Prognostic significance of hedgehog signaling network-related gene expression in breast cancer patients.

Breast cancer continues to be a serious public health problem. The role of the hedgehog pathway in normal development of the mammary gland as well as in carcinogenesis and progression of breast cancer is the subject of intense investigation, revealing functional interactions with cell surface heparan sulfate. Nevertheless, its influence on breast cancer prognosis, and its relation to specific sulfation motifs in heparan sulfate have only been poorly studied in large patient cohorts. Using the public database KMplotter that includes gene expression and survival data of 3951 patients, we found that the higher expression of SHH, HHAT, PTCH1, GLI1, GLI2, and GLI3 positively influences breast cancer prognosis. Stratifying patients according to the expression of hormone receptors, histological grade, lymph node metastasis, and systemic therapy, we observed that GLI1, GLI2, and GLI3 expression, as well as co-expression of SHH and ELP1 were associated with worse relapse-free survival in patients with HER2-positive tumors. Moreover, GLI1 expression in progesterone receptor-negative tumors and GLI3 expression in grade 3 tumors correlated with poor prognosis. SHH, in a panel of cell lines representing different breast cancer subtypes, and HHAT, PTCH1, GLI1, GLI2, and GLI3 were mostly expressed in cell lines classified as HER2-positive and basal-like. Expression of SHH, HHAT, GLI2, and GLI3 was differentially affected by overexpression of the heparan sulfate sulfotransferases HS2ST1 and HS3ST2 in vitro. Although high HS2ST1 expression was associated with poor prognosis in KMplotter analysis, high levels of HS3ST2 were associated with a good prognosis, except for ER-positive breast cancer. We suggest the GLI transcription factors as possible markers for the diagnosis, treatment, and prognosis of breast cancer especially in HER2-positive tumors, but also in progesterone receptor-negative and grade-3 tumors. The pathway interaction and prognostic impact of specific heparan sulfate sulfotransferases provide novel perspectives regarding a therapeutical targeting of the hedgehog pathway in breast cancer.

Drebrin Autoantibodies in Patients with Seizures and Suspected Encephalitis.

OBJECTIVE: Assess occurrence of the dendritic spine scaffolding protein Drebrin as a pathophysiologically relevant autoantibody target in patients with recurrent seizures and suspected encephalitis as leading symptoms. METHODS: Sera of 4 patients with adult onset epilepsy and suspected encephalitis of unresolved etiology and equivalent results in autoantibody screening were subjected to epitope identification. We combined a wide array of approaches, ranging from immunoblotting, immunoprecipitation, mass spectrometry, subcellular binding pattern analyses in primary neuronal cultures, and immunohistochemistry in brains of wild-type and Drebrin knockout mice to in vitro analyses of impaired synapse formation, morphology, and aberrant neuronal excitability by antibody exposure. RESULTS: In the serum of a patient with adult onset epilepsy and suspected encephalitis, a strong signal at ∼70kDa was detected by immunoblotting, for which mass spectrometry revealed Drebrin as the putative antigen. Three other patients whose sera also showed strong immunoreactivity around 70kDa on Western blotting were also anti-Drebrin-positive. Seizures, memory impairment, and increased protein content in cerebrospinal fluid occurred in anti-Drebrin-seropositive patients. Alterations in cerebral magnetic resonance imaging comprised amygdalohippocampal T2-signal increase and hippocampal sclerosis. Diagnostic biopsy revealed T-lymphocytic encephalitis in an anti-Drebrin-seropositive patient. Exposure of primary hippocampal neurons to anti-Drebrin autoantibodies resulted in aberrant synapse composition and Drebrin distribution as well as increased spike rates and the emergence of burst discharges reflecting network hyperexcitability. INTERPRETATION: Anti-Drebrin autoantibodies define a chronic syndrome of recurrent seizures and neuropsychiatric impairment as well as inflammation of limbic and occasionally cortical structures. Immunosuppressant therapies should be considered in this disorder. ANN NEUROL 2020;87:869-884.

Improved early outcome prediction by MRI-based 3D tumor volume assessment in patients with CNS lymphomas.

