P S Ohashi

University Health Network, Toronto, Ontario, Canada

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Publications (266)3086.76 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: During the course of many chronic viral infections, the antiviral T cell response becomes attenuated through a process that is regulated in part by the host. While elevated expression of the immunosuppressive cytokine IL-10 is involved in the suppression of viral-specific T cell responses, the relevant cellular sources of IL-10, as well as the pathways responsible for IL-10 induction, remain unclear. In this study, we traced IL-10 production over the course of chronic lymphocytic choriomeningitis virus (LCMV) infection in an IL-10 reporter mouse line. Using this model, we demonstrated that virus-specific T cells with reduced inflammatory function, particularly Th1 cells, display elevated and sustained IL-10 expression during chronic LCMV infection. Furthermore, ablation of IL-10 from the T cell compartment partially restored T cell function and reduced viral loads in LCMV-infected animals. We found that viral persistence is needed for sustained IL-10 production by Th1 cells and that the transcription factor BLIMP-1 is required for IL-10 expression by Th1 cells. Restimulation of Th1 cells from LCMV-infected mice promoted BLIMP-1 and subsequent IL-10 expression, suggesting that constant antigen exposure likely induces the BLIMP-1/IL-10 pathway during chronic viral infection. Together, these data indicate that effector T cells self-limit their responsiveness during persistent viral infection via an IL-10-dependent negative feedback loop.
    The Journal of clinical investigation. 07/2014;
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    ABSTRACT: Despite development of new antiviral drugs, viral infections are still a major health problem. The most potent antiviral defense mechanism is the innate production of type I interferon (IFN-I), which not only limits virus replication but also promotes antiviral T cell immunity through mechanisms, which remain insufficiently studied. Using the murine lymphocytic choriomeningitis virus model system, we show here that IFN-I signaling on T cells prevented their rapid elimination in vivo. Microarray analyses uncovered that IFN-I triggered the expression of selected inhibitory NK-cell-receptor ligands. Consequently, T cell immunity of IFN-I receptor (IFNAR)-deficient T cells could be restored by NK cell depletion or in NK-cell-deficient hosts (Nfil3(-/-)). The elimination of Ifnar1(-/-) T cells was dependent on NK-cell-mediated perforin expression. In summary, we identified IFN-I as a key player regulating the protection of T cells against regulatory NK cell function.
    Immunity 06/2014; · 19.80 Impact Factor
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    ABSTRACT: CD8(+) T-cell functions are critical for preventing chronic viral infections by eliminating infected cells. For healthy immune responses, beneficial destruction of infected cells must be balanced against immunopathology resulting from collateral damage to tissues. These processes are regulated by factors controlling CD8(+) T-cell function, which are still incompletely understood. Here, we show that the interferon regulatory factor 4 (IRF4) and its cooperating binding partner B-cell-activating transcription factor (BATF) are necessary for sustained CD8(+) T-cell effector function. Although Irf4(-/-) CD8(+) T cells were initially capable of proliferation, IRF4 deficiency resulted in limited CD8(+) T-cell responses after infection with the lymphocytic choriomeningitis virus. Consequently, Irf4(-/-) mice established chronic infections, but were protected from fatal immunopathology. Absence of BATF also resulted in reduced CD8(+) T-cell function, limited immunopathology, and promotion of viral persistence. These data identify the transcription factors IRF4 and BATF as major regulators of antiviral cytotoxic T-cell immunity.Cell Death and Differentiation advance online publication, 14 February 2014; doi:10.1038/cdd.2014.19.
