Extended interval dosing with ocrelizumab in multiple sclerosis.

BACKGROUND: This study investigates clinical and biomarker differences between standard interval dosing (SID) and extended interval dosing (EID) of ocrelizumab therapy in multiple sclerosis (MS). METHODS: This is a prospective, double-arm, open-label, multi-center study in Denmark. Participants diagnosed with MS on ocrelizumab therapy >12 months were included (n = 184). Clinical, radiological, and blood-based biomarker outcomes were evaluated. MRI disease activity, relapses, worsening of neurostatus, and No Evidence of Disease Activity-3 (NEDA-3) were used as a combined endpoint. RESULTS: Out of 184 participants, 107 participants received EID (58.2%), whereas 77 participants received SID (41.8%). The average extension was 9 weeks with a maximum of 78 weeks. When comparing EID to SID, we found higher levels of B-cells, lower serum concentrations of ocrelizumab, and similar levels of age-adjusted NFL and GFAP in the two groups. No difference in NEDA-3 between EID and SID was demonstrated (hazard ratio: 1.174, p = 0.69). Higher levels of NFL were identified in participants with disease activity. Body mass index correlated with levels of ocrelizumab and B-cells. CONCLUSION: Extending one treatment interval of ocrelizumab on average 9 weeks and up to 78 weeks did not result in clinical, radiological, or biomarker evidence of worsening compared with SID.

Proteomic analysis of diabetic retinopathy identifies potential plasma-protein biomarkers for diagnosis and prognosis.

OBJECTIVES: To identify molecular pathways and prognostic- and diagnostic plasma-protein biomarkers for diabetic retinopathy at various stages. METHODS: This exploratory, cross-sectional proteomics study involved plasma from 68 adults, including 15 healthy controls and 53 diabetes patients for various stages of diabetic retinopathy: non-diabetic retinopathy, non-proliferative diabetic retinopathy, proliferative diabetic retinopathy and diabetic macular edema. Plasma was incubated with peptide library beads and eluted proteins were tryptic digested, analyzed by liquid chromatography-tandem mass-spectrometry followed by bioinformatics. RESULTS: In the 68 samples, 248 of the 731 identified plasma-proteins were present in all samples. Analysis of variance showed differential expression of 58 proteins across the five disease subgroups. Protein-Protein Interaction network (STRING) showed enrichment of various pathways during the diabetic stages. In addition, stage-specific driver proteins were detected for early and advanced diabetic retinopathy. Hierarchical clustering showed distinct protein profiles according to disease severity and disease type. CONCLUSIONS: Molecular pathways in the cholesterol metabolism, complement system, and coagulation cascade were enriched in patients at various stages of diabetic retinopathy. The peroxisome proliferator-activated receptor signaling pathway and systemic lupus erythematosus pathways were enriched in early diabetic retinopathy. Stage-specific proteins for early - and advanced diabetic retinopathy as determined herein could be 'key' players in driving disease development and potential 'target' proteins for future therapies. For type 1 and 2 diabetes mellitus, the proteomic profiles were especially distinct during the early disease stage. Validation studies should aim to clarify the role of the detected molecular pathways, potential biomarkers, and potential 'target' proteins for future therapies in diabetic retinopathy.

Performance Measures and Plasma Biomarker Levels in Patients with Multiple Sclerosis after 14 Days of Fampridine Treatment: An Explorative Study.

Peripheral cytokine levels may serve as biomarkers for treatment response and disease monitoring in patients with multiple sclerosis (pwMS). The objectives were to assess changes in plasma biomarkers in PwMS after 14 days of fampridine treatment and to explore correlations between changes in performance measures and plasma biomarkers. We included 27 PwMS, 14 women and 13 men, aged 52.0 ± 11.6 years, with a disease duration of 17 ± 8.5 years, and an Expanded Disability Status Scale of 6 [IQR 5.0/6.5]. Gait and hand function were assessed using performance tests completed prior to fampridine and after 14 days of treatment. Venous blood was obtained, and chemiluminescence analysis conducted to assess plasma cytokines and neurodegenerative markers. All performance measures demonstrated improvements. Biomarkers showed decreased tumor necrosis factor (TNF) receptor-2 levels. Associations were found between change scores in (i) Six Spot Step Test and Interleukin (IL)-2, IL-8, and IL-17 levels; (ii) timed 25-foot walk and interferon-γ, IL-2, IL-8, TNF-α, and neurofilament light levels, and (iii) 12-Item Multiple Sclerosis Walking Scale and IL-17 levels. The associations may reflect increased MS-related inflammatory activity rather than a fampridine-induced response or that a higher level of inflammation induces a better response to fampridine.

