-inflammation clusters (IL-8, PROTEIN-B, GDF-15) had increased KDIGO Very High-Risk classification, greater UACR progression, and higher mortality, mainly due to cardiovascular causes. These findings highlight the prognostic value of cytokine profiling in CKD management. *Names withheld for intellectual property reasons.

Severe asthma varies in inflammation and natural history. We examined biomarker profiles in patients with different asthma durations to characterise inflammatory patterns in chronic asthma. Cross-sectional biomarker data from induced sputum, serum and plasma of adult severe asthma patients in U-BIOPRED were analysed to assess associations between asthma duration and inflammatory profiles. FeNO, blood eosinophil counts, sputum cell differentials and 74 other biomarkers (18 sputum, 56 serum/plasma) were measured. Asthma duration was defined as the time from diagnosis to study entry. Multivariable linear regression models, adjusted for age, sex, ethnicity, smoking history, BMI and OCS dose, were used. Among 408 severe asthma patients (median asthma duration: 23 years, IQR 12-38), longer disease duration was associated with higher T1/17 biomarker levels (sputum neutrophil %, sputum CXCL9 and IL-6) and lower T2 biomarker levels (FeNO, sputum eosinophil %, eosinophil-derived neurotoxin (EDN)) (Fig 1). Sputum CXCL9 was inversely associated with FEV1% (coefficient: -2.50, 95% CI: -4.22, -0.78, p=0.005), whereas other T1/17 biomarkers, such as sputum neutrophil % (coefficient -0.10, 95%CI -0.21, 0.02, p=0.096), were not. <fig><object-id>erj;66/suppl_69/PA374/F1</object-id><object-id>F1</object-id><object-id>F1</object-id><graphic></graphic></fig> Sputum findings suggest longer asthma duration is associated with a higher T1/17 and lower T2 inflammatory profile, which may contribute to reduced treatment responsiveness in severe asthma.

BackgroundThis is a multicentre, European, prospective trial evaluating the diagnostic accuracy of One Step Nucleic Acid Amplification (OSNA) compared to sentinel lymph nodes histopathological ultrastaging in endometrial cancer patients.MethodsCentres with expertise in sentinel lymph node mapping in endometrial cancer patients in Europe will be invited to participate in the study. Participating units will be trained on the correct usage of the OSNA RD-210 analyser and nucleic acid amplification reagent kit LYNOAMP CK19 E for rapid detection of metastatic nodal involvement, based on the cytokeratin 19 (CK19) mRNA detection. Endometrial cancer patients ≥ 18 years listed for surgical treatment with sentinel lymph node mapping, with no history of other types of cancer and who provide a valid written consent will be considered potentially eligible for the study. However, they will only be enrolled if a successful sentinel lymph node mapping is retrieved. Each node will be processed according to the study protocol and assessed by both OSNA and ultrastaging.DiscussionThe accuracy of OSNA (index test) will be assessed against sentinel lymph node histopathological ultrastaging (reference test). This European study has the potential to be the largest study on the use of OSNA in endometrial cancer to date. OSNA could represent a modern diagnostic alternative to sentinel lymph node ultrastaging with the added benefits of standardisation and fast results.Trial registrationThe study was registered in the German Clinical Trial Register – Nr. DRKS00021520, registration date 25th of May 2020, URL of the trial registry record: https://drks.de/search/en/trial/DRKS00021520.