    Cell death and differentiation 02/2014; · 8.24 Impact Factor
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    ABSTRACT: D.B. and A.B. contributed equally to this puplication. The ability to mount a strong immune response against pathogens is crucial for mammalian survival. However, excessive and uncontrolled immune reactions can lead to autoimmunity. Unraveling how the reactive versus tolerogenic state is controlled might point toward novel therapeutic strategies to treat autoimmune diseases. The surface receptor Toso/Faim3 has been linked to apoptosis, IgM binding, and innate immune responses. In this study, we used Toso-deficient mice to investigate the importance of Toso in tolerance and autoimmunity. We found that Toso(-/-) mice do not develop severe experimental autoimmune encephalomyelitis (EAE), a mouse model for the human disease multiple sclerosis. Toso(-/-) dendritic cells were less sensitive to Toll-like receptor stimulation and induced significantly lower levels of disease-associated inflammatory T-cell responses. Consistent with this observation, the transfer of Toso(-/-) dendritic cells did not induce autoimmune diabetes, indicating their tolerogenic potential. In Toso(-/-) mice subjected to EAE induction, we found increased numbers of regulatory T cells and decreased encephalitogenic cellular infiltrates in the brain. Finally, inhibition of Toso activity in vivo at either an early or late stage of EAE induction prevented further disease progression. Taken together, our data identify Toso as a unique regulator of inflammatory autoimmune responses and an attractive target for therapeutic intervention.
    Proceedings of the National Academy of Sciences 01/2014; · 9.74 Impact Factor
  • Evan F Lind, Pamela S Ohashi
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    ABSTRACT: The activation of T cells is a tightly regulated process that has evolved to maximize protective immune responses to pathogens while minimizing damage to self-tissues. A delicate balance of cell-intrinsic, costimulatory, and transcriptional pathways as well as micro-environmental cues such as local cytokines controls the magnitude and nature of T-cell responses in vivo. The discovery of functional small noncoding RNAs called micro-RNAs (miRNAs) has introduced new mechanisms that contribute to the regulation of protein translation and cellular responses to stimuli. miRNAs are short (approximately 22 bp) RNA species, which bind to mRNAs and suppress translation. Due to their short length and imperfect base pairing requirements, each miRNA has the potential to regulate various pathways through the translational inhibition of multiple mRNAs. The human and mouse genomes each encode hundreds of miRNAs, and studying the function of miRNAs has led to the realization that they play important roles in diverse biological processes from development and cancer to immunity. This review focuses on the function of mir-155 in T cells and the impact of this miRNA on autoimmunity, tumor immunity, and pathogen-induced immunity.
    European Journal of Immunology 01/2014; 44(1):11-5. · 4.97 Impact Factor
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    ABSTRACT: Vaccines for cancer immunotherapy are of interest but in general have not yet achieved the desired therapeutic efficacy in clinical trials. We present here a novel model to evaluate vaccine strategies by following tissue destruction in a transgenic model, where a defined antigen is expressed on pancreatic islets. We found that the transfer of syngeneic antigen-pulsed dendritic cells (DCs) resulted in autoimmune cytotoxic T-lymphocyte activation that was not observed following vaccinations that were based on peptides and adjuvants. Importantly, the induction of diabetes by DC transfer is dependent upon the maturation of DCs prior to transfer. Furthermore, diabetes induction only occurred if DCs were pulsed with the immunodominant epitope in addition to at least one other peptide, suggesting greater cytolytic activity upon engagement of multiple T-cell specificities. While the tumor environment undoubtedly will be more complex than healthy tissue, the insights gained through this model provide useful information on variables that can affect CD8-mediated tissue cytolysis in vivo.
    PLoS ONE 01/2014; 9(3):e92380. · 3.73 Impact Factor
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    ABSTRACT: Immunotherapy is a promising means to fight cancer, prompting a steady increase in clinical trials and correlative laboratory studies in this field. As antitumor T cells play central roles in immunity against malignant diseases, most immunotherapeutic protocols aim to induce and/or strengthen their function. Various treatment strategies have elicited encouraging clinical responses; however, major challenges have been uncovered that should be addressed in order to fully exploit the potential of immunotherapy. Here, we outline pitfalls for the mobilization of antitumor T cells and offer solutions to improve their therapeutic efficacy. We provide a critical perspective on the main methodologies used to characterize T-cell responses to cancer therapies, with a focus on discrepancies between T-cell attributes measured in vitro and protective responses in vivo. This review altogether provides recommendations to optimize the design of future clinical trials and highlights important considerations for the proficient analysis of clinical specimens available for research.
