An oligodendrocyte silencer element underlies the pathogenic impact of lamin B1 structural variants.

The role of non-coding regulatory elements and how they might contribute to tissue type specificity of disease phenotypes is poorly understood. Autosomal Dominant Leukodystrophy (ADLD) is a fatal, adult-onset, neurological disorder that is characterized by extensive CNS demyelination. Most cases of ADLD are caused by tandem genomic duplications involving the lamin B1 gene ( LMNB1 ) while a small subset are caused by genomic deletions upstream of the gene. Utilizing data from recently identified families that carry LMNB1 gene duplications but do not exhibit demyelination, ADLD patient tissues, CRISPR modified cell lines and mouse models, we have identified a novel silencer element that is lost in ADLD patients and that specifically targets overexpression to oligodendrocytes. This element consists of CTCF binding sites that mediate three-dimensional chromatin looping involving the LMNB1 and the recruitment of the PRC2 repressor complex. Loss of the silencer element in ADLD identifies a previously unknown role for silencer elements in tissue specificity and disease causation.

STING-Triggered CNS Inflammation in Human Neurodegenerative Diseases.

BACKGROUND: Some neurodegenerative diseases have an element of neuroinflammation that is triggered by viral nucleic acids, resulting in the generation of type I interferons. In the cGAS-STING pathway, microbial and host-derived DNA bind and activate the DNA sensor cGAS, and the resulting cyclic dinucleotide, 2'3-cGAMP, binds to a critical adaptor protein, stimulator of interferon genes (STING), which leads to activation of downstream pathway components. However, there is limited work demonstrating the activation of the cGAS-STING pathway in human neurodegenerative diseases. METHODS: Post-mortem CNS tissue from donors with multiple sclerosis (n = 4), Alzheimer's disease (n = 6), Parkinson's disease (n = 3), amyotrophic lateral sclerosis (n = 3) and non-neurodegenerative controls (n = 11) were screened by immunohistochemistry for STING and relevant protein aggregates (e.g., amyloid-ß, α-synuclein, TDP-43). Human brain endothelial cells were cultured and stimulated with the STING agonist palmitic acid (1-400 µM) and assessed for mitochondrial stress (release of mitochondrial DNA into cytosol, increased oxygen consumption), downstream regulator factors, TBK-1/pIRF3 and inflammatory biomarker interferon-ß release and changes in ICAM-1 integrin expression. RESULTS: In neurodegenerative brain diseases, elevated STING protein was observed mainly in brain endothelial cells and neurons, compared to non-neurodegenerative control tissues where STING protein staining was weaker. Interestingly, a higher STING presence was associated with toxic protein aggregates (e.g., in neurons). Similarly high STING protein levels were observed within acute demyelinating lesions in multiple sclerosis subjects. To understand non-microbial/metabolic stress activation of the cGAS-STING pathway, brain endothelial cells were treated with palmitic acid. This evoked mitochondrial respiratory stress up to a ~2.5-fold increase in cellular oxygen consumption. Palmitic acid induced a statistically significant increase in cytosolic DNA leakage from endothelial cell mitochondria (Mander's coefficient; p < 0.05) and a significant increase in TBK-1, phosphorylated transcription factor IFN regulatory factor 3, cGAS and cell surface ICAM. In addition, a dose response in the secretion of interferon-ß was observed, but it failed to reach statistical significance. CONCLUSIONS: The histological evidence shows that the common cGAS-STING pathway appears to be activated in endothelial and neural cells in all four neurodegenerative diseases examined. Together with the in vitro data, this suggests that the STING pathway might be activated via perturbation of mitochondrial stress and DNA leakage, resulting in downstream neuroinflammation; hence, this pathway may be a target for future STING therapeutics.

The role of astrocytes in brain metastasis at the interface of circulating tumour cells and the blood brain barrier.

