Cohort Of Healthy Controls Research Articles

DNA Methylation Profiles in Whole Blood of Huntington's Disease Patients.

Epigenetic mechanisms, especially DNA methylation, are suggested to play a role in the age-of-onset in Huntington's disease (HD) based on studies on patient brains, and cellular and animal models. Methylation is tissue-specific and it is not clear how HD specific methylation in the brain correlates with the blood compartment, which represents a much more clinically accessible sample. Therefore, we explored the presence of HD specific DNA methylation patterns in whole blood on a cohort of HDM and healthy controls from Slovenia. We compared CpG site-specific DNA methylation in whole blood of 11 symptomatic and 9 pre-symptomatic HDM (HDM), and 15 healthy controls, by using bisulfite converted DNA on the Infinium® Human Methylation27 BeadChip microarray (Illumina) covering 27,578 CpG sites and 14,495 genes. Of the examined 14,495 genes, 437 were differentially methylated (p < 0.01) in pre-symptomatic HDM compared to controls, with three genes (CLDN16, DDC, NXT2) retaining statistical significance after the correction for multiple testing (false discovery rate, FDR < 0.05). Comparisons between symptomatic HDM and controls, and the comparison of symptomatic and pre-symptomatic HDM further identified 260 and 198 differentially methylated genes (p < 0.01), respectively, whereas the comparison of all HDM (symptomatic and pre-symptomatic) and healthy controls identified 326 differentially methylated genes (p < 0.01), however, none of these changes retained significance (FDR < 0.05) after the correction for multiple testing. The results of our study suggest that methylation signatures in the blood compartment are not robust enough to prove as valuable biomarkers for predicting HD progression, but recognizable changes in methylation deserve further research.

CDH23 Methylation Status and Presbycusis Risk in Elderly Women.

Introduction: Presbycusis, an age-related hearing impairment (ARHI) disease, is the most common cause for HI in adults worldwide. One of the best candidate genes for ARHI susceptibility is Cadherin 23 (CDH23) which encodes stereocilia tip-links of the inner ear sensory hair cell. Although alterations in the methylation status of CpG dinucleotides across various genes were reported to be associated with HI, methylation changes in CDH23 gene have not been reported previously.Objectives: This study aimed at investigating whether DNA methylation level of CDH23 gene at intragenic CpG island overlapping an exonic-intronic region at position chr10:73565570-73565827 (GRCh37/hg19) could be risk factor associated with ARHI.Materials and Methods: We screened for methylation changes in this particular position for CDH23 gene in 50 blood samples of elderly women affected with presbycusis and healthy control cohort. Methylation of CpG sites were assessed using Quantitative methylation-specific PCR (qMSP) following sodium bisulfite DNA conversion chemistry. Methylation levels were normalized against TSH2B reference gene.Results: DNA methylation analysis for the common CpG islands in CDH23 gene revealed 3.27-folds significant increase (p < 0.0001) in methylation profile for ARHI women as compared to healthy controls with an elevated risk odds ratio (OR) of 2.219 [95% CI 1.071–4.597].Conclusion: Our study is the first of its kind to prove that higher CpG site methylation levels in CDH23 gene are likely to be associated with ARHI.

BRCA germline mutations in an unselected nationwide cohort of Chinese patients with ovarian cancer and healthy controls

ObjectiveTo investigate the BRCA status in Chinese patients with ovarian cancer (OC). Though there were two large prevalence studies in Chinese OC patients, this was the first time to observe it in healthy controls. MethodsWe performed BRCA mutation screening using next-generation sequencing to determine the prevalence of BRCA germline deleterious mutations in an unselected cohort of Chinese OC patients (n = 1331) versus healthy controls (n = 1763) and describe the types and spectrum of BRCA deleterious variants. ResultsAmong the 1331 patients with OC, 227 (17.1%) carried deleterious variants in BRCA1 and 70 (5.3%) carried deleterious variants in BRCA2. Of 1763 control subjects, 6 (0.3%) and 2 (0.1%) had deleterious variants in BRCA1 and BRCA2. No patient carried mutations in both BRCA1 and BRCA2 simultaneously. Sixty-three novel mutations were identified, and three Chinese specific hot-spot mutations were notified as BRCA1 c.5470_5477delATTGGGCA, BRCA1 c.981_982delAT, and BRCA1 c.3770_3771delAG. Interestingly, all these high-frequency recurrent mutations were distributed on exon 10, which may also be the Chinese OC BRCA mutations' distinct characteristics. In addition, in our study, the estimated odds ratio (OR) of OC associated with BRCA1 positive variants were approximately 34.6 (95% CI, 12.5–95.7) in age group under 40 and 42.4 (95% CI, 5.9–305.2) in group older than 50 in the Chinese population, respectively. ConclusionsWe recommend BRCA testing to all Chinese OC patients and those general Chinese who have family members with hereditary breast and ovarian related cancer (HBOC)-related cancers. Variants carriers would not only benefit from early prevention of OC but also for the medical management.