BACKGROUND: Central nervous system lymphomas (CNSL) display remarkable clinical heterogeneity, yet accurate prediction of outcomes remains challenging. The IPCG criteria are widely used in routine practice for the assessment of treatment response. However, the value of the IPCG criteria for ultimate outcome prediction is largely unclear, mainly due to the uncertainty in delineating complete from partial responses during and after treatment. METHODS: We explored various MRI features including semi-automated 3D tumor volume measurements at different disease milestones and their association with survival in 93 CNSL patients undergoing curative-intent treatment. RESULTS: At diagnosis, patients with more than 3 lymphoma lesions, periventricular involvement, and high 3D tumor volumes showed significantly unfavorable PFS and OS. At first interim MRI during treatment, the IPCG criteria failed to discriminate outcomes in responding patients. Therefore, we randomized these patients into training and validation cohorts to investigate whether 3D tumor volumetry could improve outcome prediction. We identified a 3D tumor volume reduction of ≥97% as the optimal threshold for risk stratification (=3D early response, 3D_ER). Applied to the validation cohort, patients achieving 3D_ER had significantly superior outcomes. In multivariate analyses, 3D_ER was independently prognostic of PFS and OS. Finally, we leveraged prognostic information from 3D MRI features and circulating biomarkers to build a composite metric that further improved outcome prediction in CNSL. CONCLUSIONS: We developed semi-automated 3D tumor volume measurements as strong and independent early predictors of clinical outcomes in CNSL patients. These radiologic features could help improve risk stratification and help guide future treatment approaches.

Comprehensive genetic profiling and molecularly guided treatment for patients with primary CNS tumors.

Despite major advances in molecular profiling and classification of primary brain tumors, personalized treatment remains limited for most patients. Here, we explored the feasibility of individual molecular profiling and the efficacy of biomarker-guided therapy for adult patients with primary brain cancers in the real-world setting within the molecular tumor board Freiburg, Germany. We analyzed genetic profiles, personalized treatment recommendations, and clinical outcomes of 102 patients with 21 brain tumor types. Alterations in the cell cycle, BRAF, and mTOR pathways most frequently led to personalized treatment recommendations. Molecularly informed therapies were recommended in 71% and implemented in 32% of patients with completed molecular diagnostics. The disease control rate following targeted treatment was 50% and the overall response rate was 30%, with a progression-free survival 2/1 ratio of at least 1.3 in 31% of patients. This study highlights the efficacy of molecularly guided treatment and the need for biomarker-stratified trials in brain cancers.

Aberrant Naive CD4-Positive T Cell Differentiation in Systemic Juvenile Idiopathic Arthritis Committed to B Cell Help.

OBJECTIVE: Systemic juvenile idiopathic arthritis (JIA) features characteristics of autoinflammation and autoimmunity, culminating in chronic arthritis. In this study, we hypothesized that aberrant or incomplete polarization of T helper cells contributes to disease pathology. METHODS: Cells or serum samples were obtained from healthy controls (n = 72) and systemic JIA patients (n = 171). Isolated naive T helper cells were cultured under Th1, Th17, and T follicular helper (Tfh) or T peripheral helper (Tph)-polarizing conditions and were partly cocultured with allogenic memory B cells. Cell samples were then analyzed for surface marker, transcription factor, and cytokine expression, as well as plasmablast generation. Serum samples were subjected to multiplexed bead and self-antigen arrays and enzyme-linked immunosorbent assays, and all data were compared to retrospective RNA profiling analyses. RESULTS: Differentiation of systemic JIA-naive T helper cells toward Th1 cells resulted in low expression levels of interferon-γ (IFNγ) and eomesodermin, which was associated in part with disease duration. In contrast, developing Th1 cells in patients with systemic JIA were found to produce elevated levels of interleukin-21 (IL-21), which negatively correlated with cellular expression of IFNγ and eomesodermin. In both in vitro and ex vivo analyses, IL-21 together with programmed cell death 1 (PD-1), inducible T cell costimulator (ICOS), and CXCR5 expression induced naive T helper cells from systemic JIA patients to polarize toward a Tfh/Tph cell phenotype. Retrospective analysis of whole-blood RNA-sequencing data demonstrated that Bcl-6, a master transcription factor in Tfh/Tph cell differentiation, was overexpressed specifically in patients with systemic JIA. Naive T helper cells from systemic JIA patients which were stimulated in vitro promoted B cellular plasmablast generation, and self-antigen array data indicated that IgG reactivity profiles of patients with systemic JIA differed from those of healthy controls. CONCLUSION: In the pathogenesis of systemic JIA, skewing of naive T helper cell differentiation toward a Tfh/Tph cell phenotype may represent an echo of autoimmunity, which may indicate the mechanisms driving progression toward chronic destructive arthritis.