Tumour metastasis to the brain is a complex process involving crosstalk between the circulating tumour cells and the blood brain barrier (BBB). Astrocytes, which reside in the abluminal surface of the microvasculature of the BBB, are now known to play an essential role in tumour cell migration and invasion into the brain parenchyma. For instance, pro-inflammatory astrocyte secretions, including TNF-α, IL-6, CXCL10 as well as polyunsaturated fatty acids interact with circulating tumour cells to promote migration and proliferation. Additionally, astrocyte and tumour cell derived MMPs play a vital role in tumour cell invasion through the BBB. Understanding these complex interactions between tumour cells and astrocytes in the tumour microenvironment may contribute to the development of novel therapeutics for brain metastasis. Therefore, in this review, we present key interactions within the neurovascular unit of the BBB in the tumour microenvironment that significantly aids cancer metastasis, focusing particularly on astrocytes.

An ancestral 10-bp repeat expansion in VWA1 causes recessive hereditary motor neuropathy.

The extracellular matrix comprises a network of macromolecules such as collagens, proteoglycans and glycoproteins. VWA1 (von Willebrand factor A domain containing 1) encodes a component of the extracellular matrix that interacts with perlecan/collagen VI, appears to be involved in stabilizing extracellular matrix structures, and demonstrates high expression levels in tibial nerve. Vwa1-deficient mice manifest with abnormal peripheral nerve structure/function; however, VWA1 variants have not previously been associated with human disease. By interrogating the genome sequences of 74 180 individuals from the 100K Genomes Project in combination with international gene-matching efforts and targeted sequencing, we identified 17 individuals from 15 families with an autosomal-recessive, non-length dependent, hereditary motor neuropathy and rare biallelic variants in VWA1. A single disease-associated allele p.(G25Rfs*74), a 10-bp repeat expansion, was observed in 14/15 families and was homozygous in 10/15. Given an allele frequency in European populations approaching 1/1000, the seven unrelated homozygote individuals ascertained from the 100K Genomes Project represents a substantial enrichment above expected. Haplotype analysis identified a shared 220 kb region suggesting that this founder mutation arose >7000 years ago. A wide age-range of patients (6-83 years) helped delineate the clinical phenotype over time. The commonest disease presentation in the cohort was an early-onset (mean 2.0 ± 1.4 years) non-length-dependent axonal hereditary motor neuropathy, confirmed on electrophysiology, which will have to be differentiated from other predominantly or pure motor neuropathies and neuronopathies. Because of slow disease progression, ambulation was largely preserved. Neurophysiology, muscle histopathology, and muscle MRI findings typically revealed clear neurogenic changes with single isolated cases displaying additional myopathic process. We speculate that a few findings of myopathic changes might be secondary to chronic denervation rather than indicating an additional myopathic disease process. Duplex reverse transcription polymerase chain reaction and immunoblotting using patient fibroblasts revealed that the founder allele results in partial nonsense mediated decay and an absence of detectable protein. CRISPR and morpholino vwa1 modelling in zebrafish demonstrated reductions in motor neuron axonal growth, synaptic formation in the skeletal muscles and locomotive behaviour. In summary, we estimate that biallelic variants in VWA1 may be responsible for up to 1% of unexplained hereditary motor neuropathy cases in Europeans. The detailed clinical characterization provided here will facilitate targeted testing on suitable patient cohorts. This novel disease gene may have previously evaded detection because of high GC content, consequential low coverage and computational difficulties associated with robustly detecting repeat-expansions. Reviewing previously unsolved exomes using lower QC filters may generate further diagnoses.

Engagement of people with multiple sclerosis to enhance research into the physiological effect of hyperbaric oxygen therapy.

