Publications by authors named "Nathali Kaushansky"

31 Publications

Dasatinib response in acute myeloid leukemia is correlated with FLT3/ITD, PTPN11 mutations and a unique gene expression signature.

Haematologica 2020 12 1;105(12):2795-2804. Epub 2020 Dec 1.

Department of Immunology, Weizmann Institute of Science, Rehovot, Israel.

Novel targeted therapies demonstrate improved survival in specific subgroups (defined by genetic variants) of acute myeloid leukemia (AML) patients, validating the paradigm of molecularly targeted therapy. However, identifying correlations between AML molecular attributes and effective therapies is challenging. Recent advances in high-throughput in vitro drug sensitivity screening applied to primary AML blasts were used to uncover such correlations; however, these methods cannot predict the response of leukemic stem cells (LSCs). Our study aimed to predict in vitro response to targeted therapies, based on molecular markers, with subsequent validation in LSCs. We performed ex vivo sensitivity screening to 46 drugs on 29 primary AML samples at diagnosis or relapse. Using unsupervised hierarchical clustering analysis we identified group with sensitivity to several tyrosine kinase inhibitors (TKIs), including the multi-TKI, dasatinib, and searched for correlations between dasatinib response, exome sequencing and gene expression from our dataset and from the Beat AML dataset. Unsupervised hierarchical clustering analysis of gene expression resulted in clustering of dasatinib responders and non-responders. In vitro response to dasatinib could be predicted based on gene expression (AUC=0.78). Furthermore, mutations in FLT3/ITD and PTPN11 were enriched in the dasatinib sensitive samples as opposed to mutations in TP53 which were enriched in resistant samples. Based on these results, we selected FLT3/ITD AML samples and injected them to NSG-SGM3 mice. Our results demonstrate that in a subgroup of FLT3/ITD AML (4 out of 9) dasatinib significantly inhibits LSC engraftment. In summary we show that dasatinib has an anti-leukemic effect both on bulk blasts and, more importantly, LSCs from a subset of AML patients that can be identified based on mutational and expression profiles. Our data provide a rational basis for clinical trials of dasatinib in a molecularly selected subset of AML patients.
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http://dx.doi.org/10.3324/haematol.2019.240705DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7726833PMC
December 2020

The Median Eminence, A New Oligodendrogenic Niche in the Adult Mouse Brain.

Stem Cell Reports 2020 06 14;14(6):1076-1092. Epub 2020 May 14.

Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.

The subventricular zone (SVZ) of the lateral ventricles and the subgranular zone (SGZ) of the dentate gyrus in the hippocampus are known as neurogenic niches. We show that the median eminence (ME) of the hypothalamus comprises BrdU newly proliferating cells co-expressing NG2 (oligodendrocyte progenitors) and RIP (pre-myelinating oligodendrocytes), suggesting their differentiation toward mature oligodendrocytes (OLs). ME cells can generate neurospheres (NS) in vitro, which differentiate mostly to OLs compared with SVZ-NS that typically generate neurons. Interestingly, this population of oligodendrocyte progenitors is increased in the ME from experimental autoimmune encephalomyelitis (EAE)-affected mice. Notably, the thrombospondin 1 (TSP1) expressed by astrocytes, acts as negative regulator of oligodendrogenesis in vitro and is downregulated in the ME of EAE mice. Importantly, transplanted ME-NS preferentially differentiate to MBP OLs compared with SVZ-NS in Shiverer mice. Hence, discovering the ME as a new site for myelin-producing cells has a great importance for advising future therapy for demyelinating diseases and spinal cord injury.
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http://dx.doi.org/10.1016/j.stemcr.2020.04.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7355143PMC
June 2020

Antagonistic Center-Surround Mechanisms for Direction Selectivity in the Retina.

Cell Rep 2020 05;31(5):107608

Department of Neurobiology, Weizmann Institute of Science, Rehovot 76100, Israel. Electronic address:

An antagonistic center-surround receptive field is a key feature in sensory processing, but how it contributes to specific computations such as direction selectivity is often unknown. Retinal On-starburst amacrine cells (SACs), which mediate direction selectivity in direction-selective ganglion cells (DSGCs), exhibit antagonistic receptive field organization: depolarizing to light increments and decrements in their center and surround, respectively. We find that a repetitive stimulation exhausts SAC center and enhances its surround and use it to study how center-surround responses contribute to direction selectivity. Center, but not surround, activation induces direction-selective responses in SACs. Nevertheless, both SAC center and surround elicited direction-selective responses in DSGCs, but to opposite directions. Physiological and modeling data suggest that the opposing direction selectivity can result from inverted temporal balance between excitation and inhibition in DSGCs, implying that SAC's response timing dictates direction selectivity. Our findings reveal antagonistic center-surround mechanisms for direction selectivity and demonstrate how context-dependent receptive field reorganization enables flexible computations.
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http://dx.doi.org/10.1016/j.celrep.2020.107608DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7221349PMC
May 2020

Circulating Picomolar Levels of CCL2 Downregulate Ongoing Chronic Experimental Autoimmune Encephalomyelitis by Induction of Regulatory Mechanisms.

J Immunol 2019 10 4;203(7):1857-1866. Epub 2019 Sep 4.

Department of Immunology, The Weizmann Institute of Science, Rehovot 76100, Israel

Multiple sclerosis is an inflammatory disease of the CNS characterized by neurologic impairment resulting from primary demyelination and axonal damage. The pathogenic mechanisms of disease development include Ag-specific T cell activation and Th1 differentiation, followed by T cell and macrophage migration into the CNS. CCL2 is a chemokine that induces migration of monocytes, memory T cells, and dendritic cells. We previously demonstrated that picomolar levels of CCL2 strongly restrict the development of inflammation in models of inflammatory bowel disease. Moreover, CCR2 deficiency in T cells promotes a program inducing the accumulation of Foxp3 regulatory T cells while decreasing the levels of Th17 cells in vivo. In the current study, the effect of picomolar levels of CCL2 on the autoimmune inflammatory response associated with a multiple sclerosis-like disease in mice was analyzed. We found that low dosages of CCL2 were effective in suppressing MOG-induced experimental autoimmune encephalomyelitis (EAE), and they downregulated chronic EAE. The modulation of EAE by CCL2 was associated with downregulation of Th1/Th17 cells and upregulation of TGF-β and induction of regulatory CD4Foxp3 T cells. Most strikingly, these low levels of CCL2 induced formation of highly functional regulatory T cells. Thus, this study strongly supports the potential use of CCL2 as a regulatory mediator for treating inflammatory autoimmune diseases.
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http://dx.doi.org/10.4049/jimmunol.1900424DOI Listing
October 2019

Daily Onset of Light and Darkness Differentially Controls Hematopoietic Stem Cell Differentiation and Maintenance.

Cell Stem Cell 2018 10 30;23(4):572-585.e7. Epub 2018 Aug 30.

