Progressive Multifocal Leukoencephalopathy Treated by Immune Checkpoint Inhibitors.

OBJECTIVE: Our aim was to assess the real-world effectiveness of immune checkpoint inhibitors for treatment of patients with progressive multifocal leukoencephalopathy (PML). METHODS: We conducted a multicenter survey compiling retrospective data from 79 PML patients, including 38 published cases and 41 unpublished cases, who received immune checkpoint inhibitors as add-on to standard of care. One-year follow-up data were analyzed to determine clinical outcomes and safety profile. Logistic regression was used to identify variables associated with 1-year survival. RESULTS: Predisposing conditions included hematological malignancy (n = 38, 48.1%), primary immunodeficiency (n = 14, 17.7%), human immunodeficiency virus/acquired immunodeficiency syndrome (n = 12, 15.2%), inflammatory disease (n = 8, 10.1%), neoplasm (n = 5, 6.3%), and transplantation (n = 2, 2.5%). Pembrolizumab was most commonly used (n = 53, 67.1%). One-year survival was 51.9% (41/79). PML-immune reconstitution inflammatory syndrome (IRIS) was reported in 15 of 79 patients (19%). Pretreatment expression of programmed cell death-1 on circulating T cells did not differ between survivors and nonsurvivors. Development of contrast enhancement on follow-up magnetic resonance imaging at least once during follow-up (OR = 3.16, 95% confidence interval = 1.20-8.72, p = 0.02) was associated with 1-year survival. Cerebrospinal fluid JC polyomavirus DNA load decreased significantly by 1-month follow-up in survivors compared to nonsurvivors (p < 0.0001). Thirty-two adverse events occurred among 24 of 79 patients (30.4%), and led to treatment discontinuation in 7 of 24 patients (29.1%). INTERPRETATION: In this noncontrolled retrospective study of patients with PML who were treated with immune checkpoint inhibitors, mortality remains high. Development of inflammatory features or overt PML-IRIS was commonly observed. This study highlights that use of immune checkpoint inhibitors should be strictly personalized toward characteristics of the individual PML patient. ANN NEUROL 2023;93:257-270.

Identification of DAGLA as an autoantibody target in cerebellar ataxia.

BACKGROUND: We aimed to investigate the clinical, imaging and fluid biomarker characteristics in patients with antidiacylglycerol lipase alpha (DAGLA)-autoantibody-associated cerebellitis. METHODS: Serum and cerebrospinal fliud (CSF) samples from four index patients were subjected to comprehensive autoantibody screening by indirect immunofluorescence assay (IIFA). Immunoprecipitation, mass spectrometry and recombinant protein assays were used to identify the autoantigen. Sera from 101 patients with various neurological symptoms and a similar tissue staining pattern as the index patient samples, and 102 healthy donors were analysed in recombinant cell-based IIFA (RC-IIFA) with the identified protein. Epitope characterisation of all positive samples was performed via ELISA, immunoblot, immunoprecipitation and RC-IIFA using different DAGLA fragments. RESULTS: All index patients were relatively young (age: 18-34) and suffered from pronounced gait ataxia, dysarthria and visual impairments. Paraclinical hallmarks in early-stage disease were inflammatory CSF changes and cerebellar cortex hyperintensity in MRI. Severe cerebellar atrophy developed in three of four patients within 6 months. All patient samples showed the same unclassified IgG reactivity with the cerebellar molecular layer. DAGLA was identified as the target antigen and confirmed by competitive inhibition experiments and DAGLA-specific RC-IIFA. In RC-IIFA, serum reactivity against DAGLA was also found in 17/101 disease controls, including patients with different clinical phenotypes than the one of the index patients, and in 1/102 healthy donors. Epitope characterisation revealed that 17/18 anti-DAGLA-positive control sera reacted with a C-terminal intracellular DAGLA 583-1042 fragment, while the CSF samples of the index patients targeted a conformational epitope between amino acid 1 and 157. CONCLUSIONS: We propose that anti-DAGLA autoantibodies detected in CSF, with a characteristic tissue IIFA pattern, represent novel biomarkers for rapidly progressive cerebellitis.

A genome-wide association study in autoimmune neurological syndromes with anti-GAD65 autoantibodies.