Serum metabolic signatures for Alzheimer's Disease reveal alterations in amino acid composition: a validation study.

INTRODUCTION: Alzheimer's Disease (AD) is complex and novel approaches are urgently needed to aid in diagnosis. Blood is frequently used as a source for biomarkers; however, its complexity prevents proper detection. The analytical power of metabolomics, coupled with statistical tools, can assist in reducing this complexity. OBJECTIVES: Thus, we sought to validate a previously proposed panel of metabolic blood-based biomarkers for AD and expand our understanding of the pathological mechanisms involved in AD that are reflected in the blood. METHODS: In the validation cohort serum and plasma were collected from 25 AD patients and 25 healthy controls. Serum was analysed for metabolites using nuclear magnetic resonance (NMR) spectroscopy, while plasma was tested for markers of neuronal damage and AD hallmark proteins using single molecule array (SIMOA). RESULTS: The diagnostic performance of the metabolite biomarker panel was confirmed using sparse-partial least squares discriminant analysis (sPLS-DA) with an area under the curve (AUC) of 0.73 (95% confidence interval: 0.59-0.87). Pyruvic acid and valine were consistently reduced in the discovery and validation cohorts. Pathway analysis of significantly altered metabolites in the validation set revealed that they are involved in branched-chain amino acids (BCAAs) and energy metabolism (glycolysis and gluconeogenesis). Additionally, strong positive correlations were observed for valine and isoleucine between cerebrospinal fluid p-tau and t-tau. CONCLUSIONS: Our proposed panel of metabolites was successfully validated using a combined approach of NMR and sPLS-DA. It was discovered that cognitive-impairment-related metabolites belong to BCAAs and are involved in energy metabolism.

Cytochrome P450 activity in rheumatoid arthritis patients during continuous IL-6 receptor antagonist therapy.

BACKGROUND: Inflammation suppresses cytochrome P450 (CYP) enzyme activity, and single-dose interleukin 6 receptor antagonists (anti-IL-6R) reverse this effect. Here, we assess the impact of continuous anti-IL-6R therapy in patients with rheumatoid arthritis. METHODS: In a clinical pharmacokinetic trial, the Basel cocktail was administered before and after 3 and 12 weeks of anti-IL-6R therapy to assess CYP enzyme activity (registered in the ClinicalTrials.gov database (identifier NCT04842981) on April 13th, 2021). In a retrospective study, the 4ß-hydroxycholesterol/cholesterol ratio was measured as a biomarker for CYP3A4 activity before and after 3 and 6 months of anti-IL-6R therapy. The control group was patients initiating a tumor necrosis factor alfa (TNF-α) inhibitor. RESULTS: In the clinical pharmacokinetic trial (n = 3), midazolam metabolic ratio (CYP3A4) was inconclusive due to the limited sample size. Midazolam AUC and Cmax indicate a weak impact on CYP3A4 activity after 3 weeks of anti-IL-6R therapy compared to baseline (AUC geometric mean ratio (GMR): 0.80, 95% CI: 0.64-0.99 and Cmax GMR: 0.58, 95% CI: 0.37-0.91), which returns to baseline levels after 12 weeks of therapy (AUC GMR 1.02, 95% CI: 0.72-1.46 and Cmax GMR 1.03, 95% CI 0.72-1.47). No effect on the 4ß-hydroxycholesterol/cholesterol ratio was observed in the retrospective study. CONCLUSION: Based on sparse data from three patients, continuous anti-IL-6R therapy seems to cause an acute but transient increase in CYP3A4 activity in rheumatoid arthritis patients, which may be due to a normalization of the inflammation-suppressed CYP activity. Further studies are warranted to understand the mechanism behind this putative transient effect. Trial registration Registered in the ClinicalTrials.gov database (identifier NCT04842981) on April 13th, 2021.

