The role of macrophage-derived granulin in pancreatic cancer metastasis - mechanisms and therapeutic opportunities

Pancreatic cancer is a very aggressive disease, and kills around 8,000 people every year in the UK and ~330,000 worldwide. Current treatments are not very effective, thus new treatment strategies are urgently needed. Among the major challenges to fight pancreatic cancer, is the fact that, soon after a pancreatic tumour develops , pancreatic cancer cells rapidly spread to distant organs in the body, where they can form additional tumours. The spreading of cancer cells from the primary tumour site, in this case the pancreas, to the distant secondary site, is called metastasis. Pancreatic cancer metastasis most often occurs to the liver. However, even though tumours can release large numbers of cancer cells into the circulation, only a small fraction of these cells are able to successfully survive and grow, at the distant metastatic site. Thus, the successful colonisation and outgrowth of metastatic tumour cells, in the new distant hostile environment, is a severe rate limiting step during metastasis. Identifying the key inter-cellular signalling pathways that allow cancer cells to successfully grow at the metastatic site could provide new therapeutic opportunities.
Although malignant cancer cells are the main drivers of metastatic spreading, recent evidence suggests that the colonisation of a new organ, by metastatic tumour cells, critically depends on the support of non- cancerous stromal cells. The stromal compartment comprises of different non-malignant cell types of the host, including cells which are involved in physiological processes associated with wound healing and tissue repair (myofibroblasts), and white blood cells from our immune system.
While it was originally thought that white blood cells would kill the cancer cells, recent evidence suggest that white blood cells lose their cell killing abilities during tumour development, and actually promote cancer progression. Thus, the reactivation of the cancer cell killing abilities of tumour infiltrating white blood cells holds great promise in the fight of many solid cancers, including pancreatic cancer. However, our understanding of how white blood cells at the metastatic sites affect our immune response against metastatic pancreatic cancer cells remains elusive.
The project proposed here focuses on understanding the role of the host stroma in pancreatic cancer metastasis. While most pancreatic cancer studies conducted so far have focused on the stromal compartment at the primary tumour site, the role of stromal cells at the secondary metastatic site remains poorly understood. We recently found that non-malignant stromal cells, including white blood cells and myofibroblasts, form a hospitable metastatic niche allowing efficient metastatic growth of disseminated pancreatic cancer cells (Nielsen et al., Nature Cell Biology, May 2016). In addition, our early work shows that a specific type of white blood cells, known as macrophages, not only promote the formation of a growth permitting environment, but also prevent the immune system from killing cancer cells.
Together our findings provide evidence that metastatic stromal cells promote pancreatic cancer metastasis, and that targeting metastatic promoting functions of metastasis associated stromal cells may provide new therapeutic opportunities to fight metastatic pancreatic cancer.

Thus, in this proposal, our overall aim is to gain a better understanding of how stromal cells promote pancreatic cancer metastasis in order to develop new therapeutic approaches for metastatic pancreatic cancer patients.

I believe that the here proposed project addresses a critical clinical need, and important emerging questions in the pancreatic cancer field. These studies will contribute to the increase of our understanding of pancreatic cancer metastasis, and consequently may lead to better diagnostic and/or more effective treatments for this devastating disease.

We have recently observed that pancreatic cancer liver metastasis critically depends on macrophage-derived granulin (Nielsen et al., Nature Cell Biology, May 2016). However, the molecular mechanisms by which granulin promotes the formation of a hospitable metastatic niche, and the therapeutic potential of targeting pro-metastatic functions of stromal cells at the metastatic site remain unknown.

Aim:
The here proposed study aims to identify and target signalling pathways that promote the pro-metastatic functions of stromal cells at the metastatic site in PDAC.

Objectives and planned methodologies:

This study will:
1. Generate and use primary hepatic stellate cells and primary macrophages in co-culture assays to characterise the interaction between these two stromal cell types.