ABSTRACT Background Primary membranous nephropathy (PMN) is usually caused by anti-phospholipase A2 receptor (PLA2R) autoantibodies. There are different therapeutic options according to baseline risk. Novel biomarkers are needed to optimize risk stratification and predict and monitor the response to therapy, as proteinuria responses may be delayed. We hypothesized that plasma or urinary cytokines may provide insights into the course and response to therapy in PMN. Methods Overall, 192 data points from 34 participants in the STARMEN trial (NCT01955187), randomized to tacrolimus–rituximab (TAC-RTX) or corticosteroids–cyclophosphamide (GC-CYC), were analysed for plasma and urine cytokines using a highly sensitive chemiluminescence immunoassay providing a high-throughput multiplex analysis. Results Baseline (pretreatment) urinary C-X-C motif chemokine ligand 13 (CXCL13) predicted the therapeutic response to TAC-RTX. Cytokine levels evolved over the course of therapy. The levels of nine plasma and six urinary cytokines correlated with analytical parameters of kidney damage and disease activity, such as proteinuria, estimated glomerular filtration rate and circulating anti-PLA2R levels. The correlation with these parameters was most consistent for plasma and urinary growth differentiation factor 15 (GDF15), plasma tumour necrosis factor α and urinary TNF-like weak inducer of apoptosis. Decreasing plasma GDF15 levels were associated with response to GC-CYC. Four clusters of cytokines were associated with different stages of response to therapy in the full cohort, with the less inflammatory cluster associated with remission. Conclusion PMN displayed characteristic plasma and urine cytokine patterns that evolved over time as patients responded to therapy. Baseline urinary CXCL13 concentration could be a prognostic marker of response to TAC-RTX.

BackgroundSepsis occurs as a result of dysregulated host response to infection. However, cytokine adsorption therapy may restore the balance of proinflammatory and anti-inflammatory mediator responses in patients with sepsis. This study aimed to determine the cytokine adsorption ability of two different types of continuous renal replacement therapy (CRRT) hemofilters for polyethyleneimine-coated polyacrylonitrile (AN69ST) (surface-treated) and polymethylmethacrylate (PMMA) CRRT.MethodsWe performed a randomized controlled trial among sepsis patients undergoing CRRT, who were randomly assigned (1:1) to receive either AN69ST or PMMA-CRRT. The primary outcome was cytokine clearance of hemofilter adsorption (CHA). The secondary endpoints were the intensive care unit (ICU) and 28-day mortalities.ResultsWe randomly selected 52 patients. Primary outcome data were available for 26 patients each in the AN69ST-CRRT and PMMA-CRRT arms. The CHA of high-mobility group box 1, tumor necrosis factor, interleukin (IL)-8, monokine induced by interferon-γ, and macrophage inflammatory protein were significantly higher in the AN69ST-CRRT group than in the PMMA-CRRT group (P < 0.001, P < 0.01, P < 0.001, P < 0.001 and P < 0.001, respectively). In contrast, the CHA of IL-6 was significantly higher in the PMMA-CRRT group than in the AN69ST-CRRT group (P < 0.001). In addition, the 28-day mortality was not significantly different between the two groups (50% in AN69ST-CRRT vs. 30.8% in PMMA-CRRT, P = 0.26).ConclusionAN69ST and PMMA membranes have different cytokine CHA in patients with sepsis. Therefore, these two hemofilters may have to be used depending on the target cytokine.Trial registration number: This study was registered in the University Hospital Medical Information Network on November 1, 2017 (Trial No: UMIN000029450, https://center6.umin.ac.jp).

Even during long-term combination antiretroviral therapy (cART), people living with HIV (PLHIV) have a dysregulated immune system, characterized by persistent immune activation, accelerated immune ageing and increased risk of non-AIDS comorbidities. A multi-omics approach is applied to a large cohort of PLHIV to understand pathways underlying these dysregulations in order to identify new biomarkers and novel genetically validated therapeutic drugs targets. The 2000HIV study is a prospective longitudinal cohort study of PLHIV on cART. In addition, untreated HIV spontaneous controllers were recruited. In-depth multi-omics characterization will be performed, including genomics, epigenomics, transcriptomics, proteomics, metabolomics and metagenomics, functional immunological assays and extensive immunophenotyping. Furthermore, the latent viral reservoir will be assessed through cell associated HIV-1 RNA and DNA, and full-length individual proviral sequencing on a subset. Clinical measurements include an ECG, carotid intima-media thickness and plaque measurement, hepatic steatosis and fibrosis measurement as well as psychological symptoms and recreational drug questionnaires. Additionally, considering the developing pandemic, COVID-19 history and vaccination was recorded. Participants return for a two-year follow-up visit. The 2000HIV study consists of a discovery and validation cohort collected at separate sites to immediately validate any finding in an independent cohort. Overall, 1895 PLHIV from four sites were included for analysis, 1559 in the discovery and 336 in the validation cohort. The study population was representative of a Western European HIV population, including 288 (15.2%) cis-women, 463 (24.4%) non-whites, and 1360 (71.8%) MSM (Men who have Sex with Men). Extreme phenotypes included 114 spontaneous controllers, 81 rapid progressors and 162 immunological non-responders. According to the Framingham score 321 (16.9%) had a cardiovascular risk of >20% in the next 10 years. COVID-19 infection was documented in 234 (12.3%) participants and 474 (25.0%) individuals had received a COVID-19 vaccine. The 2000HIV study established a cohort of 1895 PLHIV that employs multi-omics to discover new biological pathways and biomarkers to unravel non-AIDS comorbidities, extreme phenotypes and the latent viral reservoir that impact the health of PLHIV. The ultimate goal is to contribute to a more personalized approach to the best standard of care and a potential cure for PLHIV.