Initiation of glucose-lowering drugs reduces the anticoagulant effect of warfarin-But not through altered drug metabolism in patients with type 2 diabetes.

AIMS: Drug metabolism might be altered in patients with type 2 diabetes. We aimed to evaluate if initiation of glucose-lowering drugs impacts warfarin efficacy and drug metabolism. METHODS: First, we conducted a register-based self-controlled cohort study on Danish and Scottish warfarin users. Warfarin efficacy (international normalized ratio [INR]) was compared before and after initiation of glucose-lowering drugs. Second, we conducted a clinical pharmacokinetic trial comprising treatment-naïve type 2 diabetes patients. Patients ingested probe drugs for drug-metabolizing enzymes (the Basel Cocktail) before initiating glucose-lowering treatment, and after 3 and 12 weeks of treatment. Drug metabolism, glycaemic control, and inflammation were assessed on each visit. RESULTS: In the Danish and Scottish cohorts (n = 982 and n = 44, respectively), initiating glucose-lowering drugs reduced warfarin efficacy. INR decreased from 2.47 to 2.21 in the Danish cohort (mean difference -0.26; 95% CI -0.35; -0.17) and from 2.33 to 2.13 in the Scottish cohort (-0.21; 95% CI -0.52; 0.11) after initiation of glucose-lowering treatment. This impact on INR was more pronounced among individuals with stronger effects of glucose-lowering treatment. In the clinical pharmacokinetic trial (n = 10), initiating metformin did not affect drug metabolism after 3 weeks (geometric mean ratio of CYP3A metabolic ratio: 1.12 [95% CI: 0.95; 1.32]) or 12 weeks of metformin treatment. Glycaemic control improved during treatment, while inflammation remained low and unchanged during treatment. CONCLUSIONS: In conclusion, initiation of glucose-lowering drugs among chronic warfarin users seems associated with a reduction in INR, particularly among individuals with a large decrease in HbA1c . This effect seems unrelated to CYP enzyme activity and warfarin drug metabolism.

Neurofilament light chain as a biomarker of axonal damage in sensory neurons and paclitaxel-induced peripheral neuropathy in patients with ovarian cancer.

ABSTRACT: Paclitaxel-induced peripheral neuropathy (PIPN) is a barrier to effective cancer treatment and impacts quality of life among patients with cancer. We used a translational approach to assess the utility of neurofilament light chain (NFL) as a biomarker of PIPN in a human cell model and in patients with ovarian cancer. We measured NFL in medium from human induced pluripotent stem cell-derived sensory neurons (iPSC-SNs) exposed to paclitaxel. Serum NFL (sNFL) levels were quantified in 190 patients with ovarian cancer receiving paclitaxel/carboplatin chemotherapy at baseline and after each of the following 2 or 6 cycles. Adverse outcomes related to PIPN were retrospectively obtained, and Cox regression model was performed with different sNFL cut-offs after first cycle. The apparent elimination half-life of sNFL was estimated in patients who discontinued paclitaxel. Paclitaxel neurotoxicity in iPSC-SNs was accompanied by NFL release in a concentration-dependent manner ( P < 0.001, analysis of variance). Serum NFL levels increased substantially in patients during paclitaxel/carboplatin chemotherapy with considerable interindividual variability. Patients with sNFL >150 pg/mL after first cycle had increased risk to discontinue paclitaxel early (unadjusted HR: 2.47 [95% CI 1.16-5.22], adjusted HR: 2.25 [95% CI: 0.88-5.79]). Similar trends were shown for risk of severe PIPN and paclitaxel dose reduction because of PIPN. The median elimination half-life of sNFL was 43 days (IQR 27-82 days). Neurofilament light chain constitutes an objective biomarker of neurotoxicity in iPSC-SNs and in ovarian cancer patients with high sNFL predicting PIPN-related adverse outcomes. If prospectively validated, NFL can be used to study PIPN and may guide clinical decision making and personalize treatment with paclitaxel.