Autoimmune neurological syndromes (AINS) with autoantibodies against the 65 kDa isoform of the glutamic acid decarboxylase (GAD65) present with limbic encephalitis, including temporal lobe seizures or epilepsy, cerebellitis with ataxia, and stiff-person-syndrome or overlap forms. Anti-GAD65 autoantibodies are also detected in autoimmune diabetes mellitus, which has a strong genetic susceptibility conferred by human leukocyte antigen (HLA) and non-HLA genomic regions. We investigated the genetic predisposition in patients with anti-GAD65 AINS. We performed a genome-wide association study (GWAS) and an association analysis of the HLA region in a large German cohort of 1214 individuals. These included 167 patients with anti-GAD65 AINS, recruited by the German Network for Research on Autoimmune Encephalitis (GENERATE), and 1047 individuals without neurological or endocrine disease as population-based controls. Predictions of protein expression changes based on GWAS findings were further explored and validated in the CSF proteome of a virtually independent cohort of 10 patients with GAD65-AINS and 10 controls. Our GWAS identified 16 genome-wide significant (P < 5 × 10-8) loci for the susceptibility to anti-GAD65 AINS. The top variant, rs2535288 [P = 4.42 × 10-16, odds ratio (OR) = 0.26, 95% confidence interval (CI) = 0.187-0.358], localized to an intergenic segment in the middle of the HLA class I region. The great majority of variants in these loci (>90%) mapped to non-coding regions of the genome. Over 40% of the variants have known regulatory functions on the expression of 48 genes in disease relevant cells and tissues, mainly CD4+ T cells and the cerebral cortex. The annotation of epigenomic marks suggested specificity for neural and immune cells. A network analysis of the implicated protein-coding genes highlighted the role of protein kinase C beta (PRKCB) and identified an enrichment of numerous biological pathways participating in immunity and neural function. Analysis of the classical HLA alleles and haplotypes showed no genome-wide significant associations. The strongest associations were found for the DQA1*03:01-DQB1*03:02-DRB1*04:01HLA haplotype (P = 4.39 × 10-4, OR = 2.5, 95%CI = 1.499-4.157) and DRB1*04:01 allele (P = 8.3 × 10-5, OR = 2.4, 95%CI = 1.548-3.682) identified in our cohort. As predicted, the CSF proteome showed differential levels of five proteins (HLA-A/B, C4A, ATG4D and NEO1) of expression quantitative trait loci genes from our GWAS in the CSF proteome of anti-GAD65 AINS. These findings suggest a strong genetic predisposition with direct functional implications for immunity and neural function in anti-GAD65 AINS, mainly conferred by genomic regions outside the classical HLA alleles.

Elevated phospholipids and acylcarnitines C4 and C5 in cerebrospinal fluid distinguish viral CNS infections from autoimmune neuroinflammation.

BACKGROUND: Viral and autoimmune encephalitis may present with similar symptoms, but require different treatments. Thus, there is a need for biomarkers to improve diagnosis and understanding of pathogenesis. We hypothesized that virus-host cell interactions lead to different changes in central nervous system (CNS) metabolism than autoimmune processes and searched for metabolite biomarkers in cerebrospinal fluid (CSF) to distinguish between the two conditions. METHODS: We applied a targeted metabolomic/lipidomic analysis to CSF samples from patients with viral CNS infections (n = 34; due to herpes simplex virus [n = 9], varicella zoster virus [n = 15], enteroviruses [n = 10]), autoimmune neuroinflammation (n = 25; autoimmune anti-NMDA-receptor encephalitis [n = 8], multiple sclerosis [n = 17), and non-inflamed controls (n = 31; Gilles de la Tourette syndrome [n = 20], Bell's palsy with normal CSF cell count [n = 11]). 85 metabolites passed quality screening and were evaluated as biomarkers. Standard diagnostic CSF parameters were assessed for comparison. RESULTS: Of the standard CSF parameters, the best biomarkers were: CSF cell count for viral infections vs. controls (area under the ROC curve, AUC = 0.93), Q-albumin for viral infections vs. autoimmune neuroinflammation (AUC = 0.86), and IgG index for autoimmune neuroinflammation vs. controls (AUC = 0.90). Concentrations of 2 metabolites differed significantly (p < 0.05) between autoimmune neuroinflammation and controls, with proline being the best biomarker (AUC = 0.77). In contrast, concentrations of 67 metabolites were significantly higher in viral infections than controls, with SM.C16.0 being the best biomarker (AUC = 0.94). Concentrations of 68 metabolites were significantly higher in viral infections than in autoimmune neuroinflammation, and the 10 most accurate metabolite biomarkers (AUC = 0.89-0.93) were substantially better than Q-albumin (AUC = 0.86). These biomarkers comprised six phosphatidylcholines (AUC = 0.89-0.92), two sphingomyelins (AUC = 0.89, 0.91), and acylcarnitines isobutyrylcarnitine (C4, AUC = 0.92) and isovalerylcarnitine (C5, AUC = 0.93). Elevated C4 and C5 concentrations suggested dysfunctional mitochondrial ß-oxidation and correlated only moderately with CSF cell count (Spearman ρ = 0.41 and 0.44), indicating that their increase is not primarily driven by inflammation. CONCLUSIONS: Changes in CNS metabolism differ substantially between viral CNS infections and autoimmune neuroinflammation and reveal CSF metabolites as pathophysiologically relevant diagnostic biomarkers for the differentiation between the two conditions. In viral CNS infections, the observed higher concentrations of free phospholipids are consistent with disruption of host cell membranes, whereas the elevated short-chain acylcarnitines likely reflect compromised mitochondrial homeostasis and energy generation.