Immunology Department, Weizmann Institute of Science, Rehovot, Israel. Electronic address:

Hematopoietic stem and progenitor cells (HSPCs) tightly couple maintenance of the bone marrow (BM) reservoir, including undifferentiated long-term repopulating hematopoietic stem cells (LT-HSCs), with intensive daily production of mature leukocytes and blood replenishment. We found two daily peaks of BM HSPC activity that are initiated by onset of light and darkness providing this coupling. Both peaks follow transient elevation of BM norepinephrine and TNF secretion, which temporarily increase HSPC reactive oxygen species (ROS) levels. Light-induced norepinephrine and TNF secretion augments HSPC differentiation and increases vascular permeability to replenish the blood. In contrast, darkness-induced TNF increases melatonin secretion to drive renewal of HSPCs and LT-HSC potential through modulating surface CD150 and c-Kit expression, increasing COX-2/αSMA macrophages, diminishing vascular permeability, and reducing HSPC ROS levels. These findings reveal that light- and darkness-induced daily bursts of norepinephrine, TNF, and melatonin within the BM are essential for synchronized mature blood cell production and HSPC pool repopulation.
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http://dx.doi.org/10.1016/j.stem.2018.08.002DOI Listing
October 2018

Small Molecules Co-targeting CKIα and the Transcriptional Kinases CDK7/9 Control AML in Preclinical Models.

Cell 2018 09 23;175(1):171-185.e25. Epub 2018 Aug 23.

The Lautenberg Center for Immunology and Cancer Research, Institute of Medical Research Israel-Canada, Hebrew University-Hadassah Medical School, Jerusalem, Israel. Electronic address:

CKIα ablation induces p53 activation, and CKIα degradation underlies the therapeutic effect of lenalidomide in a pre-leukemia syndrome. Here we describe the development of CKIα inhibitors, which co-target the transcriptional kinases CDK7 and CDK9, thereby augmenting CKIα-induced p53 activation and its anti-leukemic activity. Oncogene-driving super-enhancers (SEs) are highly sensitive to CDK7/9 inhibition. We identified multiple newly gained SEs in primary mouse acute myeloid leukemia (AML) cells and demonstrate that the inhibitors abolish many SEs and preferentially suppress the transcription elongation of SE-driven oncogenes. We show that blocking CKIα together with CDK7 and/or CDK9 synergistically stabilize p53, deprive leukemia cells of survival and proliferation-maintaining SE-driven oncogenes, and induce apoptosis. Leukemia progenitors are selectively eliminated by the inhibitors, explaining their therapeutic efficacy with preserved hematopoiesis and leukemia cure potential; they eradicate leukemia in MLL-AF9 and Tet2;Flt3 AML mouse models and in several patient-derived AML xenograft models, supporting their potential efficacy in curing human leukemia.
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http://dx.doi.org/10.1016/j.cell.2018.07.045DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6701634PMC
September 2018

Inhomogeneous Encoding of the Visual Field in the Mouse Retina.

Curr Biol 2018 03 15;28(5):655-665.e3. Epub 2018 Feb 15.

Department of Neurobiology, Weizmann Institute of Science, 234 Herzl Street, Rehovot 7610001, Israel. Electronic address:

Stimulus characteristics of the mouse's visual field differ above and below the skyline. Here, we show for the first time that retinal ganglion cells (RGCs), the output neurons of the retina, gradually change their functional properties along the ventral-dorsal axis to allow better representation of the different stimulus characteristics. We conducted two-photon targeted recordings of transient-Offα-RGCs and found that they gradually became more sustained along the ventral-dorsal axis, revealing >5-fold-longer duration responses in the dorsal retina. Using voltage-clamp recordings, pharmacology, and genetic manipulation, we demonstrated that the primary rod pathway underlies this variance. Our findings challenge the current belief that RGCs of the same subtype exhibit the same light responses, regardless of retinal location, and suggest that networks underlying RGC responses may change with retinal location to enable optimized sampling of the visual image.
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http://dx.doi.org/10.1016/j.cub.2018.01.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6037286PMC
March 2018

CCR2 Regulates the Immune Response by Modulating the Interconversion and Function of Effector and Regulatory T Cells.

J Immunol 2017 06 15;198(12):4659-4671. Epub 2017 May 15.

Department of Immunology, Weizmann Institute of Science, Rehovot 76100, Israel; and

Chemokines and chemokine receptors establish a complex network modulating immune cell migration and localization. These molecules were also suggested to mediate the differentiation of leukocytes; however, their intrinsic, direct regulation of lymphocyte fate remained unclear. CCR2 is the main chemokine receptor inducing macrophage and monocyte recruitment to sites of inflammation, and it is also expressed on T cells. To assess whether CCR2 directly regulates T cell responses, we followed the fates of CCR2 T cells in T cell-specific inflammatory models. Our in vitro and in vivo results show that CCR2 intrinsically mediates the expression of inflammatory T cell cytokines, and its absence on T cells results in attenuated colitis progression. Moreover, CCR2 deficiency in T cells promoted a program inducing the accumulation of Foxp3 regulatory T cells, while decreasing the levels of Th17 cells in vivo, indicating that CCR2 regulates the immune response by modulating the effector/regulatory T ratio.
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http://dx.doi.org/10.4049/jimmunol.1601458DOI Listing
June 2017

Pathways and gene networks mediating the regulatory effects of cannabidiol, a nonpsychoactive cannabinoid, in autoimmune T cells.

J Neuroinflammation 2016 06 3;13(1):136. Epub 2016 Jun 3.

The Dr Miriam and Sheldon G. Adelson Center for the Biology of Addictive Diseases, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, 6997801, Israel.

Background: Our previous studies showed that the non-psychoactive cannabinoid, cannabidiol (CBD), ameliorates the clinical symptoms in mouse myelin oligodendrocyte glycoprotein (MOG)35-55-induced experimental autoimmune encephalomyelitis model of multiple sclerosis (MS) as well as decreases the memory MOG35-55-specific T cell (TMOG) proliferation and cytokine secretion including IL-17, a key autoimmune factor. The mechanisms of these activities are currently poorly understood.

Methods: Herein, using microarray-based gene expression profiling, we describe gene networks and intracellular pathways involved in CBD-induced suppression of these activated memory TMOG cells. Encephalitogenic TMOG cells were stimulated with MOG35-55 in the presence of spleen-derived antigen presenting cells (APC) with or without CBD. mRNA of purified TMOG was then subjected to Illumina microarray analysis followed by ingenuity pathway analysis (IPA), weighted gene co-expression network analysis (WGCNA) and gene ontology (GO) elucidation of gene interactions. Results were validated using qPCR and ELISA assays.

Results: Gene profiling showed that the CBD treatment suppresses the transcription of a large number of proinflammatory genes in activated TMOG. These include cytokines (Xcl1, Il3, Il12a, Il1b), cytokine receptors (Cxcr1, Ifngr1), transcription factors (Ier3, Atf3, Nr4a3, Crem), and TNF superfamily signaling molecules (Tnfsf11, Tnfsf14, Tnfrsf9, Tnfrsf18). "IL-17 differentiation" and "IL-6 and IL-10-signaling" were identified among the top processes affected by CBD. CBD increases a number of IFN-dependent transcripts (Rgs16, Mx2, Rsad2, Irf4, Ifit2, Ephx1, Ets2) known to execute anti-proliferative activities in T cells. Interestingly, certain MOG35-55 up-regulated transcripts were maintained at high levels in the presence of CBD, including transcription factors (Egr2, Egr1, Tbx21), cytokines (Csf2, Tnf, Ifng), and chemokines (Ccl3, Ccl4, Cxcl10) suggesting that CBD may promote exhaustion of memory TMOG cells. In addition, CBD enhanced the transcription of T cell co-inhibitory molecules (Btla, Lag3, Trat1, and CD69) known to interfere with T/APC interactions. Furthermore, CBD enhanced the transcription of oxidative stress modulators with potent anti-inflammatory activity that are controlled by Nfe2l2/Nrf2 (Mt1, Mt2a, Slc30a1, Hmox1).