Cholesterol efflux capacity in young acute coronary syndrome in Malaysia

Aim: In recent years, patients with acute coronary syndrome (ACS) are significantly younger. We hypothesized that the capacity of HDL to accept cholesterol from macrophages, via cholesterol efflux capacity (CEC) would serve as a predictor of atherosclerotic disease. Hence, we aim to compare CEC of young ACS to a healthy control cohort.

Elevated salivary alpha-amylase levels at awakening in patients with depression

BackgroundSpecific Major Depressive Disorder (MDD) biomarkers could help improve our understanding of MDD pathophysiology and aid in the refinement of current MDD criteria. While salivary cortisol (SC) can differentiate between healthy controls and patients with psychiatric disorders, salivary alpha amylase (sAA), may be a putative candidate biomarker for MDD specifically. MethodsIn a naturalistic cohort of consecutive out-patients and healthy controls, sAA and SC were determined in 833 participants (97 MDD patients, 142 patients with other mood, anxiety, and/or somatoform (MAS-) disorders, and 594 healthy controls). Samples were collected at 7 different time points (at awakening, after 30, 45, and 60 min, at 10:00 p.m., at 11:00 p.m., and at awakening on day 2). ResultsThe mean age of the sample was 43.8 years (SD = 12.9; 63.9% female). Concerning sAA, MDD patients had higher sAA levels upon awakening on two consecutive days (p = 0.04, p = 0.01 respectively), as well as a higher area under the curve with respect to the increase (AUCi; p = 0.04) in comparison to both controls and the other MAS-disorders group. Regarding SC, mean levels of evening SC were elevated in MDD patients (p = 0.049) in comparison to both controls and the other MAS-disorders group. SC values on day 2 after ingestion of dexamethasone were elevated in both MDD patients and the other MAS-disorders group (p = 0.04, p = 0.047 respectively). ConclusionssAA at awakening and not cortisol differentiates MDD from other psychiatric disorders in outpatients. This suggests that sAA may be a valuable candidate biomarker specifically for MDD.

Functional genetic variants in the SIRT5 gene promoter in acute myocardial infarction

Coronary artery disease (CAD) including acute myocardial infarction (AMI) is a common complex disease. To date, genetic causes for atherosclerosis remain largely unknown. It has recently been proposed that low frequency and rare genetic variants may be the main causes. Mitochondrial sirtuins, SIRT3, SIRT4 and SIRT5, function as critical regulators of mitochondrial metabolism, oxidative stress and cell survival. We speculated that altered SIRT5 level resulting from DNA sequence variants (DSVs) within SIRT5 gene regulatory regions may contribute to the CAD and AMI development. In this study, the SIRT5 gene promoter was genetically and functionally analyzed in large cohorts of AMI patients (n = 381) and healthy controls (n = 391). A total of eleven DSVs and SNPs were found. Two novel heterozygous DSVs (g.13574131C>A and g.13574287G>C) and three heterozygous SNPs [g.13573450A>G (rs573515169), g.13574110G>A (rs2804924) and g.13574259G>C (rs112443954)] were identified only in AMI patients. The DSVs and SNPs significantly decreased the transcriptional activity of the SIRT5 gene promoter in both HEK-293 and H9c2 cells (P < 0.05). Further electrophoretic mobility shift assay indicated that the SNPs significantly affected the binding of transcription factors. In contrast, the DSVs and SNPs found only in controls or in both AMI patients and controls did not significantly change the SIRT5 gene promoter activity (P > 0.05). Therefore, our data suggested that the DSVs and SNPs identified in AMI patients may change SIRT5 level by affecting activity of SIRT5 gene promoter, contributing to the AMI development as a risk factor.