Quantification of polyreactive immunoglobulin G facilitates the diagnosis of autoimmune hepatitis.

BACKGROUND AND AIMS: Detection of autoantibodies is a mainstay of diagnosing autoimmune hepatitis (AIH). However, conventional autoantibodies for the workup of AIH lack either sensitivity or specificity, leading to substantial diagnostic uncertainty. We aimed to identify more accurate serological markers of AIH with a protein macroarray. APPROACH AND RESULTS: During the search for more-precise autoantibodies to distinguish AIH from non-AIH liver diseases (non-AIH-LD), IgG antibodies with binding capacities to many human and foreign proteins were identified with a protein macroarray and confirmed with solid-phase ELISAs in AIH patients. Subsequently, polyreactive IgG (pIgG) was exemplarily quantified by reactivity against human huntingtin-interacting protein 1-related protein in bovine serum albumin blocked ELISA (HIP1R/BSA). The diagnostic fidelity of HIP1R/BSA binding pIgG to diagnose AIH was assessed in a retrospective training, a retrospective multicenter validation, and a prospective validation cohort in cryoconserved samples from 1,568 adults from 10 centers from eight countries. Reactivity against HIP1R/BSA had a 25% and 14% higher specificity to diagnose AIH than conventional antinuclear and antismooth muscle antibodies, a significantly higher sensitivity than liver kidney microsomal antibodies and antisoluble liver antigen/liver pancreas antigen, and a 12%-20% higher accuracy than conventional autoantibodies. Importantly, HIP1R/BSA reactivity was present in up to 88% of patients with seronegative AIH and in up to 71% of AIH patients with normal IgG levels. Under therapy, pIgG returns to background levels of non-AIH-LD. CONCLUSIONS: pIgG could be used as a promising marker to improve the diagnostic workup of liver diseases with a higher specificity for AIH compared to conventional autoantibodies and a utility in autoantibody-negative AIH. Likewise, pIgG could be a major source of assay interference in untreated AIH.

Does [99mTc]-3,3-diphosphono-1,2-propanodicarboxylic acid (DPD) soft tissue uptake allow the identification of patients with the diagnosis of cardiac transthyretin-related (ATTR) amyloidosis with higher risk for polyneuropathy?

BACKGROUND: With the introduction of several drugs for the therapy of transthyretin-related amyloidosis (ATTR) which slow down the disease, early detection of polyneuropathy (PNP) is becoming increasingly of interest. [99mTc]-3,3-Diphosphono-1,2-Propanodicarboxylic Acid (DPD) bone scintigraphy, which is used for the diagnosis of cardiac (c)ATTR, can possibly make an important contribution in the identification of patients at risk for PNP. METHODS: Fifty patients with cATTR, who underwent both planar whole-body DPD scintigraphy and nerve conduction studies (NCS) were retrospectively evaluated. A subgroup of 22 patients also underwent quantitative SPECT/CT of the thorax from which Standardized Uptake Values (SUVpeak) in the subcutaneous fat tissue of the left axillar region were evaluated. RESULTS: The Perugini score was significantly increased in patients with cATTR and additional diagnosis of PNP compared to patients without (2.51 ± 0.51 vs 2.13 ± 0.52; P = 0.03). Quantitative SPECT/CT revealed that DPD uptake in the subcutaneous fat of the left axillar region was significantly increased in cATTR patients with compared to patients without (1.36 ± 0.60 vs 0.74 ± 0.52; P = 0.04). CONCLUSION: This study suggests that DPD bone scintigraphy is a useful tool for identification of patients with cATTR and a risk for PNP due to increased DPD soft tissue uptake.

Neurologic manifestations of COVID-19 in critically ill patients: results of the prospective multicenter registry PANDEMIC.

