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  • Project No: #OxKEN-2022/13
  • Intake: OxKEN 2022

PROJECT OVERVIEW

Hypoxia complicates most human diseases, and the immune system operates in the resultant environment. Oxygen-homeostatic transcriptional responses are controlled by the hypoxia-inducible factor (HIF) pathways, regulated by the oxygen-sensing HIF hydroxylases (PHD 1-3 and FIH) [1]. We recently discovered that global silencing of PHD2, the major oxygen-sensitive hydroxylase controlling HIF, results in spontaneous development of systemic lupus erythematosus (SLE)-like autoimmunity, associated with impaired regulatory T cell (Treg) function in mice. Importantly this phenotype is reversible when PHD2 is re-expressed [2].

More recently, we tested the immune effects of environmental hypoxia on normal unchallenged adult mice to investigate whether the magnitude of HIF hydroxylase inhibition resulting from physiologically tolerable levels of hypoxia would be sufficient to influence immune status. Systemic hypoxia did produce a small HIF2a-dependent increase in lymph node size, milder than that seen with PHD2 silencing, but associated with an increased incidence of anti nuclear antibody (ANA) positivity (but little evidence of tissue inflammation). Furthermore, we have found that the ability of splenocytes to kill mycobacteria in vitro is enhanced following BCG immunisation combined with hypoxic exposure compared to BCG immunisation alone, mediated at least in part through HIF system effects in Tregs. Importantly, HIF induction via prolyl hydroxylase inhibition is already being used as a treatment for renal anaemia [3] and drugs inhibiting HIF2 dimerisation are showing promising results in the treatment of renal cancer [4].

In this project we will test the hypotheses that 1) HIF pathway induction can potentiate autoimmune responses/phenotypes and 2) that blocking endogenous HIF pathway induction or suppressing HIF2a can enhance immune regulation and ameliorate autoimmune phenotypes. Specifically, we will examine the effects of manipulating the HIF pathway (genetically, by altering oxygen supply, or pharmacologically) in mouse models of autoinflammatory and autoimmune conditions. Initial studies will focus on two models of SLE, TLR7 agonism with Imiquimod, which induces self-reactive antibody production and immune complex mediated renal damage consistent with lupus nephritis and MRL/lpr mice which provide a good polygenic model of multi-system human lupus.  Both models can be combined with hypoxic or pharmacological manipulation of the HIF pathway and the Imiquimod model can be applied to mice with genetic HIF pathway manipulations. Sharpin deficient and NOD mice are also available and these experiments are all covered by existing animal licence permissions.

We will then extend this work to investigate the underlying mechanisms linking changes in HIF2a activity to changes in Treg phenotype, but potentially considering effects in other cell types highlighted by the models. Mechanistic studies will combine state of the art approaches including single cell and bulk sequencing, targeted CRISPR and/or small molecule interventions using both animal (perhaps including our humanised mouse models [5]) and in vitro assays (using human or mouse leukocytes). The goal of this latter work being not only to advance knowledge and relate findings to human disease but also to identify intermediary targets that could allow the immune response to be reversibly and precisely tuned without entraining the wide effects of the entire HIF transcriptional pathways.

KEYWORDS

Hypoxia; autoimmunity; SLE; Treg; HIF

TRAINING OPPORTUNITIES

Generic skills training would be provided through access to the resources of the University’s Graduate School.This covers areas such as experimental design, literature searching, coding, statistics, research presentations and scientific writing.

The project work would involve training in specific skills including, but not restricted to:

  • use of animal models;
  • informatics relating to single cell sequencing, including RNA velocity;
  • signal pathway analysis;
  • use of tissue culture models;

and potentially

  • Cas9/CRISPR based genetic modification of cells;
  • small molecule or RNAi based screens.

Attendance at meetings run by both the Hypoxia Biology Group and Transplantation Research and Immunology Group would ensure a broad grounding in the field of studies. Attendance at seminar series run across the University and meetings held with BMS would add diversity, exposure to a commercial mind-set and exposure to other methodologies.

In addition, the recipient of the Fellowship would receive support from the Oxford University Clinical Academic Graduate School which Chris Pugh directs. This would help with career development and acquisition of skills necessary to progress a clinical academic career, including advice about future grant applications and access to Clinical Lectureships.

KEY PUBLICATIONS

  1. Pugh, C. W. & Ratcliffe, P. J. Exp Cell Res 356(2):116-121 (2017).
  2. Yamamoto et al. J Clin Invest 130, 3640-3656 (2019).
  3. Chen et al. N Engl J Med 381(11):1011-1022(2019). 
  4. Courtney et al. J Clin Oncol 36, 867-874 (2018).
  5. Adigbli et al. doi: 10.1097/TP.0000000000003177 Transplantation. (2020).

CONTACT INFORMATION OF ALL SUPERVISORS

Fadi Issa

Katherine Bull

Joanna Hester

Chris Pugh