Functional outcome in contemporary children and young adults with tetralogy of Fallot after repair

ObjectiveFunctional outcome measures are of growing importance in the aftercare of patients with congenital heart disease. This study addresses the functional status with regard to exercise capacity, health-related physical fitness...

Long-term outcomes of renal transplant in recipients with lower urinary tract dysfunction

OBJECTIVES Lower urinary tract dysfunction can lead to chronic kidney disease, which, despite surgical intervention, will progress to end-stage renal disease, requiring dialysis. Urologic pathology may damage a transplanted kidney, limiting patient and graft survival. Although smaller studies have suggested that urinary tract dysfunction does not affect graft or patient survival, this is not universally accepted. Northern Ireland has historically had the highest incidence of neural tube defects in Europe, giving rich local experience in caring for patients with lower urinary tract dysfunction. Here, we analyzed outcomes of renal transplant recipients with lower urinary tract dysfunction versus control recipients. MATERIALS AND METHODS We identified 3 groups of kidney transplant recipients treated between 2001 and 2010; those in group 1 had end-stage renal disease due to lower urinary tract dysfunction with prior intervention (urologic surgery, long-term catheter, or intermittent self-catheterization), group 2 had end-stage renal disease secondary to lower urinary tract dysfunction without intervention, and group 3 had end-stage renal disease due to polycystic kidney disease (chosen as a relatively healthy control cohort without comorbid burden of other causes of end-stage renal disease such as diabetes). The primary outcome measured, graft survival, was death censored, with graft loss defined as requirement for renal replacement therapy or retransplant. Secondary outcomes included patient survival and graft function. RESULTS In 150 study patients (16 patients in group 1, 64 in group 2, and 70 in group 3), 5-year death-censored graft survival was 93.75%, 90.6%, and 92.9%, respectively, with no significant differences in graft failure among groups (Cox proportional hazards model). Five-year patient survival was 100%, 100%, and 94.3%, respectively. CONCLUSIONS Individuals with a history of lower urinary tract dysfunction had graft and patient survival rates similar to the control group. When appropriately treated, lower urinary tract dysfunction is not a barrier to successful renal transplant.

Abstract 493: Circulating biliary tract microRNA signature discriminates cholangiocarcinoma from pancreatic cancer

Abstract Accurate diagnosis of lesions in the pancreatic head is a challenge with substantial clinical consequences in daily practice. Located in the pancreatic head, both distal cholangiocarcinoma (CCA) and pancreatic ductal adenocarcinoma (PDAC) are highly lethal malignancies with limited but distinct therapeutic options. Given these different therapeutic considerations, discovery of novel biomarkers that can ensure differential diagnostic specificity is an unmet clinical need. This study aims to identify a circulating microRNA (miRNA) signature to accurately diagnose and discriminate distal CCA from PDAC. In the initial discovery phase, microarray profiling of a panel of 752 miRNAs was performed on seven distal CCA and seven age- and sex-matched healthy control plasma samples. Raw data were normalized to the global mean. Based on probability value and fold change, candidate miRNAs were selected for further validation. In the validation phase, selected candidate miRNAs were analyzed in an independent cohort of distal CCA (n = 22) and healthy controls (n = 32) using RT-qPCR. To further assess the origin of lesions, candidate miRNA expression profiles were analyzed in malignant disease (n = 90) compared to benign disease (n = 15), and distal CCA (n = 22) compared to PDAC (n = 30). Following an empirically based selection of potential reference genes, raw data were normalized to a combination of two normalizing reference miRNAs. In the discovery phase, microarray analysis revealed nineteen miRNAs that were significantly deregulated in patients with distal CCA compared to healthy controls. Thirteen miRNAs were selected for further validation. In the validation phase, using logistic regression analysis, a three-miRNA panel was identified as the most robust diagnostic signature to discriminate malignant from benign disease (AUC = 0.881). Remarkably, two miRNAs of this panel were validated as specific classifier for distal CCA, with an AUC of 0.814 in differentiating distal CCA from PDAC. In addition, a combination of miR-93 and miR-101 was used as bona fide normalizing reference gene for these analyses. In conclusion, a combined panel of plasma miRNAs shows promising diagnostic capability to serve as unique minimally invasive distal CCA biomarker and has the potential to help clinical decision making for patients affected by pancreatobiliary disease in the pancreatic head. Citation Format: Jisce R. Puik, Laura L. Meijer, Tessa Y. Le Large, Michal Heger, Frederike Dijk, Niccola Funel, Ingrid Garajová, Elisa Giovannetti, Geert Kazemier. Circulating biliary tract microRNA signature discriminates cholangiocarcinoma from pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 493.