BACKGROUND: Thousands of people with multiple sclerosis (MS) have used self-administered oxygen therapy in the UK. Clinical trials have been performed, with scant evidence that people with MS have been consulted to explore how they benefit from or how to optimize this treatment. The conventional MS disease disability scores used in trials seldom reflect the effects individuals report when using oxygen therapy to treat their symptoms. METHODS: Three people with MS and the manager of an MS Centre formed a public involvement group and collaborated with clinicians and scientists to inform a lab-based study to investigate the physiological effects of oxygen therapy on microvascular brain endothelial cells. RESULTS: People with MS often use oxygen therapy at a later stage when their symptoms worsen and only after using other treatments. The frequency of oxygen therapy sessions and hyperbaric pressure is individualized and varies for people with MS. Despite direct comparisons of efficacy proving difficult, most individuals are exposed to 100% O2 at 1.5 atmosphere absolute (ATA; 1140 mmHg absolute) for 60 min. In a laboratory-based study human brain endothelial cells were exposed in vitro to 152 mmHg O2 for 60 min with and without pressure, as this equates to 20% O2 achievable via hyperbarics, which was then replicated at atmospheric pressure. A significant reduction in endothelial cells ICAM-1 (CD54) implicated in inflammatory cell margination across the blood brain barrier was observed under oxygen treatment. CONCLUSIONS: By collaborating with people living with MS, we were able to design laboratory-based experimental protocols that replicate their treatment regimens to advance our understanding of the physiological effects of hyperbaric oxygen treatment on brain cells and their role in neuroinflammation.

Clinical and Genetic Features in a Series of Eight Unrelated Patients with Neuropathy Due to Glycyl-tRNA Synthetase (GARS) Variants.

Pathogenic variants in the Glycyl-tRNA synthetase gene cause the allelic disorders Charcot-Marie-Tooth disease type 2D and distal hereditary motor neuropathy type V. We describe clinical features in 8 unrelated patients found to have Glycyl-tRNA synthetase variants by Next Generation Sequencing. In addition to upper limb predominant symptoms, other presentations included failure to thrive, feeding difficulties and lower limb dominant symptoms. Variability in the age at testing ranged from 14 months to 59 years. The youngest being symptomatic from 3 months and ventilator-dependent. Sequence variants were reported as pathogenic, p.(Glu125Lys), p.(His472Arg); likely pathogenic, p.(His216Arg), p.(Gly327Arg), p.(Lys510Gln), p.(Met555Val); and of uncertain significance, p.(Arg27Pro). Our case series describes novel Glycyl-tRNA synthetase variants and demonstrates the clinical utility of Next Generation Sequencing testing for patients with hereditary neuropathy. Identification of novel variants by Next Generation Sequencing illustrates that there exists a wide spectrum of clinical features and supports the newer simplified classification of neuropathies.

Astrocyte senescence may drive alterations in GFAPα, CDKN2A p14ARF, and TAU3 transcript expression and contribute to cognitive decline.

The accumulation of senescent cells in tissues is causally linked to the development of several age-related diseases; the removal of senescent glial cells in animal models prevents Tau accumulation and cognitive decline. Senescent cells can arise through several distinct mechanisms; one such mechanism is dysregulation of alternative splicing. In this study, we characterised the senescent cell phenotype in primary human astrocytes in terms of SA-ß-Gal staining and SASP secretion, and then assessed splicing factor expression and candidate gene splicing patterns. Finally, we assessed associations between expression of dysregulated isoforms and premature cognitive decline in 197 samples from the InCHIANTI study of ageing, where expression was present in both blood and brain. We demonstrate here that senescent astrocytes secrete a modified SASP characterised by increased IL8, MMP3, MMP10, and TIMP2 but decreased IL10 levels. We identified significant changes in splicing factor expression for 10/20 splicing factors tested in senescent astrocytes compared with early passage cells, as well as dysregulation of isoform levels for 8/13 brain or senescence genes tested. Finally, associations were identified between peripheral blood GFAPα, TAU3, and CDKN2A (P14ARF) isoform levels and mild or severe cognitive decline over a 3-7-year period. Our data are suggestive that some of the features of cognitive decline may arise from dysregulated splicing of important genes in senescent brain support cells, and that defects in alternative splicing or splicing regulator expression deserve exploration as points of therapeutic intervention in the future.

Cathepsin L-induced galectin-1 may act as a proangiogenic factor in the metastasis of high-grade serous carcinoma.