BACKGROUND: Neurologic manifestations are increasingly reported in patients with coronavirus disease 2019 (COVID-19). Yet, data on prevalence, predictors and relevance for outcome of neurological manifestations in patients requiring intensive care are scarce. We aimed to characterize prevalence, risk factors and impact on outcome of neurologic manifestations in critically ill COVID-19 patients. METHODS: In the prospective, multicenter, observational registry study PANDEMIC (Pooled Analysis of Neurologic DisordErs Manifesting in Intensive care of COVID-19), we enrolled COVID-19 patients with neurologic manifestations admitted to 19 German intensive care units (ICU) between April 2020 and September 2021. We performed descriptive and explorative statistical analyses. Multivariable models were used to investigate factors associated with disorder categories and their underlying diagnoses as well as to identify predictors of outcome. RESULTS: Of the 392 patients included in the analysis, 70.7% (277/392) were male and the mean age was 65.3 (SD ± 3.1) years. During the study period, a total of 2681 patients with COVID-19 were treated at the ICUs of 15 participating centers. New neurologic disorders were identified in 350 patients, reported by these centers, suggesting a prevalence of COVID-19-associated neurologic disorders of 12.7% among COVID-19 ICU patients. Encephalopathy (46.2%; 181/392), cerebrovascular (41.0%; 161/392) and neuromuscular disorders (20.4%; 80/392) were the most frequent categories identified. Out of 35 cerebrospinal fluid analyses with reverse transcriptase PCR for SARS-COV-2, only 3 were positive. In-hospital mortality was 36.0% (140/389), and functional outcome (mRS 3 to 5) of surviving patients was poor at hospital discharge in 70.9% (161/227). Intracerebral hemorrhage (OR 6.2, 95% CI 2.5-14.9, p < 0.001) and acute ischemic stroke (OR 3.9, 95% CI 1.9-8.2, p < 0.001) were the strongest predictors of poor outcome among the included patients. CONCLUSIONS: Based on this well-characterized COVID-19 ICU cohort, that comprised 12.7% of all severe ill COVID-19 patients, neurologic manifestations increase mortality and morbidity. Since no reliable evidence of direct viral affection of the nervous system by COVID-19 could be found, these neurologic manifestations may for a great part be indirect para- or postinfectious sequelae of the infection or severe critical illness. Neurologic ICU complications should be actively searched for and treated.

Phosphatidylcholine PC ae C44:6 in cerebrospinal fluid is a sensitive biomarker for bacterial meningitis.

BACKGROUND: The timely diagnosis of bacterial meningitis is of utmost importance due to the need to institute antibiotic treatment as early as possible. Moreover, the differentiation from other causes of meningitis/encephalitis is critical because of differences in management such as the need for antiviral or immunosuppressive treatments. Considering our previously reported association between free membrane phospholipids in cerebrospinal fluid (CSF) and CNS involvement in neuroinfections we evaluated phosphatidylcholine PC ae C44:6, an integral constituent of cell membranes, as diagnostic biomarker for bacterial meningitis. METHODS: We used tandem mass spectrometry to measure concentrations of PC ae C44:6 in cell-free CSF samples (n = 221) from patients with acute bacterial meningitis, neuroborreliosis, viral meningitis/encephalitis (herpes simplex virus, varicella zoster virus, enteroviruses), autoimmune neuroinflammation (anti-NMDA-receptor autoimmune encephalitis, multiple sclerosis), facial nerve and segmental herpes zoster (shingles), and noninflammatory CNS disorders (Bell's palsy, Tourette syndrome, normal pressure hydrocephalus). RESULTS: PC ae C44:6 concentrations were significantly higher in bacterial meningitis than in all other diagnostic groups, and were higher in patients with a classic bacterial meningitis pathogen (e.g. Streptococcus pneumoniae, Neisseria meningitidis, Staphylococcus aureus) than in those with less virulent or opportunistic pathogens as causative agents (P = 0.026). PC ae C44:6 concentrations were only moderately associated with CSF cell count (Spearman's ρ = 0.45; P = 0.009), indicating that they do not merely reflect neuroinflammation. In receiver operating characteristic curve analysis, PC ae C44:6 equaled CSF cell count in the ability to distinguish bacterial meningitis from viral meningitis/encephalitis and autoimmune CNS disorders (AUC 0.93 both), but had higher sensitivity (91% vs. 41%) and negative predictive value (98% vs. 89%). A diagnostic algorithm comprising cell count, lactate and PC ae C44:6 had a sensitivity of 97% (specificity 87%) and negative predictive value of 99% (positive predictive value 61%) and correctly diagnosed three of four bacterial meningitis samples that were misclassified by cell count and lactate due to low values not suggestive of bacterial meningitis. CONCLUSIONS: Increased CSF PC ae C44:6 concentrations in bacterial meningitis likely reflect ongoing CNS cell membrane stress or damage and have potential as additional, sensitive biomarker to diagnose bacterial meningitis in patients with less pronounced neuroinflammation.

Kynurenine Is a Cerebrospinal Fluid Biomarker for Bacterial and Viral Central Nervous System Infections.

BACKGROUND: The tryptophan-kynurenine-nicotinamide adenine dinucleotide (oxidized; NAD+) pathway is closely associated with regulation of immune cells toward less inflammatory phenotypes and may exert neuroprotective effects. Investigating its regulation in central nervous system (CNS) infections would improve our understanding of pathophysiology and end-organ damage, and, furthermore, open doors to its evaluation as a source of diagnostic and/or prognostic biomarkers. METHODS: We measured concentrations of kynurenine (Kyn) and tryptophan (Trp) in 221 cerebrospinal fluid samples from patients with bacterial and viral (due to herpes simplex, varicella zoster, and enteroviruses) meningitis/encephalitis, neuroborreliosis, autoimmune neuroinflammation (due to anti-N-methyl-D-aspartate receptor [NMDA] encephalitis and multiple sclerosis), and noninflamed controls (ie, individuals with Bell palsy, normal pressure hydrocephalus, or Tourette syndrome). RESULTS: Kyn concentrations correlated strongly with CSF markers of neuroinflammation (ie, leukocyte count, lactate concentration, and blood-CSF-barrier dysfunction), were highly increased in bacterial and viral CNS infections, but were low or undetectable in NMDA encephalitis, multiple sclerosis, and controls. Trp concentrations were decreased mostly in viral CNS infections and neuroborreliosis. Multiple logistic regression analysis revealed that combinations of Kyn concentration, Trp concentration, and Kyn/Trp concentration ratio with leukocyte count or lactate concentration were accurate classifiers for the clinically important differentiation between neuroborreliosis, viral CNS infections, and autoimmune neuroinflammation. CONCLUSIONS: The Trp-Kyn-NAD+ pathway is activated in CNS infections and provides highly accurate CSF biomarkers, particularly when combined with standard CSF indices of neuroinflammation.