Metabolomic Profiles for Primary Progressive Multiple Sclerosis Stratification and Disease Course Monitoring.

Primary progressive multiple sclerosis (PPMS) shows a highly variable disease progression with poor prognosis and a characteristic accumulation of disabilities in patients. These hallmarks of PPMS make it difficult to diagnose and currently impossible to efficiently treat. This study aimed to identify plasma metabolite profiles that allow diagnosis of PPMS and its differentiation from the relapsing-remitting subtype (RRMS), primary neurodegenerative disease (Parkinson’s disease, PD), and healthy controls (HCs) and that significantly change during the disease course and could serve as surrogate markers of multiple sclerosis (MS)-associated neurodegeneration over time. We applied untargeted high-resolution metabolomics to plasma samples to identify PPMS-specific signatures, validated our findings in independent sex- and age-matched PPMS and HC cohorts and built discriminatory models by partial least square discriminant analysis (PLS-DA). This signature was compared to sex- and age-matched RRMS patients, to patients with PD and HC. Finally, we investigated these metabolites in a longitudinal cohort of PPMS patients over a 24-month period. PLS-DA yielded predictive models for classification along with a set of 20 PPMS-specific informative metabolite markers. These metabolites suggest disease-specific alterations in glycerophospholipid and linoleic acid pathways. Notably, the glycerophospholipid LysoPC(20:0) significantly decreased during the observation period. These findings show potential for diagnosis and disease course monitoring, and might serve as biomarkers to assess treatment efficacy in future clinical trials for neuroprotective MS therapies.

Incidence of invasive pneumococcal disease in immunocompromised patients: A systematic review and meta-analysis.

Invasive pneumococcal disease (IPD) is associated with high morbidity and mortality, with immunocompromised patients (ICPs) at particular risk. Therefore, guidelines recommend pneumococcal vaccination for these patients. However, guidelines are scarcely underpinned with references to incidence studies of IPD in this population. This, potentially results in unawareness of the importance of vaccination and low vaccination rates. The objective of this systematic review and meta-analysis was to assess the incidence of IPD in ICPs. We systematically searched PubMed and Embase to identify studies in English published before December 6th, 2017 that included terms related to 'incidence', 'rate', 'pneumococcal', 'pneumoniae', 'meningitis', 'septicemia', or 'bacteremia'. We focused on patients with HIV, transplantation and chronic inflammatory diseases. We included 45 studies in the systematic review reporting an incidence or rate of IPD, defined as isolation of Streptococcus pneumoniae from a normally sterile site. Random effects meta-analysis of 38 studies showed a pooled IPD incidence of 331/100,000 person years in patients with HIV in the late-antiretroviral treatment era in non-African countries, and 318/100,000 in African countries; 696 and 812/100,000 in patients who underwent an autologous or allogeneic stem cell transplantation, respectively; 465/100,000 in patients with a solid organ transplantation; and 65/100,000 in patients with chronic inflammatory diseases. In healthy control cohorts, the pooled incidence was 10/100,000. ICPs are at increased risk of contracting IPD, especially those with HIV, and those who underwent transplantation. Based on our findings, we recommend pneumococcal vaccination in immunocompromised patients. ID: CRD42016048438.

Whole blood miRNA expression analysis reveals miR-3613-3p as a potential biomarker for dedifferentiated liposarcoma.