BACKGROUND: New treatment options for metastasised high-grade serous carcinoma (HGSC) are urgently needed. HGSC frequently metastasises to the omentum, inducing angiogenesis in the local omental microvasculature to facilitate tumour growth. We previously showed that HGSC-secreted cathepsin L (CathL) induces pro-angiogenic changes in disease relevant human omental microvascular endothelial cells (HOMECs), suggesting a role in tumour angiogenesis. Here we investigate whether CathL acts by inducing local production of the carbohydrate-binding protein galectin-1 (Gal1), which has been reported to be involved in tumourigenesis in other tumours. METHODS: HOMECs were used for all experiments. Gal1 mRNA and protein levels were measured by RT-PCR and ELISA respectively. Gal1-induced cell proliferation was assessed using WST-1 assay, migration using a transwell assay and in vivo Gal1 expression by immunohistochemistry. RESULTS: CathL transcriptionally regulated HOMEC production and secretion of Gal1 via activation of NFκB (significantly inhibited by sulfasalazine). Gal1 significantly enhanced HOMEC migration (p < 0.001) and proliferation (p < 0.001), suggesting an autocrine action. The latter was significantly reduced by the MEK/ERK1/2 inhibitors U0126 and PD98059 suggesting downstream activation of this pathway. Immunohistochemical analysis of omenta from HGSC patients with or without metastatic disease demonstrated a positive correlation between Gal1 expression and number of microvessels (r = 0.8702, p < 0.001), and area of vessels (r = 0.7283, p < 0.001), supporting a proangiogenic role for Gal1 in omental metastases. CONCLUSION: HOMEC Gal1 transcription and release in response to CathL secreted from metastasising HGSC acts in an autocrine manner on the local microvasculature to induce pro-angiogenic changes, highlighting a potential new therapeutic target.

Cathepsin L Induces Proangiogenic Changes in Human Omental Microvascular Endothelial Cells via Activation of the ERK1/2 Pathway.

BACKGROUND: Metastasis still remains the major cause of therapeutic failure, poor prognosis and high mortality in epithelial ovarian cancer (EOC) patients. Previously, we showed that EOC cells secrete a range of factors with potential pro-angiogenic activity, in disease-relevant human omental microvascular endothelial cells (HOMECs), including the lysosomal protease cathepsin L (CathL). Thus, the aim of this study was to examine potential pro-proliferative and pro-migratory effects of CathL in HOMECs and the activated signalling pathways, and whether these proangiogenic responses are dependent on CathL-catalytic activity. METHODS: HOMECs proliferation was investigated using WST-1, BrdU and CyQUANT assays. Cell migration was examined using a Cultrex Cell 96 transwell migration assay. Enzyme activity was assayed at various pHs using the CathL-specific fluorogenic substrate FY-CHO. Activation of cell signalling pathways was tested using a commercially available phosphokinase array and intact cellbased ELISAs. RESULTS: We showed for the first time that CathL has a potent pro-proliferative and pro-migratory effect on HOMECs. For instance, CathL significantly increases HOMEC proliferation (134.8±14.7% vs control 100%) and migration (146.6±17.3% vs control 100%). Our data strongly suggest that these proangiogenic effects of CathL are mediated via a non-proteolytic mechanism. Finally, we show that CathL-induced activation of the ERK1/2 pathway is involved in inducing these cellular effects in HOMECs. CONCLUSION: These data suggest that CathL acts as an extracellular ligand and plays an important pro-angiogenic, and thus pro-metastatic, role during EOC metastasis to the omentum, by activating the omental microvasculature, and thus can potentially be targeted therapeutically in the future.

Multifocal necrotising leucoencephalopathy following Salmonella infection in an immunocompetent patient.

Multifocal necrotising leucoencephalopathy is a rare disorder affecting the central nervous system. It is characterised pathologically by microscopic areas of necrosis with pontine predilection but also involvement of extrapontine regions, including the cerebellum, medulla and cerebral hemispheres. It usually occurs on the background of immunosuppression. Here we describe an immunocompetent patient with a recent history of Salmonella infection who presented with subacute neurological deterioration. At postmortem, she had evidence of multifocal necrotising leucoencephalopathy.