Genetic predisposition in anti-LGI1 and anti-NMDA receptor encephalitis.

We performed a genome-wide association study in 1,194 controls and 150 patients with anti-N-methyl-D-aspartate receptor (anti-NMDAR, n = 96) or anti-leucine-rich glioma-inactivated1 (anti-LGI1, n = 54) autoimmune encephalitis. Anti-LGI1 encephalitis was highly associated with 27 single-nucleotide polymorphisms (SNPs) in the HLA-II region (leading SNP rs2858870 p = 1.22 × 10-17 , OR = 13.66 [7.50-24.87]). Potential associations, below genome-wide significance, were found with rs72961463 close to the doublecortin-like kinase 2 gene (DCLK2) and rs62110161 in a cluster of zinc-finger genes. HLA allele imputation identified association of anti-LGI1 encephalitis with HLA-II haplotypes encompassing DRB1*07:01, DQA1*02:01 and DQB1*02:02 (p < 2.2 × 10-16 ) and anti-NMDAR encephalitis with HLA-I allele B*07:02 (p = 0.039). No shared genetic risk factors between encephalitides were identified. Ann Neurol 2018;83:863-869.

Lipid nanoparticle-mediated siRNA delivery for safe targeting of human CML in vivo.

Efficient and safe delivery of siRNA in vivo is the biggest roadblock to clinical translation of RNA interference (RNAi)-based therapeutics. To date, lipid nanoparticles (LNPs) have shown efficient delivery of siRNA to the liver; however, delivery to other organs, especially hematopoietic tissues still remains a challenge. We developed DLin-MC3-DMA lipid-based LNP-siRNA formulations for systemic delivery against a driver oncogene to target human chronic myeloid leukemia (CML) cells in vivo. A microfluidic mixing technology was used to obtain reproducible ionizable cationic LNPs loaded with siRNA molecules targeting the BCR-ABL fusion oncogene found in CML. We show a highly efficient and non-toxic delivery of siRNA in vitro and in vivo with nearly 100% uptake of LNP-siRNA formulations in bone marrow of a leukemic model. By targeting the BCR-ABL fusion oncogene, we show a reduction of leukemic burden in our myeloid leukemia mouse model and demonstrate reduced disease burden in mice treated with LNP-BCR-ABL siRNA as compared with LNP-CTRL siRNA. Our study provides proof-of-principle that fusion oncogene specific RNAi therapeutics can be exploited against leukemic cells and promise novel treatment options for leukemia patients.

Identification of Cerebrospinal Fluid Metabolites as Biomarkers for Enterovirus Meningitis.

Enteroviruses are among the most common causes of viral meningitis. Enteroviral meningitis continues to represent diagnostic challenges, as cerebrospinal fluid (CSF) cell numbers (a well validated diagnostic screening tool) may be normal in up to 15% of patients. We aimed to identify potential CSF biomarkers for enteroviral meningitis, particularly for cases with normal CSF cell count. Using targeted liquid chromatography-mass spectrometry, we determined metabolite profiles from patients with enteroviral meningitis (n = 10), and subdivided them into those with elevated (n = 5) and normal (n = 5) CSF leukocyte counts. Non-inflamed CSF samples from patients with Bell's palsy and normal pressure hydrocephalus (n = 19) were used as controls. Analysis of 91 metabolites revealed considerable metabolic reprogramming in the meningitis samples. It identified phosphatidylcholine PC.ae.C36.3, asparagine, and glycine as an accurate (AUC, 0.92) combined classifier for enterovirus meningitis overall, and kynurenine as a perfect biomarker for enteroviral meningitis with an increased CSF cell count (AUC, 1.0). Remarkably, PC.ae.C36.3 alone emerged as a single accurate (AUC, 0.87) biomarker for enteroviral meningitis with normal cell count, and a combined classifier comprising PC.ae.C36.3, PC.ae.C36.5, and PC.ae.C38.5 achieved nearly perfect classification (AUC, 0.99). Taken together, this analysis reveals the potential of CSF metabolites as additional diagnostic tools for enteroviral meningitis, and likely other Central nervous system (CNS) infections.