Elevated Levels of PGE2-Metabolite in Cerebrospinal Fluid and Cox-2 Gene Polymorphisms in Patients with Chronic, Post Cholecystectomy Pain and Visceral Hyperalgesia Compared to Healthy Controls. A Hypothesis-Generating Pilot Study.

Purpose: Chronic, abdominal pain remains a problem in a subset of patients after cholecystectomy. The cause is often obscure but central sensitization may be an important component and could theoretically be mediated by spinal PGE2, which is regulated by several cytokines. The aim of the study was to examine cerebrospinal fluid (CSF) of participants with post cholecystectomy syndrome and healthy volunteers for signs of PGE2 and cytokine mediated central sensitization. Patients and Methods: In phase 1 of the study, 83 subjects were included for DNA analysis, eight of these subjects with post cholecystectomy syndrome. We examined the SNPs rs5275, rs16944 and rs1800795 from the Cox-2, IL-1ß and IL-6 genes respectively. In phase 2 of the study, we examined concentrations of PGE2-metabolite (PGEM), IL-1ß and IL-6 in CSF and plasma from 6 patients with post cholecystectomy syndrome and visceral hyperalgesia and 11 pain free volunteers. Results: We found a significant difference in distribution of the rs5275 SNP of the Cox-2 enzyme (CT-genotype=88% in pain group, 45% in pain free group, TT-genotype=0 in pain group, 41% in pain free group, p=0.05) but not in the other SNPs. PGEM, but not IL-6, was significantly elevated in CSF of the pain group (3.6 pg/mL, sd=1.9 vs 2.1 pg/mL, p=0.03), IL-1ß was undetectable. Conclusion: We found elevated PGEM levels in CSF of patients with post cholecystectomy syndrome and visceral hyperalgesia, suggesting a central, possibly inflammatory component to the pain, and overrepresentation of the CT-genotype in the rs5275 SNP in the Cox2 gene, suggesting overexpression of Cox2 as a possible cause for elevated PGEM levels.

Microstructural and functional brain abnormalities in multiple sclerosis predicted by osteopontin and neurofilament light.

BACKGROUND: Osteopontin (OPN) is a proinflammatory biomarker, and neurofilament light chain (NFL) levels reflect axonal damage. Resting-state functional MRI (rs-fMRI) defines brain networks during wakeful rest. OBJECTIVE: To examine, if levels of OPN and NFL are associated on the long term with (i) lesion evolution, (ii) changes in normal-appearing white matter (NAWM) microstructure and (iii) functional connectivity in multiple sclerosis (MS). METHODS: Concentration of NFL and OPN in the blood and CSF were related to MRI findings 10.3 ± 2.8 years later in 53 patients with MS. NFL was examined by Simoa method, OPN by ELISA. Lesion volume in the brain and cervical spinal cord was examined by 3D FLAIR images. Voxel-wise images of fractional anisotropy (FA), axial diffusivity (AD), mean diffusivity (MD), and radial diffusivity (RD) were examined by tract-based spatial statistics corrected for gender, age and lesion volume. Metabolites were examined by single-voxel MR-spectroscopy in the NAWM. Fifty-five default mode network connections were examined by rs-fMRI corrected for gender, age, MS subtype and current therapy as covariates. RESULTS: While NFL in paired serum and CSF positively correlated (p = 0.019), there was no correlation between serum and CSF OPN. Higher OPN levels in the CSF but not in the serum showed association with increased brain WM lesion volume (p = 0.009) in 10.3 ± 2.8 years. Higher OPN in the CSF was associated with reduced FA, increased MD, and reduced RD in different NAWM areas 10.3 ± 2.8 years later. Higher OPN in the serum and CSF were associated with increased connectivity strength between the medial prefrontal cortex (MPFC) and other regions except with inferior parietal lobule. NFL in the CSF and in the serum was associated with decreased connectivity strength except for ventral MPFC-hippocampal formation. Neither serum OPN nor NFL at the time of the MRI were associated with functional connectivity changes. CONCLUSION: While serum NFL levels reflects CNS production, OPN in serum and CSF may have different cellular sources. OPN within the CSF but not in the serum may forecast development of lesions and microstructural abnormalities in 10 years, indicating the detrimental role of CNS inflammation on the long-term. Although both OPN and NFL in the CSF were associated with functional connectivity changes in 10 years, NFL was associated with decreased strength possibly indicating general axonal loss. In contrast, the positive association of OPN levels in the CSF with increased connectivity strength in 10 years may point to adaptive re-organization due to inflammatory WM lesions and microstructural changes.