Liposarcoma constitute about 13% of all soft tissue sarcoma and are associated with a high risk of metastases. As the preoperative differentiation between benign and malign lipomatous tumors is restricted to magnetic resonance imaging, computed tomography and biopsy, we performed a miRNA array to distinguish dedifferentiated liposarcoma patients from healthy controls and lipoma patients. Blood samples of patients with dedifferentiated liposarcoma, healthy controls and lipoma patients were collected. Whole blood RNA was extracted and samples of patients with dedifferentiated liposarcoma (n= 6) and of healthy donors (n= 4) were analyzed using an Affymetrix GeneChip miRNA Array v. 4.0. qRT-PCR was carried out to confirm the most differentially expressed miRNA; being further analyzed in an independent cohort of healthy controls as well as in lipoma patients. As shown by the microarray, two miRNAs (miR-3613-3p, miR-4668-5p) were shown to be significantly upregulated (fold change: > 2.5; p< 0.05) in patients with dedifferentiated liposarcoma (n= 6) as compared to healthy controls (n= 4). miR-3613-3p was further validated by qRT-PCR to be significantly upregulated in dedifferentiated liposarcoma patients compared to an independent cohort of healthy controls (n= 3) and lipoma patients (n= 5). We identified a specific whole blood miRNA (miR-3613-3p) that may serve to distinguish between dedifferentiated liposarcoma patients and healthy controls, thus potentially serving as a specific biomarker for dedifferentiated liposarcoma.

Associations of plasma interleukin-6 with plasma and cerebrospinal fluid monoamine biosynthetic pathway metabolites in treatment-resistant depression

BackgroundCurrent research suggests that depression is associated with several changes in the immune system, including alterations in inflammatory cytokines, the hypothalamic-pituitary-adrenal axis, and downstream effects on neurotransmitters. Specifically, plasma interleukin-6 (IL-6) is elevated in depressed individuals compared to non-depressed controls, and these same inflammatory processes may be implicated in treatment resistant depression (TR-MDD). IL-6 has also been theorized to affect central monoamine pathway metabolites. Few studies have assessed the relationship of IL-6 and plasma/CSF monoamine biosynthetic pathway metabolites and symptomatic correlates of depression. MethodsWe analyzed plasma IL-6 levels in an outpatient treatment-resistant depressed cohort (n = 67) and healthy control cohort (n = 40). We also assessed relationship of plasma IL-6 with plasma/CSF monoamine pathway metabolites and symptomatic correlates of depression. ResultsHigher levels of plasma IL-6 were found in the TR-MDD cohort compared to the control cohort. Plasma IL-6 correlated with plasma serotonin levels among all participants, although there was no association of plasma IL-6 and plasma serotonin in the TR-MDD cohort on subgroup analysis. No correlations were found between plasma IL-6 and plasma/CSF monoamine breakdown products, depressive symptomatology, or suicidality. LimitationsOur analysis was a post-hoc analysis and we were not able to exhaustively control for confounds of IL-6 levels, such as sleep or stress. ConclusionThe findings of the current study extend current evidence that plasma IL-6 levels are elevated in TR-MDD, after adjusting for body mass index. Additionally, plasma IL-6 and serotonin regulation may vary between healthy and TR-MDD populations. Lastly, our study finds no association between plasma IL-6 and CSF monoamine pathway metabolites, suggesting that further research is needed to elucidate the association of IL-6 and depression.

Functional variants in the LC3B gene promoter in acute myocardial infarction.

Acute myocardial infarction (AMI) is a common disease mainly caused by atherosclerosis, for which genetic causes remain largely unknown. Recently, low frequency and rare genetic variants have been proposed as risk factors. Autophagy has been involved in many cellular processes, such as lipid metabolism and inflammation, and implicated in human diseases, including cardiovascular diseases. In previous studies, we have reported reduced levels of LC3B, a core protein and a marker for autophagy, in AMI patients. In this study, the LC3B gene promoter was genetically and functionally analyzed in large cohorts of AMI patients (n = 383) and healthy controls (n = 390). A total of 25 DNA sequence variants (DSVs) including SNPs were found. Seven DSVs and three SNPs were only identified in AMI patients. All the DSVs and SNPs (except one) significantly decreased the transcriptional activity of the LC3B gene promoter in both HEK-293 and H9c2 cells (P < 0.05). Further electrophoretic mobility shift assay suggested that the DSVs affected the binding of transcription factors. In contrast, the DSVs and SNPs found only in controls or in both AMI patients and controls did not significantly affected LC3B gene promoter activity (P > 0.05). Therefore, our data suggested that the DSVs identified in AMI patients may change LC3B level by affecting the transcriptional activity of LC3B gene promoter, contributing to the AMI development. Upregulation of the LC3B gene expression may provide a novel and potential therapy for AMI patients.