Mass-spectrometric profiling of cerebrospinal fluid reveals metabolite biomarkers for CNS involvement in varicella zoster virus reactivation.

BACKGROUND: Varicella zoster virus (VZV) reactivation spans the spectrum from uncomplicated segmental herpes zoster to life-threatening disseminated CNS infection. Moreover, in the absence of a small animal model for this human pathogen, studies of pathogenesis at the organismal level depend on analysis of human biosamples. Changes in cerebrospinal fluid (CSF) metabolites may reflect critical aspects of host responses and end-organ damage in neuroinfection and neuroinflammation. We therefore applied a targeted metabolomics screen of CSF to three clinically distinct forms of VZV reactivation and infectious and non-infectious disease controls in order to identify biomarkers for CNS involvement in VZV reactivation. METHODS: Metabolite profiles were determined by targeted liquid chromatography-mass spectrometry in CSF from patients with segmental zoster (shingles, n = 14), facial nerve zoster (n = 16), VZV meningitis/encephalitis (n = 15), enteroviral meningitis (n = 10), idiopathic Bell's palsy (n = 11), and normal pressure hydrocephalus (n = 15). RESULTS: Concentrations of 88 metabolites passing quality assessment clearly separated the three VZV reactivation forms from each other and from the non-infected samples. Internal cross-validation identified four metabolites (SM C16:1, glycine, lysoPC a C26:1, PC ae C34:0) that were particularly associated with VZV meningoencephalitis. SM(OH) C14:1 accurately distinguished facial nerve zoster from Bell's palsy. Random forest construction revealed even more accurate classifiers (signatures comprising 2-4 metabolites) for most comparisons. Some of the most accurate biomarkers correlated only weakly with CSF leukocyte count, indicating that they do not merely reflect recruitment of inflammatory cells but, rather, specific pathophysiological mechanisms. Across all samples, only the sum of hexoses and the amino acids arginine, serine, and tryptophan correlated negatively with leukocyte count. Increased expression of the metabolites associated with VZV meningoencephalitis could be linked to processes relating to neuroinflammation/immune activation, neuronal signaling, and cell stress, turnover, and death (e.g., autophagy and apoptosis), suggesting that these metabolites might sense processes relating to end-organ damage. CONCLUSIONS: The results provide proof-of-concept for the value of CSF metabolites as (1) disease-associated signatures suggesting pathophysiological mechanisms, (2) degree and nature of neuroinflammation, and (3) biomarkers for diagnosis and risk stratification of VZV reactivation and, likely, neuroinfections due to other pathogens. TRIAL REGISTRATION: Not applicable (non-interventional study).

Varicella zoster virus infections in neurological patients: a clinical study.

BACKGROUND: Varicella zoster virus (VZV) reactivation is a common infectious disease in neurology and VZV the second most frequent virus detected in encephalitis. This study investigated characteristics of clinical and laboratory features in patients with VZV infection. METHODS: Two hundred eighty two patients with VZV reactivation that were hospitalized in the department of neurology in the time from 2005 to 2013 were retrospectively evaluated. Results from cerebrospinal fluid (CSF) analysis were available from 85 patients. RESULTS: Trigeminal rash was the most common clinical manifestation, followed by segmental rash, CNS infection, facial nerve palsy, postherpetic neuralgia, and radiculitis. MRI of the brain performed in 25/33 patients with encephalitis/meningitis did not show any signs of infection in the brain parenchyma. Only one patient showed contrast enhancement in the hypoglossal nerve. General signs of infection such as fever or elevated CRP values were found in only half of the patients. Furthermore, rash was absent in a quarter of patients with CNS infection and facial nerve palsy, and thus, infection could only be proven by CSF analysis. Although slight inflammatory CSF changes occurred in few patients with isolated rash, the frequency was clearly higher in patients with CNS infection and facial nerve palsy. CONCLUSION: Monosegmental herpes zoster is often uncomplicated and a diagnostic lumbar puncture is not essential. In contrast, CSF analysis is an essential diagnostic tool in patients with skin lesions and cranial nerve or CNS affection. In patients with neuro-psychiatric symptoms and inflammatory CSF changes analysis for VZV should be performed even in the absence of skin lesions.

Paraneoplastic cerebellar syndromes associated with antibodies against Purkinje cells.