    Expert Review of Vaccines 10/2013; · 4.22 Impact Factor
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    ABSTRACT: The American Joint Committee on Cancer/Union Internationale Contre le Cancer (AJCC/UICC)-TNM staging system provides the most reliable guidelines for the routine prognostication and treatment of colorectal carcinoma. This traditional tumor staging summarizes data on tumor burden (T), the presence of cancer cells in draining and regional lymph nodes (N) and evidence for distant metastases (M). However, it is now recognized that the clinical outcome can significantly vary among patients within the same stage. The current classification provides limited prognostic information, and does not predict response to therapy. Multiple ways to classify cancer and to distinguish different subtypes of colorectal cancer have been proposed, including morphology, cell origin, molecular pathways, mutation status, and gene expression-based stratification. These parameters rely on tumor-cell characteristics. Extensive literature investigated the host-immune response against cancer and demonstrated the prognostic impact of the in situ immune cell infiltrate in tumors. A methodology named "Immunoscore" has been defined to quantify the in situ immune infiltrate. In colorectal cancer, the Immunoscore may add to the significance of the current AJCC/UICC TNM classification since it has been demonstrated to be a prognostic factor superior to the AJCC/UICC TNM-classification. An international consortium has been initiated to validate and promote the Immunoscore in routine clinical settings. The results of this international consortium may result in the implementation of the Immunoscore as a new component for the classification of cancer, designated TNM-I (TNM-Immune).
    The Journal of Pathology 10/2013; · 7.59 Impact Factor
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    ABSTRACT: Differentiation of CD8 single-positive (SP) T cells is predicated by the ability of lymphocyte progenitors to integrate multiple signaling cues provided by the thymic microenvironment. In the thymus and the OP9-DL1 system for T cell development, Notch signals are required for progenitors to commit to the T cell lineage and necessary for their progression to the CD4(+)CD8(+) double-positive (DP) stage of T cell development. However, it remains unclear whether Notch is a prerequisite for the differentiation of DP cells to the CD8 SP stage of development. In this study, we demonstrate that Notch receptor-ligand interactions allow for efficient differentiation and selection of conventional CD8 T cells from bone marrow-derived hematopoietic stem cells. However, bone marrow-derived hematopoietic stem cells isolated from Itk(-/-)Rlk(-/-) mice gave rise to T cells with decreased IFN-γ production, but gained the ability to produce IL-17. We further reveal that positive and negative selection in vitro are constrained by peptide-MHC class I expressed on OP9 cells. Finally, using an MHC class I-restricted TCR-transgenic model, we show that the commitment of DP precursors to the CD8 T cell lineage is dependent on Notch signaling. Our findings further establish the requirement for Notch receptor-ligand interactions throughout T cell differentiation, including the final step of CD8 SP selection.
    The Journal of Immunology 07/2013; · 5.52 Impact Factor
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    ABSTRACT: The protein-tyrosine phosphatase Shp1 is expressed ubiquitously in hematopoietic cells and is generally viewed as a negative regulatory molecule. Mutations in Ptpn6, which encodes Shp1, result in widespread inflammation and premature death, known as the motheaten (me) phenotype. Previous studies identified Shp1 as a negative regulator of TCR signaling, but the severe systemic inflammation in me mice may have confounded our understanding of Shp1 function in T cell biology. To define the T cell-intrinsic role of Shp1, we characterized mice with a T cell-specific Shp1 deletion (Shp1(fl/fl) CD4-cre). Surprisingly, thymocyte selection and peripheral TCR sensitivity were unaltered in the absence of Shp1. Instead, Shp1(fl/fl) CD4-cre mice had increased frequencies of memory phenotype T cells that expressed elevated levels of CD44. Activation of Shp1-deficient CD4(+) T cells also resulted in skewing to the Th2 lineage and increased IL-4 production. After IL-4 stimulation of Shp1-deficient T cells, Stat 6 activation was sustained, leading to enhanced Th2 skewing. Accordingly, we observed elevated serum IgE in the steady state. Blocking or genetic deletion of IL-4 in the absence of Shp1 resulted in a marked reduction of the CD44(hi) population. Therefore, Shp1 is an essential negative regulator of IL-4 signaling in T lymphocytes.