Elevated Neurofilament Light Chain in Cerebrospinal Fluid and Plasma Reflect Inflammatory MRI Activity in Neurosarcoidosis.

BACKGROUND: Damage to axonal cells releases neurofilament light chain (NFL) into the cerebrospinal fluid and plasma. The objective of this study was to investigate NFL as a potential biomarker of disease activity in neurosarcoidosis. MRIs were graded according to enhancing lesions at different central nervous system (CNS) sites. RESULTS: In cerebrospinal fluid, levels of NFL were higher in neurosarcoidosis patients (n = 20) median 2304 pg/mL (interquartile range (IQR) 630-19,612) compared to 426 pg/mL (IQR 261-571) in extra-neurologic sarcoidosis patients (n = 20) and 336 pg/mL (IQR 194-402) in healthy controls (n = 11) (p = 0.0002). In plasma, levels of NFL were higher in neurosarcoidosis patients median 28.2 pg/mL (IQR 11.5-49.3) compared to 6.2 pg/mL (IQR 4.3-8.2) in extra-neurologic sarcoidosis patients and 7.1 pg/mL (IQR 6.2-9.0) in healthy controls (p = 0.0001). Levels in both cerebrospinal fluid and plasma were higher in neurosarcoidosis patients with moderate/severe enhancement than patients with mild enhancement on MRI (p = 0.009 and p = 0.005, respectively). To distinguish neurosarcoidosis patients from extra-neurologic patients and healthy controls, a cut-off level of 630 pg/mL in cerebrospinal fluid had 94% specificity and 79% sensitivity, while a cut-off level of 11.4 pg/mL in plasma had 97% specificity and 75% sensitivity. CONCLUSIONS: NFL levels in cerebrospinal fluid and plasma are significantly higher in neurosarcoidosis patients compared to extra-neurologic patients and healthy controls, and the levels correlate to the extent of inflammation on MRI.

Rapid SARS-CoV-2 Detection Using Electrochemical Immunosensor.

The outbreak of the coronavirus disease (COVID-19) pandemic caused by the novel coronavirus (SARS-CoV-2) has been declared an international public health crisis. It is essential to develop diagnostic tests that can quickly identify infected individuals to limit the spread of the virus and assign treatment options. Herein, we report a proof-of-concept label-free electrochemical immunoassay for the rapid detection of SARS-CoV-2 virus via the spike surface protein. The assay consists of a graphene working electrode functionalized with anti-spike antibodies. The concept of the immunosensor is to detect the signal perturbation obtained from ferri/ferrocyanide measurements after binding of the antigen during 45 min of incubation with a sample. The absolute change in the [Fe(CN)6]3-/4- current upon increasing antigen concentrations on the immunosensor surface was used to determine the detection range of the spike protein. The sensor was able to detect a specific signal above 260 nM (20 µg/mL) of subunit 1 of recombinant spike protein. Additionally, it was able to detect SARS-CoV-2 at a concentration of 5.5 × 105 PFU/mL, which is within the physiologically relevant concentration range. The novel immunosensor has a significantly faster analysis time than the standard qPCR and is operated by a portable device which can enable on-site diagnosis of infection.

CSF-Neurofilament Light Chain Levels in NMDAR and LGI1 Encephalitis: A National Cohort Study.