Detection of alpha-synuclein conformational variants from gastro-intestinal biopsy tissue as a potential biomarker for Parkinson's disease.

AimsGastrointestinal (GI) α‐synuclein (aSyn) detection as a potential biomarker of Parkinson's disease (PD) is challenged by conflicting results of recent studies. To increase sensitivity and specificity, we applied three techniques to detect different conformations of aSyn in GI biopsies obtained from a longitudinal, clinically well‐characterized cohort of PD patients and healthy controls (HC).MethodsWith immunohistochemistry (IHC), we used antibodies reactive for total, phosphorylated and oligomeric aSyn; with aSyn proximity ligation assay (AS‐PLA), we targeted oligomeric aSyn species specifically; and with paraffin‐embedded tissue blot (AS‐PET‐blot) we aimed to detect fibrillary, synaptic aSyn.ResultsA total of 163 tissue blocks were collected from 51 PD patients (113 blocks) and 21 HC (50 blocks). In 31 PD patients, biopsies were taken before the PD diagnosis (Prodromal); while in 20 PD patients biopsies were obtained after diagnosis (Manifest). The majority of tissues blocks were from large intestine (62%), followed by small intestine (21%), stomach (10%) and oesophagus (7%). With IHC, four staining patterns were detected (neuritic, ganglionic, epithelial and cellular), while two distinct staining patterns were detected both with AS‐PLA (cellular and diffuse signal) and with AS‐PET‐blot (aSyn‐localized and pericrypt signal). The level of agreement between different techniques was low and no single technique or staining pattern reliably distinguished PD patients (Prodromal or Manifest) from HC.ConclusionsOur study suggests that detection of aSyn conformational variants currently considered pathological is not adequate for the diagnosis or prediction of PD. Future studies utilizing novel ultrasensitive amyloid aggregation assays may increase sensitivity and specificity.

T77. Automated Reconstruction of White Matter Pathways in Attention-Deficit / Hyperactivity Disorder Using Anatomical Priors

ADHD has repeatedly been associated with alterations in white matter microstructure, as measured through fractional anisotropy (FA) and acquired through diffusion-weighted MRI (DWI). Due to methodological heterogeneity of analysis techniques, there have been inconsistent results regarding anatomical specificity of changes in FA. The aim of this study was to determine, in an automated manner, whether FA is related to ADHD in a large cohort of patients and healthy controls.

Circulating MicroRNA Profile as a Potential Predictive Biomarker for Early Diagnosis of Spontaneous Abortion in Patients With Subclinical Hypothyroidism.

An increasing number of studies suggest that subclinical hypothyroidism (SCH) is associated with complications of gestation, including spontaneous abortion (SA). However, the underlying mechanism is not clear. MicroRNA (miRNA) has been demonstrated to be closely related to gynecological reproductive diseases. We determined miRNA expression in patients with SCH, SCH with SA (SCH + SA), and in those with SA as well as healthy controls (HCs), and analyzed whether dysregulation in several miRNAs was specific to these cohorts. An Agilent Human miRNA array was used to explore miRNA levels in pooled serum samples as a pilot study, followed by a validation of selected miRNAs by real-time polymerase chain reaction in SCH (N = 24), SA (N = 19), SCH + SA (N = 21), and HC cohorts (N = 18). The relative expression of miR-940 was elevated in the SCH + SA group compared with SCH, SA, and HC groups. In addition, miR-486-5p was upregulated in the SCH + SA group compared with SA and HC groups, without a difference noted between SCH + SA and SCH groups. Further analysis suggested that miR-940 or miR-486-5p may be potential predictive biomarkers for the early diagnosis of SA in patients with SCH.

Circulating programmed death ligand-1 (cPD-L1) in non-small-cell lung cancer (NSCLC).