The paraneoplastic cerebellar syndrome presents as severe neuroimmunological disease associated with malignancies. Antibodies against antigens expressed by tumor cells cross-react with proteins of cerebellar Purkinje cells leading to neuroinflammation and neuronal loss. These antineuronal antibodies are preferentially investigated by serological analyses while examination of the cerebrospinal fluid is only performed infrequently. We retrospectively investigated 12 patients with antineuronal antibodies against Purkinje cells with a special focus on cerebrospinal fluid. Our results confirm a subacute disease with a severe cerebellar syndrome in 10 female patients due to anti-Yo antibodies associated mostly with gynecological malignancies. While standard cerebrospinal fluid parameters infrequently revealed pathological results, all patients presented oligoclonal bands indicating intrathecal IgG synthesis. Analyses of anti-Yo antibodies in cerebrospinal fluid by calculating the antibody specific index revealed intrathecal synthesis of anti-Yo antibodies in these patients. In analogy to anti-Yo syndrome, an intrathecal production of anti-Tr antibodies in one patient who presented with a paraneoplastic cerebellar syndrome was detected. In an additional patient, anti-Purkinje cell antibodies of unknown origin in the cerebrospinal fluid but not in serum were determined suggesting an isolated immune reaction within the central nervous system (CNS) and underlining the importance of investigating the cerebrospinal fluid. In conclusion, patients with a cerebellar syndrome display a distinct immune reaction within the cerebrospinal fluid including intrathecal synthesis of disease-specific antibodies. We emphasize the importance of a thorough immunological work up including investigations of both serum and cerebrospinal fluid.

The Persisting Significance of Oligoclonal Bands in the Dawning Era of Kappa Free Light Chains for the Diagnosis of Multiple Sclerosis.

The latest revision of the McDonald criteria of 2017 considers the evidence of an intrathecal immunoglobulin (IgG) synthesis as a diagnostic criterion for dissemination in time in multiple sclerosis. While the detection of oligoclonal bands is considered as the gold standard, determination of kappa free light chains might be a promising tool as a less technically demanding and cost saving method. However, data on the direct comparison between kappa free light chains and oligoclonal bands are limited and no study to date has used the highly sensitive method of polyacrylamide gels with consecutive silver staining for the demonstration of oligoclonal bands. Furthermore, the impact of the revised McDonald criteria of 2017 on the role of kappa free light chains as a biomarker has not been investigated. Nephelometry was used to determine kappa free light chains in cerebrospinal fluid (CSF) and serum from 149 patients with their first demyelinating event between 2010 and 2015. Clinical data, kappa free light chains, and oligoclonal band status were compared at the time of initial diagnosis and after follow-up to identify converters from clinically isolated syndrome to multiple sclerosis. An elevated kappa free light chain index (>5.9) was found in 79/83 patients (95%) with multiple sclerosis diagnosed at baseline, slightly less frequent than oligoclonal bands (98.8%). 18/25 (72%) patients who converted from clinically isolated syndrome to multiple sclerosis showed an elevated kappa free light chain index compared to 20/25 (80%) patients with positive oligoclonal bands. In patients with stable clinically isolated syndrome 7/41 (17%) displayed an elevated kappa free light chain index against 11/41 (27%) oligoclonal band positive patients. Only two patients with stable clinically isolated syndrome showed an elevated kappa free light chain index but were oligoclonal bands negative. In conclusion, determination of the kappa free light chain index is a promising diagnostic approach to assess intrathecal immunoglobulin synthesis in multiple sclerosis. Nevertheless, oligoclonal bands are highly prevalent in multiple sclerosis and can detect an intrathecal synthesis of IgG even when the kappa free light chain index is below the threshold. We consider sequential use of both methods as reasonable.

Common and uncommon neurological manifestations of neuroborreliosis leading to hospitalization.

BACKGROUND: Neuroborreliosis represents a relevant infectious disease and can cause a variety of neurological manifestations. Different stages and syndromes are described and atypical symptoms can result in diagnostic delay or misdiagnosis. The aim of this retrospective study was to define the pivotal neurological deficits in patients with neuroborreliosis that were the reason for admission in a hospital. METHODS: We retrospectively evaluated data of patients with neuroborreliosis. Only patients who fulfilled the diagnostic criteria of an intrathecal antibody production against Borrelia burgdorferi were included in the study. RESULTS: Sixty-eight patients were identified with neuroborreliosis. Cranial nerve palsy was the most frequent deficit (50%) which caused admission to a hospital followed by painful radiculitis (25%), encephalitis (12%), myelitis (7%), and meningitis/headache (6%). In patients with a combination of deficits, back pain was the first symptom, followed by headache, and finally by cranial nerve palsy. Indeed, signs of meningitis were often found in patients with neuroborreliosis, but usually did not cause admission to a hospital. Unusual cases included patients with sudden onset paresis that were initially misdiagnosed as stroke and one patient with acute delirium. Cerebrospinal fluid (CSF) analysis revealed typical changes including elevated CSF cell count in all but one patient, a blood-CSF barrier dysfunction (87%), CSF oligoclonal bands (90%), and quantitative intrathecal synthesis of immunoglobulins (IgM in 74%, IgG in 47%, and IgA in 32% patients). Importantly, 6% of patients did not show Borrelia specific antibodies in the blood. CONCLUSION: In conclusion, the majority of patients presented with typical neurological deficits. However, unusual cases such as acute delirium indicate that neuroborreliosis has to be considered in a wide spectrum of neurological diseases. CSF analysis is essential for a reliable diagnosis of neuroborreliosis.