Conclusions: Microarray-based gene expression profiling demonstrated that CBD exerts its immunoregulatory effects in activated memory TMOG cells via (a) suppressing proinflammatory Th17-related transcription, (b) by promoting T cell exhaustion/tolerance, (c) enhancing IFN-dependent anti-proliferative program, (d) hampering antigen presentation, and (d) inducing antioxidant milieu resolving inflammation. These findings put forward mechanism by which CBD exerts its anti-inflammatory effects as well as explain the beneficial role of CBD in pathological memory T cells and in autoimmune diseases.
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http://dx.doi.org/10.1186/s12974-016-0603-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4891926PMC
June 2016

Post-CNS-inflammation expression of CXCL12 promotes the endogenous myelin/neuronal repair capacity following spontaneous recovery from multiple sclerosis-like disease.

J Neuroinflammation 2016 Jan 8;13. Epub 2016 Jan 8.

Department of Immunology, The Weizmann Institute of Science, 234 Herzl Street, Rehovot, 7610001, Israel.

Background: Demyelination and axonal degeneration, hallmarks of multiple sclerosis (MS), are associated with the central nervous system (CNS) inflammation facilitated by C-X-C motif chemokine 12 (CXCL12) chemokine. Both in MS and in experimental autoimmune encephalomyelitis (EAE), the deleterious CNS inflammation has been associated with upregulation of CXCL12 expression in the CNS. We investigated the expression dynamics of CXCL12 in the CNS with progression of clinical EAE and following spontaneous recovery, with a focus on CXCL12 expression in the hippocampal neurogenic dentate gyrus (DG) and in the corpus callosum (CC) of spontaneously recovered mice, and its potential role in promoting the endogenous myelin/neuronal repair capacity.

Methods: CNS tissue sections from mice with different clinical EAE phases or following spontaneous recovery and in vitro differentiated adult neural stem cell cultures were analyzed by immunofluorescent staining and confocal imaging for detecting and enumerating neuronal progenitor cells (NPCs) and oligodendrocyte precursor cells (OPCs) and for expression of CXCL12.

Results: Our expression dynamics analysis of CXCL12 in the CNS with EAE progression revealed elevated CXCL12 expression in the DG and CC, which persistently increases following spontaneous recovery even though CNS inflammation has subsided. Correspondingly, the numbers of NPCs and OPCs in the DG and CC, respectively, of EAE-recovered mice increased compared to that of naïve mice (NPCs, p < 0.0001; OPCs, p < 0.00001) or mice with active disease (OPCs, p < 0.0005). Notably, about 30 % of the NPCs and unexpectedly also OPCs (~50 %) express CXCL12, and their numbers in DG and CC, respectively, are higher in EAE-recovered mice compared with naïve mice and also compared with mice with ongoing clinical EAE (CXCL12(+) NPCs, p < 0.005; CXCL12(+) OPCs, p < 0.0005). Moreover, a significant proportion (>20 %) of the CXCL12(+) NPCs and OPCs co-express the CXCL12 receptor, CXCR4, and their numbers significantly increase with recovery from EAE not only relative to naïve mice (p < 0.0002) but also to mice with ongoing EAE (p < 0.004).

Conclusions: These data link CXCL12 expression in the DG and CC of EAE-recovering mice to the promotion of neuro/oligodendrogenesis generating CXCR4(+) CXCL12(+) neuronal and oligodendrocyte progenitor cells endowed with intrinsic neuro/oligondendroglial differentiation potential. These findings highlight the post-CNS-inflammation role of CXCL12 in augmenting the endogenous myelin/neuronal repair capacity in MS-like disease, likely via CXCL12/CXCR4 autocrine signaling.
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http://dx.doi.org/10.1186/s12974-015-0468-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4706716PMC
January 2016

Anti-inflammatory effects of the cannabidiol derivative dimethylheptyl-cannabidiol - studies in BV-2 microglia and encephalitogenic T cells.

J Basic Clin Physiol Pharmacol 2016 May;27(3):289-96

Background: Dimethylheptyl-cannabidiol (DMH-CBD), a non-psychoactive, synthetic derivative of the phytocannabinoid cannabidiol (CBD), has been reported to be anti-inflammatory in RAW macrophages. Here, we evaluated the effects of DMH-CBD at the transcriptional level in BV-2 microglial cells as well as on the proliferation of encephalitogenic T cells.

Methods: BV-2 cells were pretreated with DMH-CBD, followed by stimulation with the endotoxin lipopolysaccharide (LPS). The expression levels of selected genes involved in stress regulation and inflammation were determined by quantitative real-time PCR. In addition, MOG35-55-reactive T cells (TMOG) were cultured with antigen-presenting cells in the presence of DMH-CBD and MOG35-55 peptide, and cell proliferation was determined by measuring [3H]thymidine incorporation.

Results: DMH-CBD treatment downregulated in a dose-dependent manner the mRNA expression of LPS-upregulated pro-inflammatory genes (Il1b, Il6, and Tnf) in BV-2 microglial cells. The expression of these genes was also downregulated by DMH-CBD in unstimulated cells. In parallel, DMH-CBD upregulated the expression of genes related to oxidative stress and glutathione homeostasis such as Trb3, Slc7a11/xCT, Hmox1, Atf4, Chop, and p8 in both stimulated and unstimulated microglial cells. In addition, DMH-CBD dose-dependently inhibited MOG35-55-induced TMOG proliferation.

Conclusions: The results show that DMH-CBD has similar anti-inflammatory properties to those of CBD. DMH-CBD downregulates the expression of inflammatory cytokines and protects the microglial cells by inducing an adaptive cellular response against inflammatory stimuli and oxidative injury. In addition, DMH-CBD decreases the proliferation of pathogenic activated TMOG cells.
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http://dx.doi.org/10.1515/jbcpp-2015-0071DOI Listing
May 2016

CD151 Regulates T-Cell Migration in Health and Inflammatory Bowel Disease.

Inflamm Bowel Dis 2016 Feb;22(2):257-67

*Department of Immunology, Weizmann Institute of Science, Rehovot, Israel; ‡IBD Unit, Department of Gastroenterology and Liver Diseases, Tel Aviv Medical Center, Tel Aviv, Israel (affiliated with the Sackler Faculty of Medicine, Tel Aviv, Israel); and §Division of Cell Biology, The Netherlands Cancer Institute, Amsterdam, the Netherlands.

The continuous recirculation of mature lymphocytes and their entry into the peripheral lymph nodes are crucial for the development of an immune response to foreign antigens. Occasionally, the entry and the subsequent response of T lymphocytes in these sites lead to severe inflammation and pathological conditions. Here, we characterized the tetraspanin molecule, CD151, as a regulator of T cell motility in health and in models of inflammatory bowel disease. CD151 formed a cell surface complex with VLA-4 and LFA-1 integrins, and its activation led to enhanced migration of T cells. Picomolar levels of CCL2 that were previously shown to inhibit T-cell migration to lymph nodes suppressed CD151 expression and dissociated CD151-integrin complexes in T lymphocytes, resulting in attenuated migration toward T-cell attractant chemokines. To directly inhibit CD151 function, a truncated CD151 peptide fragment mimicking of the CD151 extracellular loop was designed. CD151 extracellular loop inhibited T-cell migration in vitro and in vivo and attenuated the development of dextrane sulfate sodium-induced colitis. Thus, CD151 is a key orchestrator of T cell motility; interference with its proper function results in attenuated progression of inflammatory bowel disease.
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http://dx.doi.org/10.1097/MIB.0000000000000621DOI Listing
February 2016

HU-446 and HU-465, Derivatives of the Non-psychoactive Cannabinoid Cannabidiol, Decrease the Activation of Encephalitogenic T Cells.