    Journal of Experimental Medicine 06/2013; · 13.21 Impact Factor
  • Dylan J Johnson, Pamela S Ohashi
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    ABSTRACT: Dendritic cells are master regulators of immunity. Immature dendritic cells are essential for maintaining self-tolerance, while mature dendritic cells initiate a variety of specialized immune responses. Dendritic cell quiescence is often viewed as a default state that requires exogenous stimuli to induce maturation. However, recent studies have identified dendritic cell quiescence factors that actively program dendritic cells to an immature state. In the absence of these factors, dendritic cells spontaneously become immunogenic and can induce autoimmune responses. Herein we discuss two such factors, NF-κB1 and A20, that preserve dendritic cell immaturity through their regulation of NF-κB signaling. Loss of either of these factors increases dendritic cell immunogenicity, suggesting that they may be important targets for enhancing dendritic cell-based cancer immunotherapies. Alternatively, defects in molecules critical for maintaining steady-state DCs may provide novel biomarkers that identify patients who have enhanced natural antitumor immunity or that correlate with better responses to various immunotherapies.
    Annals of the New York Academy of Sciences 05/2013; 1284(1):46-51. · 4.38 Impact Factor
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    ABSTRACT: Natural killer (NK) cells are important mediators of the immune response against microbial pathogens and tumors. There is growing evidence from mouse and human studies that, NK cells exhibit immunoregulatory functions and can limit T cell immunity. NK cell regulatory activity has been demonstrated in a variety of disease models including chronic viral infection, autoimmunity, and transplantation. Depending on the nature of the immune challenge, NK cells use different strategies to limit T cell function, including via cytokines, interactions with NK receptors NKG2D and NKp46, or by perforin-mediated T cell death. Future work should address whether specific subsets of NK cells inhibit T cell responses, and how NK cells acquire immunosuppressive functions.
    Trends in Immunology 04/2013; · 9.49 Impact Factor
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    ABSTRACT: Cluster of differentiation (CD)8(+) T cells are like a double edged sword during chronic viral infections because they not only promote virus elimination but also induce virus-mediated immunopathology. Elevated levels of reactive oxygen species (ROS) have been reported during virus infections. However, the role of ROS in T-cell-mediated immunopathology remains unclear. Here we used the murine lymphocytic choriomeningitis virus to explore the role of ROS during the processes of virus elimination and induction of immunopathology. We found that virus infection led to elevated levels of ROS producing granulocytes and macrophages in virus-infected liver and spleen tissues that were triggered by the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Lack of the regulatory subunit p47phox of the NADPH oxidase diminished ROS production in these cells. While CD8(+) T cells exhibited ROS production that was independent of NADPH oxidase expression, survival and T-cell function was elevated in p47phox-deficient (Ncf1(-/-)) mice. In the absence of p47phox, enhanced T-cell immunity promoted virus elimination and blunted corresponding immunopathology. In conclusion, we find that NADPH-mediated production of ROS critically impairs the immune response, impacting elimination of virus and outcome of liver cell damage.
    Cell death and differentiation 04/2013; 40(4):649-58. · 8.24 Impact Factor
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    ABSTRACT: Background Wiskott-Aldrich syndrome (WAS) is a rare X-linked primary immunodeficiency caused by absence of Wiskott-Aldrich syndrome protein (WASP) expression, resulting in defective function of many immune cell lineages and susceptibility to severe bacterial, viral, and fungal infections. Despite a significant proportion of patients with WAS having recurrent viral infections, surprisingly little is known about the effects of WASP deficiency on antiviral immunity. Objective We sought to evaluate the antiviral immune response in patients with WASP deficiency in vivo. Methods Viral clearance and associated immunopathology were measured after infection of WASP-deficient (WAS KO) mice with lymphocytic choriomeningitis virus (LCMV). Induction of antiviral CD8+ T-cell immunity and cytotoxicity was documented in WAS KO mice by means of temporal enumeration of total and antigen-specific T-cell numbers. Type I interferon (IFN-I) production was measured in serum in response to LCMV challenge and characterized in vivo by using IFN-I reporter mice crossed with WAS KO mice. Results WAS KO mice showed reduced viral clearance and enhanced immunopathology during LCMV infection. This was attributed to both an intrinsic CD8+ T-cell defect and defective priming of CD8+ T cells by dendritic cells (DCs). IFN-I production by WAS KO DCs was reduced both in vivo and in vitro. Conclusions These studies use a well-characterized model of persistence-prone viral infection to reveal a critical deficiency of CD8+ T-cell responses in murine WASP deficiency, in which abrogated production of IFN-I by DCs might play an important contributory role. These findings might help us to understand the immunodeficiency of WAS.
