Dependence Receptors in Health and Disease: demonstrating whether serine proteases influence the proliferation and migration of specified cell populations

Reference Number: NDORMS 2018/11

Epidemiological evidence supports a role for diet in inflammation and the initiation or development of cancer. There are positive associations between red and processed meat consumption and carcinogenesis and negative associations (protection) from fruits and vegetables. In addition, chronic inflammation contributes significantly to the initiation and development of cancers [1-3]. This proposal examines a potential explanation of these phenomena with substantial implications for ameliorating chronic inflammatory disease and reducing cancer incidence.

Dependence receptors regulate cell viability and survival. When deprived of their local, extracellular ligands the dependence receptors induce cell apoptosis, preventing cells escaping from their home tissue and establishing an inappropriate colony elsewhere. In the absence of the dependence receptors, ligand-induced over-stimulation of cells leads to hyper-proliferation and increased migration.

A major group of dependence receptors includes Deleted in Colorectal Cancer (DCC), neogenin and Uncoordinated-5 (unc5), all of which have the extracellular netrin proteins as primary ligands and all were originally described as tumour suppressors until additional roles in cell guidance were recognised: the netrin/DCC or netrin/neogenin interactions generate cell attraction, while unc5 or the complex of DCC with unc5 generates cell repulsion. All these receptors also exist in lymphocytes: DCC is found in T and B cell populations, where levels are increased by anti-CD40 activation [4] and the expression of netrin is increased by interferon-ɣ during immune stimulation.

Serine proteases such as chymotrypsin increase cell proliferation and migration with high levels associated with several cancers, correlating with the development of malignant disease. Conversely, serine protease inhibition or down-regulation reduces cell migration and invasiveness, while reduced concentrations of serine protease inhibitors promote oncogenesis.

Recent evidence indicates that two serine proteases - the mammalian digestive protease chymotrypsin and the bacterial chymotrypsin-like subtilisin - selectively deplete cells of their DRs and this effect may underlie changes in proliferation and migration. The potential importance of this lies in the increased secretion of chymotrypsin with over-eating and the use of subtilisin or Bacillus subtilis in food processing (e.g. meat tenderisation) and farm animal probiotics.

There are several other chymotryptic serine proteases secreted by cells attracted to regions of infection or inflammation: mast cells and neutrophils secrete chymotrypsin, chymase, cathepsin G and granzyme. Other serine proteases linked with cancer incidence include Prostate Specific Antigen (PSA), the pro-protein convertases and the plasminogen families. If these serine proteases share the activity of chymotrypsin and subtilisin, the results could help to explain their association with cancer progression. Finally we shall examine the effects of the kynurenines on DR expression since these inflammation-induced compounds modulate T cell differentiation and activity as well as the migration of cancer cell lines. The compounds are proving to have clinical relevance in neurobiology, transplantation, and graft rejection but little is understood about their relationship to cancer.

Our hypothesis is that chymotryptic serine proteases produced during inflammation deplete cell DRs, leading to increased proliferation and cell migration and that these serine proteases also act on DRs in T cells and macrophages to induce a differential accumulation of pro-inflammatory and carcinogenic cells [5, 6].

The experimental aim is to demonstrate whether serine proteases influence the proliferation and migration of specified cell populations promoting or suppressing cancer development in parallel with changes in DR expression.

In this study work will be performed on cell cultures prepared from human blood and in animal models of inflammation [7] and cancer. We shall examine the effects of chymotryptic enzymes on the proliferation and migration of different populations of monocytes and lymphocytes. The functional studies will include proliferation assays and cell migration assays. In parallel we shall quantify the expression of DR in the different cell populations to examine correlations between DR presence and cell function. We shall also consider that serine protease treatment may have differential effects on cytokine or chemokine production and release under resting or stimulated conditions and we shall therefore quantify an appropriate panel of cytokines and chemokines using ELISAs and RT-PCR as necessary. We shall also address the question of whether serine protease involvement in inflammation and cancer directly using in vivo models.

The Williams group (https://www.kennedy.ox.ac.uk/research/targeting-disease-pathways-in-rheumatoid-arthritis) includes two senior scientists (Richard Williams and Trevor Stone), two post-docs (Felix Clanchy and I-Shu Huang) and two DPhil students (Wen-Yi Tseng and Federica Borghese). The project will be jointly supervised by Richard Williams [Email Address Removed] and Trevor Stone [Email Address Removed].

Training
The Kennedy Institute is a world-renowned research centre, housed in a brand new, state-of-the-art facility at the University of Oxford which enables and encourages research and education into the causes and treatment of musculoskeletal and other diseases. Training will be provided in techniques including in vivo imaging, flow cytometry, assays of immune cell function, migration assays, immunohistochemistry and animal models.

A core curriculum of lectures will be taken in the first term to provide a solid foundation in a broad range of subjects including musculoskeletal biology, inflammation, epigenetics, translational immunology and data analysis.

Students will attend weekly seminars within the department and those relevant in the wider University.

Students will be expected to present data regularly to the department, the Williams group and to attend external conferences to present their research globally.

How to Apply
The department accepts applications throughout the year but it is recommended that, in the first instance, you contact the relevant supervisor(s) or the Directors of Graduate Studies who will be able to advise you of the essential requirements.

Interested applicants should have or expect to obtain a first or upper second class BSc degree or equivalent, and will also need to provide evidence of English language competence. The University requires candidates to formally apply online and for their referees to submit online references via the online application system.

The application guide and form is found online and the DPhil or MSc by research will commence in October 2018.

When completing the online application, please read the University Guide: https://www.ox.ac.uk/admissions/graduate/applying-to-oxford/application-guide?wssl=1