Aims The yield of colonoscopy surveillance after colorectal cancer screening is limited. To lower patient burden and healthcare costs, there is a need to reduce colonoscopies in which no advanced neoplasia is detected. The MOlecular stool testing for Colorectal CAncer Surveillance (MOCCAS) study evaluated whether stool-based testing could safely reduce the number of surveillance colonoscopies.

Coronavirus disease 2019 (COVID-19) is an acute respiratory disease; approximately 5% of patients developing severe COVID-19. It is known that cytokine release is associated with disease severity, but the relationship between the different clinical phenotypes and inflammatory endotypes is not well understood. This study investigated the association between inflammatory biomarker-based endotypes and severe COVID-19 phenotypes. Interleukin (IL) -6, C-reactive protein (CRP), C-X-C motif chemokine (CXCL) 9, IL-18, C-C motif chemokine (CCL) 3, CCL17, IL-10, and vascular endothelial growth factor (VEGF) were measured in 57 COVID-19 patients, and their association with clinical characteristics was examined using a cluster analysis. Significantly higher blood levels of the eight inflammatory markers were noted in patients who developed acute respiratory distress syndrome (ARDS) than in those who did not develop ARDS (non-ARDS). Using a cluster analysis, the patient groups were classified into four clusters, of which two had patients with high IL-6 and CRP levels. In the cluster with high levels of Type 1 (T1) inflammatory markers such as CXCL9 and IL-18, 85% of the patients had ARDS, 65% of the patients developed acute kidney injury (AKI), and 78% of the patients developed pulmonary fibrosis. In the cluster with high levels of T1 inflammatory markers, the patients frequently suffered from tissue damage, manifested as ARDS and AKI. Our findings identified distinct T1 inflammatory endotypes of COVID-19 and suggest the importance of controlling inflammation by monitoring T1 biomarkers and treating accordingly to limit the severity of the disease.

Circulating tumor DNA (ctDNA) has emerged as a non-invasive “liquid biopsy” for early breast cancer diagnosis. We evaluated the suitability of ctDNA analysis in the diagnosis of early breast cancer after mammography findings, comparing PIK3CA and TP53 mutations between tumor biopsies and pre-biopsy circulating DNA. Matched plasma and frozen fresh tissue biopsies from patients with Breast Imaging-Reporting and Data System (BIRADS) 4c/5 mammography findings and subsequent diagnosis of primary breast cancer were analyzed using NGS TruSeq Custom Amplicon Low Input Panel (Illumina) and plasma SafeSEQ (Sysmex Inostics). The same plasma and tumor mutations were observed in eight of 29 patients (27.6%) with four in TP53 and five in PIK3CA mutations. Sequencing analysis also revealed four additional ctDNA mutations (three in TP53 and one in PIK3CA) previously not identified in three patients tissue biopsy. One of these patients had mutations in both genes. Age, tumor grade and size, immunohistochemical (IHC) subtype, BIRADS category, and lymph node positivity were significantly associated with the detectability of these blood tumor-derived mutations. In conclusion, ctDNA analysis could be used in early breast cancer diagnosis, providing critical clinical information to improve patient diagnosis.