Chem Biol Drug Des 2016 Jan 16;87(1):143-53. Epub 2015 Sep 16.

The Dr Miriam and Sheldon G. Adelson Center for the Biology of Addictive Diseases, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, 69978, Israel.

Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease peripheral inflammation and neuroinflammation in mouse experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Here we have studied the anti-inflammatory effects of newly synthesized derivatives of natural (-)-CBD ((-)-8,9-dihydro-7-hydroxy-CBD; HU-446) and of synthetic (+)-CBD ((+)-8,9-dihydro-7-hydroxy-CBD; HU-465) on activated myelin oligodendrocyte glycoprotein (MOG)35-55-specific mouse encephalitogenic T cells (T(MOG) ) driving EAE/MS-like pathologies. Binding assays followed by molecular modeling revealed that HU-446 has negligible affinity toward the cannabinoid CB1 and CB2 receptors while HU-465 binds to both CB1 and CB2 receptors at the high nanomolar concentrations (Ki = 76.7 ± 5.8 nm and 12.1 ± 2.3 nm, respectively). Both, HU-446 and HU-465, at 5 and 10 μm (but not at 0.1 and 1 μm), inhibited the MOG35-55-induced proliferation of autoreactive T(MOG) cells via CB1/CB2 receptor independent mechanisms. Moreover, both HU-446 and HU-465, at 5 and 10 μm, inhibited the release of IL-17, a key autoimmune cytokine, from MOG35-55-stimulated T(MOG) cells. These results suggest that HU-446 and HU-465 have anti-inflammatory potential in inflammatory and autoimmune diseases.
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http://dx.doi.org/10.1111/cbdd.12637DOI Listing
January 2016

Role of a Novel Human Leukocyte Antigen-DQA1*01:02;DRB1*15:01 Mixed Isotype Heterodimer in the Pathogenesis of "Humanized" Multiple Sclerosis-like Disease.

J Biol Chem 2015 Jun 24;290(24):15260-78. Epub 2015 Apr 24.

From the Departments of Immunology and

Gene-wide association and candidate gene studies indicate that the greatest effect on multiple sclerosis (MS) risk is driven by the HLA-DRB1*15:01 allele within the HLA-DR15 haplotype (HLA-DRB1*15:01-DQA1*01:02-DQB1*0602-DRB5*01:01). Nevertheless, linkage disequilibrium makes it difficult to define, without functional studies, whether the functionally relevant effect derives from DRB1*15:01 only, from its neighboring DQA1*01:02-DQB1*06:02 or DRB5*01:01 genes of HLA-DR15 haplotype, or from their combinations or epistatic interactions. Here, we analyzed the impact of the different HLA-DR15 haplotype alleles on disease susceptibility in a new "humanized" model of MS induced in HLA-transgenic (Tg) mice by human oligodendrocyte-specific protein (OSP)/claudin-11 (hOSP), one of the bona fide potential primary target antigens in MS. We show that the hOSP-associated MS-like disease is dominated by the DRB1*15:01 allele not only as the DRA1*01:01;DRB1*15:01 isotypic heterodimer but also, unexpectedly, as a functional DQA1*01:02;DRB1*15:01 mixed isotype heterodimer. The contribution of HLA-DQA1/DRB1 mixed isotype heterodimer to OSP pathogenesis was revealed in (DRB1*1501xDQB1*0602)F1 double-Tg mice immunized with hOSP(142-161) peptide, where the encephalitogenic potential of prevalent DRB1*1501/hOSP(142-161)-reactive Th1/Th17 cells is hindered due to a single amino acid difference in the OSP(142-161) region between humans and mice; this impedes binding of DRB1*1501 to the mouse OSP(142-161) epitope in the mouse CNS while exposing functional binding of mouse OSP(142-161) to DQA1*01:02;DRB1*15:01 mixed isotype heterodimer. This study, which shows for the first time a functional HLA-DQA1/DRB1 mixed isotype heterodimer and its potential association with disease susceptibility, provides a rationale for a potential effect on MS risk from DQA1*01:02 through functional DQA1*01:02;DRB1*15:01 antigen presentation. Furthermore, it highlights a potential contribution to MS risk also from interisotypic combination between products of neighboring HLA-DR15 haplotype alleles, in this case the DQA1/DRB1 combination.
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http://dx.doi.org/10.1074/jbc.M115.641209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4463466PMC
June 2015

Cannabidiol, a non-psychoactive cannabinoid, leads to EGR2-dependent anergy in activated encephalitogenic T cells.

J Neuroinflammation 2015 Mar 15;12:52. Epub 2015 Mar 15.

The Dr Miriam and Sheldon G. Adelson Center for the Biology of Addictive Diseases, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel.

Background: Cannabidiol (CBD), the main non-psychoactive cannabinoid, has been previously shown by us to ameliorate clinical symptoms and to decrease inflammation in myelin oligodendrocyte glycoprotein (MOG)35-55-induced mouse experimental autoimmune encephalomyelitis model of multiple sclerosis as well as to decrease MOG35-55-induced T cell proliferation and IL-17 secretion. However, the mechanisms of CBD anti-inflammatory activities are unclear.

Methods: Here we analyzed the effects of CBD on splenocytes (source of accessory T cells and antigen presenting cells (APC)) co-cultured with MOG35-55-specific T cells (TMOG) and stimulated with MOG35-55. Using flow cytometry, we evaluated the expression of surface activation markers and inhibitory molecules on T cells and B cells. TMOG cells were purified using CD4 positive microbead selection and submitted for quantitative PCR and microarray of mRNA transcript analyzes. Cell signaling studies in purified TMOG were carried out using immunoblotting.

Results: We found that CBD leads to upregulation of CD69 and lymphocyte-activation gene 3 (LAG3) regulatory molecules on CD4(+)CD25(-) accessory T cells. This subtype of CD4(+)CD25(-)CD69(+)LAG3(+) T cells has been recognized as induced regulatory phenotype promoting anergy in activated T cells. Indeed, we observed that CBD treatment results in upregulation of EGR2 (a key T cell anergy inducer) mRNA transcription in stimulated TMOG cells. This was accompanied by elevated levels of anergy promoting genes such as IL-10 (anti-inflammatory cytokine), STAT5 (regulatory factor), and LAG3 mRNAs, as well as of several enhancers of cell cycle arrest (such as Nfatc1, Casp4, Cdkn1a, and Icos). Moreover, CBD exposure leads to a decrease in STAT3 and to an increase in STAT5 phosphorylation in TMOG cells, positive and negative regulators of Th17 activity, respectively. In parallel, we observed decreased levels of major histocompatibility complex class II (MHCII), CD25, and CD69 on CD19(+) B cells following CBD treatment, showing diminished antigen presenting capabilities of B cells and reduction in their pro-inflammatory functions.