BackgroundThis study aimed at investigating feasibility of programmed death ligand-1 (PD-L1) testing in plasma samples of advanced NSCLC patients receiving first-line treatment, assessing whether circulating (c)PD-L1 levels were modified by the therapy and whether baseline cPD-L1 levels were associated with patients’ clinical responses and survival outcome.MethodsPeripheral blood samples were collected from 16 healthy volunteers and 56 newly diagnosed NSCLC patients before and at 12th week during the course of first-line therapy. The level of PD-L1 was measured in plasma samples using the human (PD-L1/CD274) ELISA kit (CUSABIO, MD, USA). The Mann Whitney test or Fisher’s test were used for comparisons. Survival analysis was performed using Kaplan Meyer method, providing median and p-value.ResultsBaseline median cPD-L1 was 42.21 pg/ml (range 12.00-143.49) in NSCLC patients and 37.81 pg/ml (range 9.73-90.21) in healthy control cohort (p = 0.78). Median cPD-L1 increased in patients treated with first-line chemotherapy (63.20 pg/ml vs 39.34 pg/ml; p = 0.002), with no changes in patients exposed to non-chemotherapy drugs (42.39 pg/ml vs 50.67 pg/ml; p = 0.398). Time to progression and overall survival were 4.4 vs 6.9 months (p = 0.062) and 8.8 vs 9.3 months (p = 0.216) in cPD-L1 positive vs cPD-L1 negative patients. Baseline cPD-L1 levels increased with the ascending number of metastatic sites, even if the association was not statistically significant (p = 0.063).ConclusionsThis study showed that cPD-L1 testing is feasible, with chemotherapy influencing PD-L1 plasma levels. The possibility of using such test for predicting or monitoring the effect of immunotherapy or combination of chemotherapy and immunotherapy warrant further investigations.

T69. PERFORMANCE OF PATIENTS WITH NMDAR-AB ENCEPHALITIS COMPARED TO PATIENTS WITH SCHIZOPHRENIA AND HEALTHY CONTROLS USING FUNCTIONAL MAGNETIC RESONANCE IMAGING (F-MRI) AND A WORKING MEMORY PARADIGM

BackgroundN-methyl-D-aspartate receptor (NMDAR) encephalitis has prominent early psychotic features. It has been rarely misdiagnosed as schizophrenia. NMDAR encephalitis causes deficiencies in working memory in the short & long term. Early immunotherapy/tumour removal improves outcomes. Schizophrenia can cause significant cognitive performance deficits with working memory often severely affected.Clinical MRI in NMDAR encephalitis is usually normal, despite severe clinical course. Resting state fMRI studies have shown reduced functional connectivity of the left & right hippocampus. To date, to the best of our knowledge, functional neuroimaging utilising a working memory task such as the 1-back task (participant has to identify the number onscreen immediately before the number currently on screen) has not been utilised in NMDAR encephalitis. Potentially this may highlight alterations in brain regions through altered neuronal activity underlying working memory.We hypothesised that patients with NMDAR encephalitis would demonstrate a difference in BOLD response (a proxy for neuronal activity) compared to healthy controls during fMRI while performing the 1-back task. Additionally, we wished to compare task evoked BOLD response in patients with NMDAR-Ab encephalitis compared to patients with schizophrenia.MethodsUsing G*Power 3.1 software with effect size f (0.8), total sample size was calculated from data obtained from a previous study on working memory performance in NMDAR encephalitis for the 3 groups (NMDAR encephalitis, healthy controls & schizophrenia) (N=30). Following ethical approval, 12 individuals diagnosed with NMDAR encephalitis were recruited from neurological centres. Participants were trained in the 1-back task prior to scanning. Results were compared to previously collected data from cohorts of healthy controls (N=14) and schizophrenia (N=14). Cohorts were matched insofar as possible. Data was analysed using SPSS and ART toolbox.ResultsIn the NMDAR encephalitis (N=12) cohort, gender ratio F:M was (11:1), mean(s.d.) age 37.5(12.2) years. 50% identified ongoing memory problems. Schizophrenia cohort (N=14) gender ratio (7:7), mean (s.d.) age 39 (12.3) years. Healthy controls gender ratio (10.4) mean (s.d.) age 29.8 (6.5) years. There was no difference for gender, age or educational level between cohorts. FSIQ was higher (p<0.01) in controls & NMDAR encephalitis compared to schizophrenia cohorts. NMDAR encephalitis cohort had acute symptoms [mean (s.d.)] for 7.09 (2.43) weeks until treatment. 9 individuals were still on immunotherapy at time of entry.Duration since diagnosis to entry into this study was 55.18(23.13) months for NMDAR encephalitis and 12.6 (10.91) years for schizophrenia. There were no significant differences in number of ART motion outliers (p >0.05) between cohorts. Age & gender were added to second level analyses as covariates.One-way ANOVA was used to compare N-back accuracy & reaction time. There was a significant difference (p<0.01) between groups on accuracy but not reaction time, with only the schizophrenia group showing lower accuracy.1-back working memory condition was associated with significantly increased BOLD response across several brain regions that play a role in working memory, including the dorsolateral prefrontal cortex. However, there were no significant main effects on BOLD response for the 1-back task for any cohort.DiscussionPatients with NMDAR encephalitis report ongoing memory difficulties. However, in this study, there were no identifiable deficits compared to healthy controls using fMRI. This may relate to regeneration of NMDA receptors, the relatively long time until from diagnosis to follow up and early treatment with immunotherapy.