Cerebrospinal Fluid Findings in Neurological Diseases Associated with Sjögren's Syndrome.

BACKGROUND: Sjögren's syndrome is a chronic autoimmune-mediated disease that can cause a variety of neurological manifestations. METHODS: This study investigated characteristics of clinical and cerebrospinal fluid (CSF) features in patients with neurological diseases associated with Sjögren's syndrome. Eighty-two patients were examined separately according to the presence of Sjögren's syndrome alone or in combination with other autoimmune diseases. RESULTS: In the 47 patients with primary Sjögren's syndrome, peripheral neuropathy (57%) was found most frequently, followed by the involvement of the central nervous system (CNS; 17%), cranial neuropathy (15%), and myalgia (11%). These patients did not display consistent signs of inflammation in the CSF. Slight pleocytosis of 8-107 cells/µL was found in patients with peripheral neuropathy (9%), cranial neuropathy (20%), and CNS involvement (25%). Oligoclonal bands indicating intrathecal IgG synthesis occurred in 26% of patients with peripheral neuropathy, 20% of patients with cranial neuropathy, and 25% of patients with CNS involvement. CONCLUSIONS: In patients with Sjögren's syndrome and neurological manifestations, inflammatory CSF changes were rarely found and did not show a characteristic pattern irrespective of peripheral or central genesis of neurological deficits. Analysis of the CSF presents therefore an important diagnostic procedure to exclude other autoimmune and infectious diseases.

Clinically Isolated Syndrome According to McDonald 2010: Intrathecal IgG Synthesis Still Predictive for Conversion to Multiple Sclerosis.

While the revised McDonald criteria of 2010 allow for the diagnosis of multiple sclerosis (MS) in an earlier stage, there is still a need to identify the risk factors for conversion to MS in patients with clinically isolated syndrome (CIS). Since the latest McDonald criteria were established, the prognostic role of cerebrospinal fluid (CSF) and visual evoked potentials (VEP) in CIS patients is still poorly defined. We conducted a monocentric investigation including patients with CIS in the time from 2010 to 2015. Follow-ups of 120 patients revealed that 42% converted to MS. CIS patients with positive oligoclonal bands (OCB) were more than twice as likely to convert to MS as OCB negative patients (hazard ratio = 2.6). The probability to develop MS was even higher when a quantitative intrathecal IgG synthesis was detected (hazard ratio = 3.8). In patients with OCB, VEP did not add further information concerning the conversion rate to MS. In patients with optic neuritis and negative OCB, a significantly higher rate converted to MS when VEP were delayed. In conclusion, the detection of an intrathecal IgG synthesis increases the conversion probability to MS. Pathological VEP can help to predict the conversion rate to MS in patients with optic neuritis without an intrathecal IgG synthesis.

Deregulation of microRNA-181c in cerebrospinal fluid of patients with clinically isolated syndrome is associated with early conversion to relapsing-remitting multiple sclerosis.

BACKGROUND: MiRNA-181c, miRNA-633 and miRNA-922 have been reported to be deregulated in multiple sclerosis. OBJECTIVES: To investigate the association between miRNA-181c, miRNA-633 and miRNA-922 and conversion from clinically isolated syndrome (CIS) to relapsing-remitting multiple sclerosis (RRMS); and to compare microRNAs in cerebrospinal fluid (CSF) and serum with regard to dysfunction of the blood-CSF barrier. METHODS: CSF and serum miRNA-181c, miRNA-633 and miRNA-922 were retrospectively determined by quantitative real-time polymerase chain reaction in CIS patients with (CIS-RRMS) and without (CIS-CIS) conversion to RRMS within 1 year. RESULTS: Thirty of 58 CIS patients developed RRMS. Cerebrospinal fluid miRNA-922, serum miRNA-922 and cerebrospinal fluid miRNA-181c were significantly higher in CIS-RRMS compared to CIS-CIS (P=0.027, P=0.048, P=0.029, respectively). High levels of cerebrospinal fluid miRNA-181c were independently associated with conversion from CIS to RRMS in multivariate Cox regression analysis (hazard ratio 2.99, 95% confidence interval 1.41-6.34, P=0.005). A combination of high cerebrospinal fluid miRNA-181c, younger age and more than nine lesions on magnetic resonance imaging showed the highest specificity (96%) and positive predictive value (94%) for conversion from CIS to RRMS. MiRNA-181c was higher in serum than in cerebrospinal fluid (P <0.001), while miRNA-633 and miRNA-922 were no different in cerebrospinal fluid and serum. Cerebrospinal fluid/serum albumin quotients did not correlate with microRNAs in cerebrospinal fluid (all P>0.711). CONCLUSIONS: Cerebrospinal fluid miRNA-181c might serve as a biomarker for early conversion to RRMS. Moreover, our data suggest an intrathecal origin of microRNAs detected in the cerebrospinal fluid.