Conclusions: Our data suggests that CBD exerts its immunoregulatory effects via induction of CD4(+)CD25(-)CD69(+)LAG3(+) cells in MOG35-55-activated APC/TMOG co-cultures. This is accompanied by EGR2-dependent anergy of stimulated TMOG cells as well as a switch in their intracellular STAT3/STAT5 activation balance leading to the previously observed decrease in Th17 activity.
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http://dx.doi.org/10.1186/s12974-015-0273-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4363052PMC
March 2015

DQB1*06:02-Associated Pathogenic Anti-Myelin Autoimmunity in Multiple Sclerosis-Like Disease: Potential Function of DQB1*06:02 as a Disease-Predisposing Allele.

Front Oncol 2014 16;4:280. Epub 2014 Oct 16.

Department of Immunology, The Weizmann Institute of Science , Rehovot , Israel.

Susceptibility to multiple sclerosis (MS) has been linked mainly to the HLA-DRB1 locus, with the HLA-DR15 haplotype (DRB1*1501-DQA1*0102-DQB1*0602-DRB5*0101) dominating MS risk in Caucasians. Although genes in the HLA-II region, particularly DRB1*1501, DQA1*0102-DQB1*0602, are in tight linkage disequilibrium, genome-wide-association, and gene candidate studies identified the DRB1*15:01 allele as the primary risk factor in MS. Many genetic and immune-functional studies have indicated DRB1*15:01 as a primary risk factor in MS, while only some functional studies suggested a disease-modifying role for the DRB5*01 or DQB1*06 alleles. In this respect, the susceptibility of DRB1*15:01-transgenic (Tg) mice to myelin basic protein- or myelin oligodendrocyte glycoprotein-induced MS-like disease is consistent with primary contribution of DRB1*15:01 to HLA-DR15+ MS. The studies summarized here show that susceptibility to MS-like disease, induced in HLA-"humanized" mice by myelin oligodendrocytic basic protein or by the proteolipid protein, one of the most prominent encephalitogenic target antigens implicated in human MS, is determined by DQB1*06:02, rather than by the DRB1*15:01 allele. These findings not only offer a rationale for a potential role for DQB1*06:02 in predisposing susceptibility to MS, but also suggest a more complex and differential functional role for HLA-DR15 alleles, depending on the primary target myelin antigen. However, the conflict between these findings in HLA-Tg mice and the extensive genome-wide-association studies, which could not detect any significant effect from the DQB1*06:02 allele on MS risk, is rather puzzling. Functional analysis of MS PBLs for DQB1*06:02-associated anti-myelin autoimmunity may indicate whether or not DQB1*06:02 is associated with MS pathogenesis.
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http://dx.doi.org/10.3389/fonc.2014.00280DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4199271PMC
October 2014

From classic to spontaneous and humanized models of multiple sclerosis: impact on understanding pathogenesis and drug development.

J Autoimmun 2014 Nov 28;54:33-50. Epub 2014 Aug 28.

Department of Neuroimmunology, Max Planck Institute of Neurobiology, Martinsried 82152, Germany. Electronic address:

Multiple sclerosis (MS), a demyelinating disease of the central nervous system (CNS), presents as a complex disease with variable clinical and pathological manifestations, involving different pathogenic pathways. Animal models, particularly experimental autoimmune encephalomyelitis (EAE), have been key to deciphering the pathophysiology of MS, although no single model can recapitulate the complexity and diversity of MS, or can, to date, integrate the diverse pathogenic pathways. Since the first EAE model was introduced decades ago, multiple classic (induced), spontaneous, and humanized EAE models have been developed, each recapitulating particular aspects of MS pathogenesis. The advances in technologies of genetic ablation and transgenesis in mice of C57BL/6J background and the development of myelin-oligodendrocyte glycoprotein (MOG)-induced EAE in C57BL/6J mice yielded several spontaneous and humanized EAE models, and resulted in a plethora of EAE models in which the role of specific genes or cell populations could be precisely interrogated, towards modeling specific pathways of MS pathogenesis/regulation in MS. Collectively, the numerous studies on the different EAE models contributed immensely to our basic understanding of cellular and molecular pathways in MS pathogenesis as well as to the development of therapeutic agents: several drugs available today as disease modifying treatments were developed from direct studies on EAE models, and many others were tested or validated in EAE. In this review, we discuss the contribution of major classic, spontaneous, and humanized EAE models to our understanding of MS pathophysiology and to insights leading to devising current and future therapies for this disease.
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http://dx.doi.org/10.1016/j.jaut.2014.06.004DOI Listing
November 2014

An immunomodulating motif of the HIV-1 fusion protein is chirality-independent: implications for its mode of action.

J Biol Chem 2013 Nov 27;288(46):32852-60. Epub 2013 Sep 27.

From the Departments of Biological Chemistry and.

An immunosuppressive motif was recently found within the HIV-1 gp41 fusion protein (termed immunosuppressive loop-associated determinant core motif (ISLAD CM)). Peptides containing the motif interact with the T-cell receptor (TCR) complex; however, the mechanism by which the motif exerts its immunosuppressive activity is yet to be determined. Recent studies showed that interactions between protein domains in the membrane milieu are not always sterically controlled. Therefore, we utilized the unique membrane leniency toward association between D- and L-stereoisomers to investigate the detailed mechanism by which ISLAD CM inhibits T-cell activation. We show that a D-enantiomer of ISLAD CM (termed ISLAD D-CM) inhibited the proliferation of murine myelin oligodendrocyte glycoprotein (MOG)-(35-55)-specific line T-cells to the same extent as the l-motif form. Moreover, the D- and L-forms preferentially bound spleen-derived T-cells over B-cells by 13-fold. Furthermore, both forms of ISLAD CM co-localized with the TCR on activated T-cells and interacted with the transmembrane domain of the TCR. FRET experiments revealed the importance of basic residues for the interaction between ISLAD CM forms and the TCR transmembrane domain. Ex vivo studies demonstrated that ISLAD D-CM administration inhibited the proliferation (72%) and proinflammatory cytokine secretion of pathogenic MOG(35-55)-specific T-cells. This study provides insights into the immunosuppressive mechanism of gp41 and demonstrates that chirality-independent interactions in the membrane can take place in diverse biological systems. Apart from HIV pathogenesis, the D-peptide reported herein may serve as a potential tool for treating T-cell-mediated pathologies.
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http://dx.doi.org/10.1074/jbc.M113.512038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3829137PMC
November 2013

Cannabinoids decrease the th17 inflammatory autoimmune phenotype.

J Neuroimmune Pharmacol 2013 Dec 28;8(5):1265-76. Epub 2013 Jul 28.