    The Journal of allergy and clinical immunology 03/2013; 131(3):815-24. · 12.05 Impact Factor
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    ABSTRACT: Rapid activation of immune responses is necessary for antibacterial defense, but excessive immune activation can result in life-threatening septic shock. Understanding how these processes are balanced may provide novel therapeutic potential in treating inflammatory disease. Fc receptors are crucial for innate immune activation. However, the role of the putative Fc receptor for IgM, known as Toso/Faim3, has to this point been unclear. In this study, we generated Toso-deficient mice and used them to uncover a critical regulatory function of Toso in innate immune activation. Development of innate immune cells was intact in the absence of Toso, but Toso-deficient neutrophils exhibited more reactive oxygen species production and reduced phagocytosis of pathogens compared with controls. Cytokine production was also decreased in Toso(-/-) mice compared with WT animals, rendering them resistant to septic shock induced by lipopolysaccharide. However, Toso(-/-) mice also displayed limited cytokine production after infection with the bacterium Listeria monocytogenes that was correlated with elevated presence of Listeria throughout the body. Accordingly, Toso(-/-) mice succumbed to infections of L. monocytogenes, whereas WT mice successfully eliminated the infection. Taken together, our data reveal Toso to be a unique regulator of innate immune responses during bacterial infection and septic shock.
    Proceedings of the National Academy of Sciences 01/2013; · 9.74 Impact Factor
  • Journal of Pathology review. 01/2013; In Press:2013.
  • Evan F Lind, Alisha R Elford, Pamela S Ohashi
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    ABSTRACT: Recent studies have begun to define the role of micro-RNAs in regulating the immune response. Micro-RNA155 (mir-155) has been shown to play a role in germinal center formation, T cell inflammation, and regulatory T cell development. In this study, we evaluated the role of mir-155 in cytotoxic T cell function. We report in this study that mice lacking mir-155 have impaired CD8(+) T cell responses to infections with lymphocytic choriomeningitis virus and the intracellular bacteria Listeria monocytogenes. We show by a series of adoptive transfer studies that the impaired CD8(+) T cell response to L. monocytogenes is T cell intrinsic. In addition, we observed that CD8(+) T cells lacking mir-155 have impaired activation of the prosurvival Akt pathway after TCR cross-linking. These data suggest that mir-155 may be a good target for therapies aimed at modulating immune responses.
    The Journal of Immunology 12/2012; · 5.52 Impact Factor
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    ABSTRACT: PURPOSE: Breast carcinomas, including basal and hereditary cases, often present with a prominent tumoral lymphocytic infiltrate (LI). Chemokines could play a role in attracting these cells and contribute to tumor progression. We explored tumoral expression of CXCL10 and determined the relationship between CXCL10 and LI in a cohort of breast cancers. EXPERIMENTAL DESIGN: Using tissue microarrays of 364 breast tumors we evaluated expression of CXCL10 and its receptor, CXCR3, in relation to histopathologic features, biomarkers and lymphocyte markers. Additionally, we overexpressed CXCL10 and CXCR3 in MCF7 breast cancer cells and monitored T-lymphocyte migration and invasion. RESULTS: Forty-five percent of tumors expressed CXCL10 and a significant association was found with CXCR3 and LI. Further characterization of the LI revealed an association with CXCL10 expression for peritumoral CD4+ and CD8+ lymphocytes. CD8+ intratumoral lymphocytes, FOXP3+ Tregs and T-BET+ Th1 cells were associated with BRCA1 and basal tumors. Conditioned media from MCF7 cells overexpressing both CXCL10 and CXCR3 increased T-lymphocyte migration and invasion. CONCLUSIONS: Our findings suggest that CXCL10 may act in a paracrine manner, affecting the tumor microenvironment, and in an autocrine manner, acting on the tumor cells themselves and may play a role in tumor invasiveness and progression. The CXCL10-CXCR3 axis can serve as a potential target in BRCA1 and basal breast cancers which present with a prominent LI and a poor prognosis.