2. Use a proteomic approach to identify how macrophage-derived granulin induces pro-metastatic functions of hStCs.

3. Evaluate in vitro and in vivo the identified pro-metastatic factors for their ability to sustain metastatic cancer cell growth and tumour immunity.

4. Use pre-clinical pancreatic tumour metastasis models in combination with genetic tools to ablate stromal cell populations (i.e. MAMs and activated hStCs) to characterize their role in tumour immunity.

5. Test in pre-clinical metastatic PDAC models the therapeutic potential of combining stroma-targeted therapies with a T cell checkpoint inhibitor.

Results:
The pre-clinical testing of potential therapies will identify treatment options that can be translated into clinical trials. In using both genetic and pharmacological approaches, this programme of work maximises the likelihood of successfully identifying key targets for metastatic pancreatic cancer treatment.

My research team is part of the Pancreas Research Group in Liverpool. The Pancreas Research Group concentrates on investigating fundamental cellular mechanisms relevant to the pathophysiology of acute pancreatitis and pancreatic ductal adenocarcinoma (PDAC) and the translation into clinical outputs. Clinical members of the group (including Prof Palmer, Prof Neoptolemos, Prof Ghaneh) hold prominent roles in the pancreatic cancer community, and will ensure the translation of our findings into the clinics. The MRC funding requested in this application will further underpin conceptual and technical developments in the Pancreas Research Group necessary for translational progression.

In the framework of this project, all information and knowledge acquired will be immediately shared with the Pancreas Research Group and the Liverpool NIHR Pancreas Biomedical Research Unit, and could be used for the informed design of pharmacological approaches for the treatment of PDAC. All technical information and cell-based assays performed will also be shared with my colleagues, and could be utilised for the development of procedures to test new pharmacological compounds in pancreatic cancer patients. The Pancreas Research Group and the Liverpool NIHR Pancreas Biomedical Research Unit have established contacts with pharmaceutical companies interested in developing or testing compounds which could alleviate PDAC. The relevant conceptual knowledge and methodological expertise generated in the course of our project will be conveyed to collaborating pharmaceutical companies. Collaborators are also actively involved in communicating the results of their research to the general public (see Pathways to Impact).

The advanced equipment (e.g. flow and mass cytometry, in vivo imaging techniques, high resolution nanoLC-MS/MS on an Orbitrap Q-Exactive instrument) available at the University of Liverpool (UoL) and the technical expertise accumulated at UoL (e.g. Centre for Proteome Research, Centre for Pre-clinical Imaging, Centre for Genomic Research) will complement high throughput instruments and techniques established in the NIHR Pancreas Biomedical Research Unit to create an optimal technology platform, necessary for both, discovery and translation. Importantly, the Pancreas Research Group serves as a training base for MRC, Wellcome Trust and North West Cancer Research PhD students, as well as for clinical fellows undertaking research projects.

The MRC support will allow me to further strengthen and expand my team here at UoL, and to further develop my expertise and interest in the tumour microenvironment of PDAC, which ideally complement existing research efforts here at UoL. The postdoctoral research scientist, the research associate funded by this application, and myself, will strongly be involved in the collaboration with existing research groups at UoL as well as with our national (Prof Jeffrey Pollard, University of Edinburgh; Prof Duncan Jodrell, CRUK Cambridge Institute) and international collaborators (Prof David Tuveson, Cold Spring Harbor, USA). The MRC support for the salary of a postdoctoral scientist and a research associate will also allow the establishment and maintenance of technical skills and transfer of expertise to future PhD students.

The MRC support of the current grant application will therefore not only provide resources necessary for the specific project, aiming to characterise crucial interactions of stromal cells in PDAC metastasis, but will also i) enhance local, national, and international collaborative efforts to fight PDAC, ii) help the developmental of interdisciplinary approaches, and iii) strengthen the training base here in Liverpool for PhD students with an interest in tumour inflammation, cancer cell biology, and metastasis.