Background and Objectives: The two most common autoimmune encephalitides (AE), N-methyl-D-Aspartate receptor (NMDAR) and Leucine-rich Glioma-Inactivated 1 (LGI1) encephalitis, have been known for more than a decade. Nevertheless, no well-established biomarkers to guide treatment or estimate prognosis exist. Neurofilament light chain (NfL) has become an unspecific screening marker of axonal damage in CNS diseases, and has proven useful as a diagnostic and disease activity marker in neuroinflammatory diseases. Only limited reports on NfL in AE exist. We investigated NfL levels at diagnosis and follow-up in NMDAR and LGI1-AE patients, and evaluated the utility of CSF-NfL as a biomarker in AE. Methods: Patients were included from the National Danish AE cohort (2009-present) and diagnosed based upon autoantibody positivity and diagnostic consensus criteria. CSF-NfL was analyzed by single molecule array technology. Clinical and diagnostic information was retrospectively evaluated and related to NfL levels at baseline and follow-up. NMDAR-AE patients were subdivided into: idiopathic/teratoma associated or secondary NMDAR-AE (post-viral or concomitant with malignancies/demyelinating disease). Results: A total of 74 CSF samples from 53 AE patients (37 NMDAR and 16 LGI1 positive) were included in the study. Longitudinal CSF-NfL levels was measured in 21 patients. Median follow-up time was 23.8 and 43.9 months for NMDAR and LGI1-AE respectively. Major findings of this study are: i) CSF-NfL levels were higher in LGI1-AE than in idiopathic/teratoma associated NMDAR-AE at diagnosis; ii) CSF-NfL levels in NMDAR-AE patients distinguished idiopathic/teratoma cases from cases with other underlying etiologies (post-viral or malignancies/demyelinating diseases) and iii) Elevated CSF-NfL at diagnosis seems to be associated with worse long-term disease outcomes in both NMDAR and LGI1-AE. Discussion: CSF-NfL measurement may be beneficial as a prognostic biomarker in NMDAR and LGI1-AE, and high CSF-NfL could foster search for underlying etiologies in NMDAR-AE. Further studies on larger cohorts, using standardized methods, are warranted.

Prognostic impact of serum levels of EGFR and EGFR ligands in early-stage breast cancer.

Epidermal growth factor receptor (EGFR) and its ligands are involved in cancer pathogenesis. The emerging role of treatments co-targeting the EGFR system in breast cancer has increased the need to identify companion biomarkers. The aim of this study is to investigate whether pretreatment serum levels of EGFR and EGFR ligands in early-stage breast cancer patients might provide prognostic information as a stepping stone for further investigation. The study, which included 311 early-stage breast cancer patients, investigated associations between preoperative serum levels of EGFR and EGFR ligands (epidermal growth factor, heparin-binding epidermal growth factor (HBEGF), amphiregulin, transforming growth factor-α and betacellulin) and survival. Cutoffs were determined using Youden's method, and overall survival (OS) and invasive disease-free survival (IDFS) were evaluated using Cox regression. Preoperative S-EGFR < 60.3 ng/mL was associated with shorter OS and IDFS in both univariate analyses and when adjusting for standard prognostic factors (p < 0.05). Preoperative S-HBEGF < 21.4 pg/mL was associated with shorter OS in both univariate and multivariate analyses, whereas association with shorter IDFS could only be demonstrated in the univariate analysis. In conclusion, our study demonstrated shorter survival in early-stage breast cancer patients who had low pretreatment levels of either S-EGFR or S-HBEGF.

Neurofilament light chain as biomarker in idiopathic intracranial hypertension.

BACKGROUND: Damage of the optic nerve is the major complication of idiopathic intracranial hypertension. A biomarker indicative for optic nerve damage would help identifying high-risk patients requiring surgical procedures. Here, we studied the potential of cerebrospinal fluid neurofilament to predict idiopathic intracranial hypertension-induced optic nerve damage. METHODS: In two centers, serum and cerebrospinal fluid of 61 patients with clinically suspected idiopathic intracranial hypertension were prospectively collected. Neurofilament concentrations were measured and related to ophthalmological assessment. RESULTS: The average cerebrospinal fluid neurofilament concentration in patients with moderate and severe papilledema was increased compared to patients with minor and no papilledema (1755 ± 3507 pg/ml vs. 244 ± 102 pg/ml; p < 0.001). Cerebrospinal fluid neurofilament concentrations correlated with the maximal lumbar puncture opening pressure (r = 0.67, p < 0.001). In patients fulfilling the Friedman criteria for idiopathic intracranial hypertension with or without papilledema (n = 35), development of bilateral visual field defects and bilateral atrophy of the optic nerve were associated with increased average age-adjusted cerebrospinal fluid neurofilament concentrations. At last follow-up (n = 30), 8/13 of patients with increased, but only 3/17 with normal, cerebrospinal fluid neurofilament had developed bilateral visual field defects and/or bilateral optic nerve atrophy resulting in a sensitivity of 72.7% and a specificity of 73.7% of cerebrospinal fluid neurofilament to detect permanent optic nerve damage. CONCLUSIONS: Cerebrospinal fluid neurofilament is a putative biomarker for optical nerve damage in idiopathic intracranial hypertension.