The Dr Miriam and Sheldon G. Adelson Center for the Biology of Addictive Diseases, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel,

Cannabinoids, the Cannabis constituents, are known to possess anti-inflammatory properties but the mechanisms involved are not understood. Here we show that the main psychoactive cannabinoid, Δ-9-tetrahydrocannabinol (THC), and the main nonpsychoactive cannabinoid, cannabidiol (CBD), markedly reduce the Th17 phenotype which is known to be increased in inflammatory autoimmune pathologies such as Multiple Sclerosis. We found that reactivation by MOG35-55 of MOG35-55-specific encephalitogenic T cells (cells that induce Experimental Autoimmune Encephalitis when injected to mice) in the presence of spleen derived antigen presenting cells led to a large increase in IL-17 production and secretion. In addition, we found that the cannabinoids CBD and THC dose-dependently (at 0.1-5 μM) suppressed the production and secretion of this cytokine. Moreover, the mRNA and protein of IL-6, a key factor in Th17 induction, were also decreased. Pretreatment with CBD also resulted in increased levels of the anti-inflammatory cytokine IL-10. Interestingly, CBD and THC did not affect the levels of TNFα and IFNγ. The downregulation of IL-17 secretion by these cannabinoids does not seem to involve the CB1, CB2, PPARγ, 5-HT1A or TRPV1 receptors. In conclusion, the results show a unique cannabinoid modulation of the autoimmune cytokine milieu combining suppression of the pathogenic IL-17 and IL-6 cytokines along with boosting the expression of the anti-inflammatory cytokine IL-10.
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http://dx.doi.org/10.1007/s11481-013-9493-1DOI Listing
December 2013

A highly conserved sequence associated with the HIV gp41 loop region is an immunomodulator of antigen-specific T cells in mice.

Blood 2013 Mar 16;121(12):2244-52. Epub 2013 Jan 16.

Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.

Modulation of T-cell responses by HIV occurs via distinct mechanisms, 1 of which involves inactivation of T cells already at the stage of virus-cell fusion. Hydrophobic portions of the gp41 protein of the viral envelope that contributes to membrane fusion may modulate T-cell responsiveness. Here we found a highly conserved sequence (termed "ISLAD") that is associated with the membranotropic gp41 loop region. We showed that ISLAD has the ability to bind the T-cell membrane and to interact with the T-cell receptor (TCR) complex. Furthermore, ISLAD inhibited T-cell proliferation and interferon-γ secretion that resulted from TCR engagement through antigen-presenting cells. Moreover, administering ISLAD (10 µg per mouse) to an experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis reduced the severity of the disease. This was related to the inhibition of pathogenic T-cell proliferation and to reduced pro-inflammatory cytokine secretion in the lymph nodes of ISLAD-treated EAE mice. The data suggest that T-cell inactivation by HIV during membrane fusion may lie in part in this conserved sequence associated with the gp41 loop region.
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http://dx.doi.org/10.1182/blood-2012-11-468900DOI Listing
March 2013

DQB1*0602 rather than DRB1*1501 confers susceptibility to multiple sclerosis-like disease induced by proteolipid protein (PLP).

J Neuroinflammation 2012 Feb 8;9:29. Epub 2012 Feb 8.

Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.

Background: Multiple sclerosis (MS) is associated with pathogenic autoimmunity primarily focused on major CNS-myelin target antigens including myelin basic protein (MBP), proteolipidprotein (PLP), myelin oligodendrocyte protein (MOG). MS is a complex trait whereby the HLA genes, particularly class-II genes of HLA-DR15 haplotype, dominate the genetic contribution to disease-risk. Due to strong linkage disequilibrium in HLA-II region, it has been hard to establish precisely whether the functionally relevant effect derives from the DRB1*1501, DQA1*0102-DQB1*0602, or DRB5*0101 loci of HLA-DR15 haplotype, their combinations, or their epistatic interactions. Nevertheless, most genetic studies have indicated DRB1*1501 as a primary risk factor in MS. Here, we used 'HLA-humanized' mice to discern the potential relative contribution of DRB1*1501 and DQB1*0602 alleles to susceptibility to "humanized" MS-like disease induced by PLP, one of the most prominent and encephalitogenic target-antigens implicated in human MS.

Methods: The HLA-DRB1*1501- and HLA-DQB1*0602-Tg mice (MHC-II(-/-)), and control non-HLA-DR15-relevant-Tg mice were immunized with a set of overlapping PLP peptides or with recombinant soluble PLP for induction of "humanized" MS-like disease, as well as for ex-vivo analysis of immunogenic/immunodominant HLA-restricted T-cell epitopes and associated cytokine secretion profile.

Results: PLP autoimmunity in both HLA-DR15-Tg mice was focused on 139-151 and 175-194 epitopes. Strikingly, however, the HLA-DRB1*1501-transgenics were refractory to disease induction by any of the overlapping PLP peptides, while HLA-DQB1*0602 transgenics were susceptible to disease induction by PLP139-151 and PLP175-194 peptides. Although both transgenics responded to both peptides, the PLP139-151- and PLP175-194-reactive T-cells were directed to Th1/Th17 phenotype in DQB1*0602-Tg mice and towards Th2 in DRB1*1501-Tg mice.

Conclusions: While genome studies map a strong MS susceptibility effect to the region of DRB1*1501, our findings offer a rationale for potential involvement of pathogenic DQ6-associated autoimmunity in MS. Moreover, that DQB1*0602, but not DRB1*1501, determines disease-susceptibility to PLP in HLA-transgenics, suggests a potential differential, functional role for DQB1*0602 as a predisposing allele in MS. This, together with previously demonstrated disease-susceptibility to MBP and MOG in DRB1*1501-transgenics, also suggests a differential role for DRB1*1501 and DQB1*0602 depending on target antigen and imply a potential complex 'genotype/target antigen/phenotype' relationship in MS heterogeneity.
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http://dx.doi.org/10.1186/1742-2094-9-29DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3344688PMC
February 2012

'Multi-epitope-targeted' immune-specific therapy for a multiple sclerosis-like disease via engineered multi-epitope protein is superior to peptides.

PLoS One 2011 29;6(11):e27860. Epub 2011 Nov 29.

Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.

Antigen-induced peripheral tolerance is potentially one of the most efficient and specific therapeutic approaches for autoimmune diseases. Although highly effective in animal models, antigen-based strategies have not yet been translated into practicable human therapy, and several clinical trials using a single antigen or peptidic-epitope in multiple sclerosis (MS) yielded disappointing results. In these clinical trials, however, the apparent complexity and dynamics of the pathogenic autoimmunity associated with MS, which result from the multiplicity of potential target antigens and "epitope spread", have not been sufficiently considered. Thus, targeting pathogenic T-cells reactive against a single antigen/epitope is unlikely to be sufficient; to be effective, immunospecific therapy to MS should logically neutralize concomitantly T-cells reactive against as many major target antigens/epitopes as possible. We investigated such "multi-epitope-targeting" approach in murine experimental autoimmune encephalomyelitis (EAE) associated with a single ("classical") or multiple ("complex") anti-myelin autoreactivities, using cocktail of different encephalitogenic peptides vis-a-vis artificial multi-epitope-protein (designated Y-MSPc) encompassing rationally selected MS-relevant epitopes of five major myelin antigens, as "multi-epitope-targeting" agents. Y-MSPc was superior to peptide(s) in concomitantly downregulating pathogenic T-cells reactive against multiple myelin antigens/epitopes, via inducing more effective, longer lasting peripheral regulatory mechanisms (cytokine shift, anergy, and Foxp3+ CTLA4+ regulatory T-cells). Y-MSPc was also consistently more effective than the disease-inducing single peptide or peptide cocktail, not only in suppressing the development of "classical" or "complex EAE" or ameliorating ongoing disease, but most importantly, in reversing chronic EAE. Overall, our data emphasize that a "multi-epitope-targeting" strategy is required for effective immune-specific therapy of organ-specific autoimmune diseases associated with complex and dynamic pathogenic autoimmunity, such as MS; our data further demonstrate that the "multi-epitope-targeting" approach to therapy is optimized through specifically designed multi-epitope-proteins, rather than myelin peptide cocktails, as "multi-epitope-targeting" agents. Such artificial multi-epitope proteins can be tailored to other organ-specific autoimmune diseases.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0027860PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3226621PMC
April 2012

Cannabidiol inhibits pathogenic T cells, decreases spinal microglial activation and ameliorates multiple sclerosis-like disease in C57BL/6 mice.