Adult-onset temporal lobe epilepsy suspicious for autoimmune pathogenesis: Autoantibody prevalence and clinical correlates.

Temporal lobe adult-onset seizures (TAOS) related to autoimmunity represent an increasingly recognized disease syndrome within the spectrum of epilepsies. In this context, certain autoantibodies (autoABs) were often associated with limbic encephalitis (LE). Here, we aimed to gain insights into (a) the distribution of 'neurological' autoABs (neuroABs, defined as autoABs targeting neuronal surface structures or 'onconeuronal' ABs or anti-glutamate acid decarboxylase 65 (GAD65) autoABs) in a large consecutive TAOS patient cohort, to characterize (b) clinical profiles of seropositive versus seronegative individuals and to find (c) potential evidence for other autoABs. Blood sera/cerebrospinal fluid (CSF) of TAOS patients (n = 800) and healthy donors (n = 27) were analyzed for neuroABs and screened for other autoABs by indirect immunofluorescence on hippocampal/cerebellar sections and immunoblots of whole brain and synaptosome lysates. Serological results were correlated with clinico-neuropsychological features. 13% of TAOS patients (n = 105) were neuroAB+, with anti-GAD65 and anti-N-methyl-D-aspartate receptors (NMDAR) as most frequent autoABs in this group. In our screening tests 25% of neuroAB- patients (n = 199) were positive (screening+), whereas all control samples were negative (n = 27). Intriguingly, key clinico-neuropsychological characteristics including magnetic resonance imaging (MRI) findings, epileptiform electroencephalographic (EEG) activity, and inflammatory cellular infiltrates in CSF were shared to a greater extent by neuroAB+ with neuroAB-/screening+ patients than with neuroAB-/screening- patients. Serological testing in a large consecutive TAOS patient series revealed seropositivity for anti-GAD65 autoABs as the most frequent neuroAB. Intriguingly, neuroAB+ individuals were virtually indistinguishable from neuroAB-/screening+ patients in several major clinical features. In contrast, neuroAB-/screening- TAOS patients differed in many parameters. These data support the potential presence of so far unrecognized autoABs in patients with TAOS.

Anti-epileptogenic and Anti-convulsive Effects of Fingolimod in Experimental Temporal Lobe Epilepsy.

Temporal lobe epilepsy (TLE) represents a devastating neurological condition, in which approximately 4/5 of patients remain refractory for anti-convulsive drugs. Epilepsy surgery biopsies often reveal the damage pattern of "hippocampal sclerosis" (HS) characterized not only by neuronal loss but also pronounced astrogliosis and inflammatory changes. Since TLE shares distinct pathogenetic aspects with multiple sclerosis (MS), we have here scrutinized therapeutic effects in experimental TLE of the immunmodulator fingolimod, which is established in MS therapy. Fingolimod targets sphingosine-phosphate receptors (S1PRs). mRNAs of fingolimod target S1PRs were augmented in two experimental post status epilepticus (SE) TLE mouse models (suprahippocampal kainate/pilocarpine). SE frequently induces chronic recurrent seizures after an extended latency referred to as epileptogenesis. Transient fingolimod treatment of mice during epileptogenesis after suprahippocampal kainate-induced SE revealed substantial reduction of chronic seizure activity despite lacking acute attenuation of SE itself. Intriguingly, fingolimod exerted robust anti-convulsive activity in kainate-induced SE mice treated in the chronic TLE stage and had neuroprotective and anti-gliotic effects and reduced cytotoxic T cell infiltrates. Finally, the expression profile of fingolimod target-S1PRs in human hippocampal biopsy tissue of pharmacoresistant TLE patients undergoing epilepsy surgery for seizure relief suggests repurposing of fingolimod as novel therapeutic perspective in focal epilepsies.