Dysregulated EGFR pathway in serum in early-stage breast cancer patients: A case control study.

The epidermal growth factor receptor (EGFR) and its ligands are involved in cancer pathogenesis and they might serve as circulating biomarkers. The current study aims to investigate if abnormal pre-treatment serum levels of EGFR and EGFR ligands are present in women with early-stage breast cancer and if up- or downregulation of EGFR and EGFR ligands occur in defined patient subgroups. Pre-treatment serum samples were obtained from 311 women with newly diagnosed early-stage breast cancer and from 419 healthy women and analysed for EGFR and the ligands: Epidermal growth factor (EGF), heparin-binding epidermal growth factor (HBEGF), betacellulin (BTC), amphiregulin (AREG), and transforming growth factor α (TGF-α). Previously, age-dependent 95% reference intervals for EGFR and the EGFR ligands have been established based on the healthy women population. S-EGFR, S-EGF, S-HBEGF, S-AREG, and S-TGFα were all significantly different in women with breast cancer compared to healthy women (p < 0.05). Elevated S-EGFR, according to the reference intervals, was present in 11.3% of breast cancer patients, whereas decreased S-EGF was found in 11.6%. Elevated S-EGFR was associated with estrogen receptor positivity of tumor (ER+) and a subgroup of ER + breast cancer patients showed markedly elevated S-EGFR (>120 ng/mL).

Comparison of neurofilament light chain results between two independent facilities.

OBJECTIVES: To examine levels of neurofilament light chain (NFL) in identical cerebrospinal fluid (CSF) and blood samples at two different facilities, and how differences affect interpretation of levels within and above the normal range. METHODS: CSF and plasma from patients with multiple sclerosis (MS) and healthy controls (HCs) were analysed by Simoa (Quanterix) for levels of NFL providing a total of 165 CSF samples (119 from MS) and 225 plasma samples (180 from MS). RESULTS: CSF and plasma concentrations highly correlated between NFL laboratory facilities (R: 0.92 and 0.84, <0.0001, respectively), and there were no differences between facilities. A bias between the two sites for plasma was -0.95 pg/mL and for CSF -73.53 pg/mL. The cut-offs for CSF were 807.5 and 571.0 pg/mL at site 1 and site 2, respectively; the cut-offs for plasma were 13.0 and 11.8 pg/mL, respectively. Seven out of 180 plasma samples (3.9%) and 3 out of 119 CSF samples (2.5%) from MS patients could be reclassified as normal/abnormal, that is, below/above cut-off, when measured at different facilities. CONCLUSION: Our study demonstrates that results of NFL in CSF and blood measured with SIMOA are comparable between facilities. Nevertheless, healthcare practitioners should consider reference values at different laboratories, since different sensitivity/specificity can affect interpretation when low values are adjacent to cut-offs.

Effects of ß-hydroxybutyrate on cognition in patients with type 2 diabetes.