    Clinical Cancer Research 12/2012; · 7.84 Impact Factor
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    ABSTRACT: Despite efforts to understand and treat acute myeloid leukemia (AML), there remains a need for more comprehensive therapies to prevent AML-associated relapses. To identify new therapeutic strategies for AML, we screened a library of on- and off-patent drugs and identified the antimalarial agent mefloquine as a compound that selectively kills AML cells and AML stem cells in a panel of leukemia cell lines and in mice. Using a yeast genome-wide functional screen for mefloquine sensitizers, we identified genes associated with the yeast vacuole, the homolog of the mammalian lysosome. Consistent with this, we determined that mefloquine disrupts lysosomes, directly permeabilizes the lysosome membrane, and releases cathepsins into the cytosol. Knockdown of the lysosomal membrane proteins LAMP1 and LAMP2 resulted in decreased cell viability, as did treatment of AML cells with known lysosome disrupters. Highlighting a potential therapeutic rationale for this strategy, leukemic cells had significantly larger lysosomes compared with normal cells, and leukemia-initiating cells overexpressed lysosomal biogenesis genes. These results demonstrate that lysosomal disruption preferentially targets AML cells and AML progenitor cells, providing a rationale for testing lysosomal disruption as a novel therapeutic strategy for AML.
    The Journal of clinical investigation 12/2012; · 15.39 Impact Factor
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    ABSTRACT: The E3 ligase ARIH2 has an unusual structure and mechanism of elongating ubiquitin chains. To understand its physiological role, we generated gene-targeted mice deficient in ARIH2. ARIH2 deficiency resulted in the embryonic death of C57BL/6 mice. On a mixed genetic background, the lethality was attenuated, with some mice surviving beyond weaning and then succumbing to an aggressive multiorgan inflammatory response. We found that in dendritic cells (DCs), ARIH2 caused degradation of the inhibitor IκBβ in the nucleus, which abrogated its ability to sequester, protect and transcriptionally coactivate the transcription factor subunit p65 in the nucleus. Loss of ARIH2 caused dysregulated activation of the transcription factor NF-κB in DCs, which led to lethal activation of the immune system in ARIH2-sufficent mice reconstituted with ARIH2-deficient hematopoietic stem cells. Our data have therapeutic implications for targeting ARIH2 function.
    Nature Immunology 11/2012; · 26.20 Impact Factor

Publication Stats

20k Citations
3,086.76 Total Impact Points

Institutions

  • 2003–2014
    • University Health Network
      • Campbell Family Institute for Breast Cancer Research
      Toronto, Ontario, Canada
  • 1988–2014
    • University of Toronto
      • • Department of Immunology
      • • Department of Medical Biophysics
      Toronto, Ontario, Canada
  • 1988–2013
    • Ontario Institute for Cancer Research
      Toronto, Ontario, Canada
  • 2012
    • Cancer Research Institute
      New York City, New York, United States
    • University of Duisburg-Essen
      Essen, North Rhine-Westphalia, Germany
  • 2010–2012
    • The Princess Margaret Hospital
      Toronto, Ontario, Canada
  • 2010–2011
    • Walter And Eliza Hall Institute For Medical Research
      Melbourne, Victoria, Australia
  • 1989–2010
    • University of Zurich
      • • Institut für Experimentelle Immunologie
      • • Institute of Veterinary Pathology
      Zürich, ZH, Switzerland
  • 2005
    • Austrian Academy of Sciences
      • Institut für Molekulare Biotechnologie (IMBA)
      Vienna, Vienna, Austria
  • 2004
    • La Jolla Institute for Allergy & Immunology
      • Division of Cell Biology
      La Jolla, CA, United States
  • 2001
    • Osaka University
      • Division of Molecular and Cellular Pharmacology
      Ōsaka-shi, Osaka-fu, Japan
  • 1994–2000
    • Amgen Canada
      Mississauga, Ontario, Canada
  • 1999
    • University of Wuerzburg
      • Institute for Medical Radiation and Cell Research
      Würzburg, Bavaria, Germany
  • 1998
    • University of Lausanne
      Lausanne, Vaud, Switzerland
    • University Hospital of Lausanne
      Lausanne, Vaud, Switzerland