Cathepsin D non-proteolytically induces proliferation and migration in human omental microvascular endothelial cells via activation of the ERK1/2 and PI3K/AKT pathways.

Epithelial ovarian cancer (EOC) frequently metastasises to the omentum, a process that requires pro-angiogenic activation of human omental microvascular endothelial cells (HOMECs) by tumour-secreted factors. We have previously shown that ovarian cancer cells secrete a range of factors that induce pro-angiogenic responses e.g. migration, in HOMECs including the lysosomal protease cathepsin D (CathD). However, the cellular mechanism by which CathD induces these cellular responses is not understood. The aim of this study was to further examine the pro-angiogenic effects of CathD in HOMECs i.e. proliferation and migration, to investigate whether these effects are dependent on CathD catalytic activity and to delineate the intracellular signalling kinases activated by CathD. We report, for the first time, that CathD significantly increases HOMEC proliferation and migration via a non-proteolytic mechanism resulting in activation of ERK1/2 and AKT. These data suggest that EOC cancer secreted CathD acts as an extracellular ligand and may play an important pro-angiogenic, and thus pro-metastatic, role by activating the omental microvasculature during EOC metastasis to the omentum.

Manipulation of Oxygen and Endoplasmic Reticulum Stress Factors as Possible Interventions for Treatment of Multiple Sclerosis: Evidence for and Against.

Multiple sclerosis (MS) is normally considered a chronic inflammatory disease of the central nervous system (CNS), where T-cells breaching the blood brain barrier react against proteins of the axonal myelin sheaths, leading to focal plaques and demyelination in the brain and spinal cord. Many current therapies are immunosuppressive in nature and are designed to target the immune system at an early stage of the disease. But there is no cure and MS may evolve into a neurodegenerative disease, where immunomodulatory treatments appear less effective. Neurodegeneration is influenced by oxidative and endoplasmic reticulum (ER) mediated stress which can be induced independently of immune processes. Since 1970, MS patients have been self-managing their long term symptoms using hyperbaric oxygen and reporting improvement in their symptoms, especially bladder control. In contrast, the majority of clinical trial evidence does not support the views of patients. Therefore does oxygen under pressure affect brain tissue by modulating oxidative or ER stress at the cellular level resulting in CNS tissue repair or deterioration? This chapter reviews our understanding and the role of oxidative and ER stress in the context of employing hyperoxia treatments to treat MS and evaluate its effects on neural cells.

Rab32 connects ER stress to mitochondrial defects in multiple sclerosis.

BACKGROUND: Endoplasmic reticulum (ER) stress is a hallmark of neurodegenerative diseases such as multiple sclerosis (MS). However, this physiological mechanism has multiple manifestations that range from impaired clearance of unfolded proteins to altered mitochondrial dynamics and apoptosis. While connections between the triggering of the unfolded protein response (UPR) and downstream mitochondrial dysfunction are poorly understood, the membranous contacts between the ER and mitochondria, called the mitochondria-associated membrane (MAM), could provide a functional link between these two mechanisms. Therefore, we investigated whether the guanosine triphosphatase (GTPase) Rab32, a known regulator of the MAM, mitochondrial dynamics, and apoptosis, could be associated with ER stress as well as mitochondrial dysfunction. METHODS: We assessed Rab32 expression in MS patient and experimental autoimmune encephalomyelitis (EAE) tissue, via observation of mitochondria in primary neurons and via monitoring of survival of neuronal cells upon increased Rab32 expression. RESULTS: We found that the induction of Rab32 and other MAM proteins correlates with ER stress proteins in MS brain, as well as in EAE, and occurs in multiple central nervous system (CNS) cell types. We identify Rab32, known to increase in response to acute brain inflammation, as a novel unfolded protein response (UPR) target. High Rab32 expression shortens neurite length, alters mitochondria morphology, and accelerates apoptosis/necroptosis of human primary neurons and cell lines. CONCLUSIONS: ER stress is strongly associated with Rab32 upregulation in the progression of MS, leading to mitochondrial dysfunction and neuronal death.