OBJECTIVE: Cognitive impairment in type 2 diabetes is associated with cerebral glucose hypometabolism. Providing a glucose substitute such as ketone bodies might restore metabolic balance in glucose-compromised neurones and improve cognitive performance. We aimed to investigate if ß-hydroxybutyrate (ketone body) infusion acutely affects cognitive performance, measured by a neuropsychological test battery, in patients with type 2 diabetes. DESIGN: Randomised, placebo-controlled, double-blind cross-over trial. METHODS: Eighteen patients with type 2 diabetes received i.v. ketone body (ß-hydroxybutyrate) and placebo (saline) infusion in a randomised order on two separate occasions. On both days of examination, blood glucose was clamped at 7.5 mmol/L and a neuropsychological test battery was used to assess global cognitive performance (primary outcome) and specialized cognitive measures of verbal memory, working memory, executive function, psychomotor speed, and sustained attention. RESULTS: During neurocognitive testing, ß-hydroxybutyrate concentrations were 2.4 vs 0.1 mmol/L. Working memory assessed by Wechsler Adult Intelligence Scale letter-number-sequencing significantly improved by 1.6 points (95% CI: 0.7, 2.4; non-adjusted P < 0.001) corresponding to a 17% increase in performance during ketone infusion compared to placebo. There was no change for global cognitive performance or any other cognitive measure after adjusting for multiple comparisons. Blood concentrations of ß-hydroxybutyrate and glycaemic status did not associate with test performance; however, insulin resistance measured by HOMA was related to improved working memory performance during ketone infusion (ß = 4%; 95% CI: 1.1, 7.7; P = 0.012). CONCLUSIONS: Ketone infusion specifically improved working memory performance in patients with type 2 diabetes in the absence of changes in global cognition.

EGFR and EGFR ligands in serum in healthy women; reference intervals and age dependency.

Background The epidermal growth factor receptor (EGFR) system is involved in cancer pathogenesis and serves as an important target for multiple cancer treatments. EGFR and its ligands epidermal growth factor (EGF), heparin-binding epidermal growth factor (HB-EGF), betacellulin (BTC), amphiregulin (AREG) and transforming growth factor α (TGF-α) have potential applications as prognostic or predictive serological biomarkers in cancer. The aim was to establish EGFR and EGFR ligand reference intervals in healthy women. Methods EGFR and EGFR ligands were measured in serum from 419 healthy women aged 26-78 years. The need for age partitioned reference intervals was evaluated using Lahti's method. EGFR and EGF were analyzed using ELISA assays, whereas HB-EGF, BTC, AREG and TGF-α were analyzed using the highly sensitive automated single molecule array (Simoa) enabling detection below the lower reference limit for all six biomarkers. Results Reference intervals for EGFR and the EGFR ligands were determined as the 2.5th and 97.5th percentiles. All six biomarkers were detectable in all serum samples. For EGFR, EGF, HB-EGF and TGF-α, reference intervals were established for women <55 years and for women >55 years, whilst common reference intervals were established for AREG and BTC including women aged 26-78 years. Conclusions Age specific reference intervals were determined for EGFR, EGF, HB-EGF, BTC, AREG and TGF-α.

Locally advanced rectal cancer transcriptomic-based secretome analysis reveals novel biomarkers useful to identify patients according to neoadjuvant chemoradiotherapy response.

Most patients with locally advanced rectal cancer (LARC) present incomplete pathological response (pIR) to neoadjuvant chemoradiotherapy (nCRT). Despite the efforts to predict treatment response using tumor-molecular features, as differentially expressed genes, no molecule has proved to be a strong biomarker. The tumor secretome analysis is a promising strategy for biomarkers identification, which can be assessed using transcriptomic data. We performed transcriptomic-based secretome analysis to select potentially secreted proteins using an in silico approach. The tumor expression profile of 28 LARC biopsies collected before nCRT was compared with normal rectal tissues (NT). The expression profile showed no significant differences between complete (pCR) and incomplete responders to nCRT. Genes with increased expression (pCR = 106 and pIR = 357) were used for secretome analysis based on public databases (Vesiclepedia, Human Cancer Secretome, and Plasma Proteome). Seventeen potentially secreted candidates (pCR = 1, pIR = 13 and 3 in both groups) were further investigated in two independent datasets (TCGA and GSE68204) confirming their over-expression in LARC and association with nCRT response (GSE68204). The expression of circulating amphiregulin and cMET proteins was confirmed in serum from 14 LARC patients. Future studies in liquid biopsies could confirm the utility of these proteins for personalized treatment in LARC patients.