Br J Pharmacol 2011 Aug;163(7):1507-19

The Dr. Miriam and Sheldon G. Adelson Center for Biology of Addictive Diseases, Physiology and Pharmacology Department, Sackler School of Medicine, Tel Aviv University, Tel Aviv, Israel.

Background And Purpose: Cannabis extracts and several cannabinoids have been shown to exert broad anti-inflammatory activities in experimental models of inflammatory CNS degenerative diseases. Clinical use of many cannabinoids is limited by their psychotropic effects. However, phytocannabinoids like cannabidiol (CBD), devoid of psychoactive activity, are, potentially, safe and effective alternatives for alleviating neuroinflammation and neurodegeneration.

Experimental Approach: We used experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein (MOG) in C57BL/6 mice, as a model of multiple sclerosis. Using immunocytochemistry and cell proliferation assays we evaluated the effects of CBD on microglial activation in MOG-immunized animals and on MOG-specific T-cell proliferation.

Key Results: Treatment with CBD during disease onset ameliorated the severity of the clinical signs of EAE. This effect of CBD was accompanied by diminished axonal damage and inflammation as well as microglial activation and T-cell recruitment in the spinal cord of MOG-injected mice. Moreover, CBD inhibited MOG-induced T-cell proliferation in vitro at both low and high concentrations of the myelin antigen. This effect was not mediated via the known cannabinoid CB(1) and CB(2) receptors.

Conclusions And Implications: CBD, a non-psychoactive cannabinoid, ameliorates clinical signs of EAE in mice, immunized against MOG. Suppression of microglial activity and T-cell proliferation by CBD appeared to contribute to these beneficial effects.
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http://dx.doi.org/10.1111/j.1476-5381.2011.01379.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3165959PMC
August 2011

The myelin-associated oligodendrocytic basic protein (MOBP) as a relevant primary target autoantigen in multiple sclerosis.

Autoimmun Rev 2010 Feb 13;9(4):233-6. Epub 2009 Aug 13.

Department of Immunology, The Weizmann Institute of Science, P.O. Box 26, Rehovot 76000, Israel.

Multiple sclerosis (MS) is a disease of the human CNS, characterized by perivascular inflammation, demyelination and axonal damage. Although the etiology of MS is unknown, it is believed that the disease results from destructive autoimmune mechanisms, presumably initiated by abnormal activation of potentially pathogenic autoimmune T-cells recognizing CNS components. The myelin-associated oligodendrocyte basic protein (MOBP), a relatively abundant CNS-specific myelin protein, which plays a role in stabilizing the myelin sheath in the CNS, has recently been implicated in the pathogenesis of MS. Here we review studies showing that MOBP is as an important candidate target antigen in MS as the other widely studied target antigens, myelin basic protein (MBP), proteolipid protein (PLP), and myelin oligodendrocyte glycoprotein (MOG). The studies summarized below indicate that T-cell autoimmunity against MOBP can be detected in MS patients; T-cells reactive against MOBP can be pathogenic in several mouse strains as well as in the "humanized" HLA-DR15-Tg mice; and, that the HLA-DQ6-restricted, but not HLA-DR15-restricted, MOBP-reactive T-cells cause in HLA-DR15-Tg mice MS-like clinical disease associated with perivascular and parenchymal infiltration, demyelination, axonal loss, and optic neuritis. Accordingly, the MOBP should be considered a bona fide primary target antigen in MS, in addition to MBP, PLP, and MOG.
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http://dx.doi.org/10.1016/j.autrev.2009.08.002DOI Listing
February 2010

HLA-DQB1*0602 determines disease susceptibility in a new "humanized" multiple sclerosis model in HLA-DR15 (DRB1*1501;DQB1*0602) transgenic mice.

J Immunol 2009 Sep 31;183(5):3531-41. Epub 2009 Jul 31.

Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.

The susceptibility to multiple sclerosis (MS), a chronic neurological autoimmune disease that primarily targets CNS myelin, has long been associated with HLA class-II genes. Although several other HLA and non-HLA disease predisposing alleles have been identified, alleles of the HLA-DR15 haplotype (DRB1*1501, DRB5*0101, and DQB1*0602) remain the strongest susceptibility factor. Many studies have suggested that the HLA-DRB1*1501 allele determines MS-associated susceptibility. However, due to strong linkage disequilibrium within the HLA class II region, it has been difficult to unequivocally determine the relative roles of the DRB1*1501 and DQB1*0602 products. In this study we use HLA class-II transgenic mice to illuminate the relative contributions of the DRB1*1501 and DQB1*0602 alleles or their combination to susceptibility toward a new "humanized" MS-like disease induced by myelin-associated oligodendrocytic basic protein (MOBP). Although many immunological studies have focused overwhelmingly on the role of the HLA-DRB1*1501 product in MS, we show that HLA-DRB1*1501 transgenics are refractory to MOBP disease induction, whereas the HLA-DQB1*0602 transgenics are susceptible via T cells reactive against MOBP15-36 and MOBP55-77 encephalitogenic epitopes. Although both transgenics react against these epitopes, the MOBP15-36- and MOBP55-77-reactive T cells are of Th2-type in HLA-DRB1*1501 transgenics and are pathogenic Th1/Th17 cells in the HLA-DQB1*0602 transgenic mice. This new humanized model of MS further implicates autoimmunity against MOBP in MS pathogenesis, provides the first evidence of pathogenic HLA-DQ-associated anti-myelin autoimmunity, and is the first to offer a rationale for HLA-DQB1*0602 association with MS. These findings have important bearing on the candidacy of the DQB1*0602 allele as a genetic risk factor for MS.
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http://dx.doi.org/10.4049/jimmunol.0900784DOI Listing
September 2009

Activation and control of pathogenic T cells in OSP/claudin-11-induced EAE in SJL/J mice are dominated by their focused recognition of a single epitopic residue (OSP58M).

Int Immunol 2008 Nov 18;20(11):1439-49. Epub 2008 Sep 18.

Department of Immunology, Weizmann Institute of Science, Rehovot, Israel.