Mutations in the histone methyltransferase gene KMT2B cause complex early-onset dystonia.

Histone lysine methylation, mediated by mixed-lineage leukemia (MLL) proteins, is now known to be critical in the regulation of gene expression, genomic stability, cell cycle and nuclear architecture. Despite MLL proteins being postulated as essential for normal development, little is known about the specific functions of the different MLL lysine methyltransferases. Here we report heterozygous variants in the gene KMT2B (also known as MLL4) in 27 unrelated individuals with a complex progressive childhood-onset dystonia, often associated with a typical facial appearance and characteristic brain magnetic resonance imaging findings. Over time, the majority of affected individuals developed prominent cervical, cranial and laryngeal dystonia. Marked clinical benefit, including the restoration of independent ambulation in some cases, was observed following deep brain stimulation (DBS). These findings highlight a clinically recognizable and potentially treatable form of genetic dystonia, demonstrating the crucial role of KMT2B in the physiological control of voluntary movement.

The Potential Role of the Proteases Cathepsin D and Cathepsin L in the Progression and Metastasis of Epithelial Ovarian Cancer.

Epithelial ovarian cancer (EOC) is the leading cause of death from gynecologic malignancies and has a poor prognosis due to relatively unspecific early symptoms, and thus often advanced stage, metastasized cancer at presentation. Metastasis of EOC occurs primarily through the transcoelomic route whereby exfoliated tumor cells disseminate within the abdominal cavity, particularly to the omentum. Primary and metastatic tumor growth requires a pool of proangiogenic factors in the microenvironment which propagate new vasculature in the growing cancer. Recent evidence suggests that proangiogenic factors other than the widely known, potent angiogenic factor vascular endothelial growth factor may mediate growth and metastasis of ovarian cancer. In this review we examine the role of some of these alternative factors, specifically cathepsin D and cathepsin L.

Loss of the endothelial glycocalyx is associated with increased E-selectin mediated adhesion of lung tumour cells to the brain microvascular endothelium.

BACKGROUND: Arrest of metastasising lung cancer cells to the brain microvasculature maybe mediated by interactions between ligands on circulating tumour cells and endothelial E-selectin adhesion molecules; a process likely to be regulated by the endothelial glycocalyx. Using human cerebral microvascular endothelial cells and non-small cell lung cancer (NSCLC) cell lines, we describe how factors secreted by NSCLC cells i.e. cystatin C, cathepsin L, insulin-like growth factor-binding protein 7 (IGFBP7), vascular endothelial growth factor (VEGF) and tumour necrosis factor-alpha (TNF-α), damage the glycocalyx and enhance initial contacts between lung tumour and cerebral endothelial cells. METHODS: Endothelial cells were treated with tumour secreted-proteins or lung tumour conditioned medium (CM). Surface levels of E-selectin were quantified by ELISA. Adhesion of A549 and SK-MES-1 cells was examined under flow conditions (1 dyne/cm(2)). Alterations in the endothelial glycocalyx were quantified by binding of fluorescein isothiocyanate-linked wheat germ agglutinin (WGA-FITC). RESULTS: A549 and SK-MES-1 CM and secreted-proteins significantly enhanced endothelial surface E-selectin levels after 30 min and 4 h and tumour cell adhesion after 30 min, 4 and 24 h. Both coincided with significant glycocalyx degradation; A549 and SK-MES-1 CM removing 55 ± 12 % and 58 ± 18.7 % of WGA-FITC binding, respectively. Inhibition of E-selectin binding by monoclonal anti-E-selectin antibody completely attenuated tumour cell adhesion. CONCLUSION: These data suggest that metastasising lung cancer cells facilitate their own adhesion to the brain endothelium by secreting factors that damage the endothelial glycocalyx, resulting in exposure of the previously shielded adhesion molecules and engagement of the E-selectin-mediated adhesion axis.

The difficulty in diagnosing X linked adrenoleucodystrophy and the importance of identifying cerebral involvement.