38.3 ONGOING GERMINAL CENTRE REACTIONS CONTRIBUTE TO N-METHYL-D-ASPARTATE RECEPTOR (NMDAR) ANTIBODY PRODUCTION IN NMDAR-ANTIBODY ENCEPHALITIS

BackgroundImmunoglobulin G (IgG) against the NR1-subunit of the N-methyl-D-aspartate (NMDAR) receptor mediates NMDAR-antibody encephalitis (NMDAR-Ab-E). This multi-stage illness presents with an acute severe psychiatric syndrome, alongside other neurological features, similar to human and animal NMDAR antagonist models. The disease is associated with an ovarian teratoma in around 20% of cases. The cellular immunity underlying this disease is not well understood. While antibody-modifying immunotherapies often promote disease resolution, the illness can be refractory to these treatments correlating with sub-optimal outcomes.NR1-IgG can be detected several years after clinical resolution, which may be via ongoing germinal centre reactions or the establishment of antibody-secreting cells as long-lived plasma cells in bone marrow niches. These two divergent models implicate use of differing immunotherapies to target these cells. Here we investigate the contribution of ongoing germinal centre reactions to disease progression, potentially informing disease mechanisms and guide targeted immunotherapy.MethodsWe hypothesised that recurrent antigen-driven germinal centre reactions would be associated with active generation of NR1-specific IgM and IgG and NR1-specific circulating B cells. We validated a NR1-IgM cell based assay establishing specificity cut-offs by screening healthy and disease control cohorts alongside a previously collected NMDAR-Ab-E cohort (n=46). Following this we went on to explore the temporal evolution of NR1-IgG and NR1-IgM titres in a prospective cohort (n=12).To investigate the lymphocyte characteristics, we stimulated ovarian teratoma lymphocytes and peripheral blood mononuclear cells (PBMCs) from multiple time points under varying cytokine conditions to understand whether these circulating cells showed capacity for NR1-IgG and IgM generation.ResultsWe found a 43% prevalence rate of NR1-IgM in the historic cohort. We then confirmed that NR1-IgM binding was specific by its selective depletion after anti-IgM precipitation but not with protein G. In the prospective cohort, we noted often high titres of IgM (up to 1:500) most commonly early in the disease but persisting for around 2 years. NR1-IgM levels varied in titre alongside NR1-IgG spikes. Consistently, culture experiments of patient lymphocytes (PBMCs and tumour-derived) produced varying degrees of NR1-IgM and NR1-IgG under conditions associated with B cell proliferation. The NR1-IgG levels correlated with serum NR1-titres suggesting these circulating B cells made a proportional contribution to serum levels.DiscussionOngoing germinal centre reactions likely contribute much of the circulating NR1-specific B cell population in NMDAR-Ab-E. Autoimmunisation at these centres represents an as yet unexplored therapeutic target in this and potentially other autoimmune encephalopathies. Regional specificity of these reactions including lymph nodes draining sources of NR1-antigen require further direct evaluation.