Oligodendrocyte-specific protein (OSP)/claudin-11 has been recently implicated in multiple sclerosis pathophysiology. Yet, the pathogenic autoimmunity against OSP has been poorly investigated. We previously showed that OSP-induced experimental autoimmune encephalomyelitis (EAE) and optic neuritis in SJL/J mice are primarily associated with CD4+ T cells reactive against OSP55-80. Dissecting the fine epitope specificity to the level of epitopic residues recognized by OSP-specific encephalitogenic T cells revealed their focused recognition of OSP58M. Accordingly, OSP58M predicted by computer modeling to be a major TCR contact residue shared by the three nonameric core epitopes within OSP55-80, albeit at different MHC-II pockets, was experimentally determined as the primary TCR contact residue crucial for activation and control of encephalitogenic T cells reactive against OSP55-80 or against recombinant OSP. Ala substitution of OSP58M impaired the functional TCR recognition/activation of pathogenic OSP-reactive T cells. Accordingly, the non-stimulatory/non-encephalitogenic pOSP55-58A-66 analogue not only treated EAE induced by pOSP55-80 but also effectively reversed EAE induced by whole OSP. Thus, the selection/activation and control of OSP-pathogenic T cells in H-2(s) mice appeared to be dominated by their predetermined focused recognition of OSP58M. Such a focused recognition by OSP-pathogenic T cells, despite their extensive TCR heterogeneity (Kaushansky, N., Zhong, M. C., Kerlero de Rosbo, N., Hoeftberger, R., Lassmann, H. and Ben-Nun, A. 2006. Epitope specificity of autoreactive T and B cells associated with experimental autoimmune encephalomyelitis and optic neuritis induced by oligodendrocyte-specific protein in SJL/J mice. J. Immunol. 177:7364), may impact profoundly on peripheral self-tolerance to OSP and on altered peptide ligand-mediated immune-specific modulation of the recently described OSP-related autoimmune pathogenesis.
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http://dx.doi.org/10.1093/intimm/dxn099DOI Listing
November 2008

Pathogenic T cells in MOBP-induced murine EAE are predominantly focused to recognition of MOBP21F and MOBP27P epitopic residues.

Eur J Immunol 2007 Nov;37(11):3281-92

Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.

Myelin-associated oligodendrocytic basic protein (MOBP) is a central nervous system (CNS)-specific myelin constituent important in stabilizing the unique multi-layered structure of the CNS-myelin sheath. Although autoimmunity against MOBP has been associated with multiple sclerosis pathogenesis, anti-MOBP pathogenic T cells have been poorly investigated as compared to T cells against other encephalitogenic myelin proteins. We have recently determined MOBP15-36 as the major encephalitogenic epitope of MOBP for SJL/J mice. In this study, epitope-specificity was dissected to the level of residues crucial for activation/control of MOBP-specific encephalitogenic T cells. Structural bioinformatic analysis of MOBP15-36/I-A(s) interaction and experimental data have determined MOBP21F and MOBP27P of I-A(s)-binding nonameric core epitope (MOBP20-28) as TCR-contact residues critical for functional activation of encephalitogenic MOBP-specific T cells. Accordingly, the non-encephalitogenic/nonstimulatory pMOBP16-21A27A-33 inhibited encephalitogenic MOBP-reactive T cells, shifted their cytokine secretion profile, and effectively suppressed pMOBP15-36-induced EAE. Moreover, pMOBP16-21A27A-33 fully reversed ongoing clinical EAE induced by whole MOBP as well as by pMOBP15-36. Data show that encephalitogenic MOBP-reactive T cells are predominantly focused to recognition of MOBP21F and MOBP27P, and suggest that both their selection and control are governed by MOBP21F and MOBP27P epitopic residues. Such focused epitopic recognition may affect profoundly on peripheral self-tolerance to MOBP and on potential altered peptide ligand-mediated therapy of MOBP-related autoimmune pathogenesis.
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http://dx.doi.org/10.1002/eji.200737438DOI Listing
November 2007

OSP/claudin-11-induced EAE in mice is mediated by pathogenic T cells primarily governed by OSP192Y residue of major encephalitogenic region OSP179-207.

Eur J Immunol 2007 Jul;37(7):2018-31

Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.

Pathogenic autoimmunity against oligodendrocyte-specific protein (OSP/claudin-11), recently implicated in multiple sclerosis (MS) pathophysiology, has been poorly investigated as compared to that against other myelin encephalitogens. Using recombinant soluble mouse OSP (smOSP) and overlapping peptides thereof, we show that smOSP-induced chronic EAE in C57BL/6J mice is primarily associated with CD4(+) T cells reactive against OSP179-207 and OSP22-46, the major and minor encephalitogenic regions, respectively, and with a predominant B cell response against OSP22-46. The encephalitogenic OSP179-207-specific T cells recognized OSP190-202 as minimal stimulatory epitope, while minimal encephalitogenic sequence was OSP191-199. Further delineation and structural bioinformatic analysis of the major encephalitogenic region suggested four overlapping potential I-A(b) core epitopes, predicting OSP192Y as major TCR-contact residue shared by OSP 188-196, OSP190-198, and OSP191-199 cores, albeit at different MHC-II pockets. Accordingly, substitution at OSP192Y yielded OSP188-192A-202, a non-stimulatory/non-encephalitogenic altered peptide ligand (APL) that was antagonistic for OSP188-202-specific encephalitogenic T cells. Systemic administration of OSP188-192A-202 suppressed OSP188-202-induced EAE and fully reversed smOSP-induced EAE. These data suggest that a single epitopic residue (OSP192Y) governs the selection and control of most pathogenic T cells associated with smOSP-induced EAE in H-2(b) mice. This may impact profoundly on peripheral self-tolerance to OSP and on potential APL-mediated therapy of OSP-related autoimmune pathogenesis.
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http://dx.doi.org/10.1002/eji.200636965DOI Listing
July 2007

Epitope specificity of autoreactive T and B cells associated with experimental autoimmune encephalomyelitis and optic neuritis induced by oligodendrocyte-specific protein in SJL/J mice.

J Immunol 2006 Nov;177(10):7364-76

Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.

The encephalitogenic potential of oligodendrocyte-specific protein (OSP) in mice, its specific localization in the intralamellar tight junctions in CNS myelin, and the detection of autoreactivity against OSP in multiple sclerosis (MS) strongly suggest the relevance of autoreactivity against OSP in the pathogenesis of MS. In this study, we have characterized the autoimmune T and B cells that are associated with clinicopathological manifestations of OSP-induced MS-like disease in mice by using recombinant soluble mouse OSP (smOSP) and synthetic overlapping peptides spanning smOSP. SJL/J mice immunized with smOSP developed chronic relapsing clinical experimental autoimmune encephalomyelitis accompanied with intense perivascular and parenchymal inflammatory infiltrates, widespread demyelination, axonal loss, and remarkable optic neuritis. The smOSP-primed lymph node cells reacted predominantly against OSP55-80 and to a lesser extent also to OSP22-46 and OSP179-207. Unexpectedly, in vitro selection with smOSP resulted in pathogenic smOSP-specific CD4+ T cells that reacted equally well against OSP55-80, OSP22-46, OSP45-66, and OSP179-207. Fine analysis of the anti-OSP autoimmunity revealed that the disease is primarily associated with CD4+ T cells directed against the major (OSP55-80) and the minor (OSP179-207) encephalitogenic regions that were further delineated, both in vitro and in vivo, to OSP55-66 and OSP194-207, respectively. In contrast, the OSP-induced Abs were predominantly directed against OSP22-46; these Abs were mostly of IgG1 isotype, but high levels of IgG2a and IgG2b and significant levels of IgE were also observed. The reactivity of pathogenic T cells to two encephalitogenic regions, OSP55-80 and OSP179-207, and their diverse TCRVbeta gene repertoire may impose difficulties for epitope-directed or TCR-targeting approaches to immune-specific modulation of OSP-related pathogenesis.
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http://dx.doi.org/10.4049/jimmunol.177.10.7364DOI Listing
November 2006