Two patients are described, a mother and son, who were initially clinically diagnosed with hereditary spastic paraparesis. This was rectified after very long chain fatty acid testing confirmed adrenomyeloneuropathy (AMN). The son's initial symptoms were characteristic of AMN (the commonest phenotype) but progressed to show symptoms of cerebral involvement. This evolution from non-cerebral to cerebral AMN is recognised in the medical literature and is increasingly important to consider in light of the availability of potential treatments such as haematopoietic stem cell transplantation.

Clinical Relevance of Increased Endothelial and Mesothelial Expression of Proangiogenic Proteases and VEGFA in the Omentum of Patients with Metastatic Ovarian High-Grade Serous Carcinoma.

Epithelial ovarian cancer (EOC) metastasis to the omentum requires implantation and angiogenesis. We propose that prometastatic changes in the omental endothelium (for angiogenesis) and mesothelium (for implantation) are critical. We investigated the expression of angiogenic proteases [cathepsin D (CD), cathepsin L (CL), and matrix metalloproteinase 2 (MMP2) and MMP9] and vascular endothelial growth factor A (VEGFA) in the mesothelium and endothelium of omentum from patients with EOC with omental metastases and control patients with benign ovarian tumors. Endothelial expression of CL, VEGFA, and MMP9 and mesothelial expression of VEGFA, MMP9, and CD were significantly increased in patients with metastasized EOC. High expression of MMP9 and VEGFA in endothelium and mesothelium and CD in mesothelium was positively associated with poor disease-specific survival (DSS). High MMP9 expression in either endothelium or mesothelium and presence of ascites prospectively showed the greatest risk of shorter DSS [hazard ratio (HR)= 6.16, 95% confidence interval (CI) = 1.76-21.6, P = .0045; HR = 11.42, 95% CI = 2.59-50.35, P = .0013; and HR = 6.35, 95% CI = 2.01-20.1, P = .002, respectively]. High endothelial MMP9 expression and ascites were independent predictors of reduced DSS and overall survival, together resulting in worst patient prognosis. Our data show that omental metastasis of EOC is associated with increased proangiogenic protein expression in the omental endothelium and mesothelium.

CD20+inflammatory T-cells are present in blood and brain of multiple sclerosis patients and can be selectively targeted for apoptotic elimination.

BACKGROUND: A subset of T-cells expresses the B-cell marker CD20 and in rheumatoid arthritis secretes Interleukin (IL)-17. IL-17 secreting T-cells (Th17) have also been implicated in the inflammatory response in the central nervous system in multiple sclerosis (MS) and may be a potential target for elimination by biologic therapeutics. ScFvRit:sFasL comprises of a rituximab-derived antibody fragment scFvRit genetically fused to human soluble FasL that specifically eliminated T-cells. OBJECTIVE: To determine the presence and phenotype of CD20+T-cells in blood and brain of MS patients. Second, to determine whether scFvRit:sFasL can selectively eliminate CD20+T-cells. After CD20-selective binding, scFvRit:sFasL is designed to trigger FasL-mediated activation-induced cell death of T-cells, but not B-cells. METHODS: Flow cytometry and immunohistochemistry were used to screen for CD20+inflammatory T-cells in MS blood and brain tissue. ScFvRit:sFasL pro-apoptotic activity was evaluated by Annexin-V/PI staining followed by flow cytometry assessment. RESULTS: Peripheral blood (n=11) and chronic but not active lesions of MS patient brains (n=5) contained CD20+inflammatory T-cells. Activated CD20+T-cells were predominantly CD4+and secreted both IL-17 and INF-γ. ScFvRit:sFasL triggered CD20-restricted FasL-mediated activation-induced cell death in peripheral blood CD20+T-cells, but not CD20+B-cells. CONCLUSION: CD20+inflammatory T-cells are present in blood and chronic brain lesions of MS patients. ScFvRit:sFasL selectively eliminated CD20+T-cells and may eliminate pathogenic T-cells without B-cell depletion.