Impact of targeting myeloid PI3Kg on the pancreatic tumour stroma compartment and the response to chemotherapy.

Pancreatic cancer is very aggressive and kills around 8000 people every year in the UK. New treatment strategies are urgently needed. During tumour (cancer) formation, cancer cells attract cells from the immune system (white blood cells), which progressively accumulate in the tumour environmental bed (or stroma), producing an appearance of inflammation. Although it was originally thought that tumour infiltrating immune cells would kill the cancer cells, recent evidence suggests that they may actually promote tumour growth. In the tumour stroma certain immune cells, secrete high amounts of stimulatory factors that aid the cancer cells to multiply, survive, and spread. The abundance of white blood cells may determine whether the patient will die of the cancer or not.

The tumour infiltrating white blood cells still have the capability to kill cancer cells, but their anti-cancer functions are inhibited by cancer cells themselves and are skewed towards cancer promoting functions. Our preliminary results indicate that blockade of a specific molecule (known as PI3K gamma) present in certain white cells called macrophages, inhibits their cancer promoting functions and restores their initial cancer cell killing ability. Since macrophages are critical players in regulating the immune response, studying and targeting this recently identified function of PI3K gamma represents a promising approach to re-activate the immune system to fight against cancer.

The pancreatic cancer stroma comprises of two other types of normally resident cells known as pancreatic stellate cells and fibroblasts. These cells are responsible for secreting a high amount of substances called extracelluar matrix proteins that provide the scaffolding (or matrix) for the shape of the tissue and also the glue to hold the cells to the scaffolding. This is the stroma. In pancreatic cancer there is far too much of this stroma which means that anti-cancer drugs cannot properly reach the cancer cells to kill them.

Our early work shows that in pancreatic cancer patients, macrophages are found side by side with stellate cells, indicating a potential interaction between these cell types. Since we are able to change macrophage function by targeting PI3K gamma, we are now in the position to use this tool to study a so far uncharacterized interaction of stellate cells with macrophages.

Therefore, these studies will increase our knowledge concerning the cancer promoting functions of tumour associated macrophages and might reveal new key proteins that regulate their function and contribute to the poor response to chemotherapy. Our findings will also open new strategies to improve the treatments of pancreatic cancer.

The project proposed here focuses on the role of the host immune cells during pancreatic cancer progression and its response to chemotherapy. While most research has so far focused on the cancer cells themselves and their precursor cells, it has become clear that the immune system and the formation of a tumour microenvironment also influences the course of pancreatic cancer progression and its response to chemotherapy.

I believe that the here proposed project addresses important emerging questions in the pancreatic cancer field and that these studies will contribute to increase the understanding of how tumour inflammation impacts on pancreatic cancer progression.

This will help lead to better diagnosis and better treatments.

Pancreatic ductal adenocarcinoma (PDA) has one of the highest mortality rates of any malignancy and effective treatment is major unmet need. A marked infiltration of myeloid cells into the stromal compartment and the generation of a desmoplastic stromal reaction is a particular characteristic of PDA and is thought to play a key role in disease progression and the poor response to therapy.Consequently, there is an urgent need to develop effective therapies targeting the stromal compartment in PDA.

Aim:
The proposed study aims to identify and target myeloid signalling pathways that promote pro-tumourigenic myeloid cell functions and desmoplasia in pancreatic cancer.

Objectives and planned methodologies:
This study will:

1. Use pre-clinical pancreatic tumour models in combination with genetic tools to inhibit gene expression (e.g. PI3Kg) in tumour associated macrophages.

2. Generate and use primary pancreatic stellate cells and primary macrophages in co-culture assays to characterise the interaction between these two stromal cell types.

3. Use a proteomic approach to identify PI3Kg downstream effectors regulating macrophage conversion, tumour immunity, PSC activation and desmoplasia.

4. Evaluate identified factors in vitro and in vivofor their impact on tumour immunity and desmoplasia formation.

5. Test in pre-clinical mouse PDA models whether blockade of PI3Kg and/or the newly identified downstream effectors in combination with standard chemotherapy (gemcitabine or 5FU) improves therapeutic outcome.

Results:
The preclinical 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 specific novel targets that are necessary for the tumour promoting functions of macrophages and provide the rationale for developing new anti-cancer therapeutic approaches.

The Pancreas Research Group in Liverpool concentrates on investigating fundamental cellular mechanisms relevant to the pathophysiology of acute pancreatitis and pancreatic ductal adenocarcinoma (PDA) and translation into clinical outputs. The work of our team is closely co-ordinated through the Centre of Excellence for Pancreas Cancer of the Liverpool Cancer Research UK Centre and the Liverpool NIHR Pancreas Biomedical Research Unit, which is the only NIHR Research Unit focused on translational pancreatic research. The MRC funding requested in this application will further underpin conceptual and technical developments in the Pancreas Research Group necessary for translational progression. This Group coordinates the efforts of clinicians, cell biologists and physiologists, organic chemists, structural biologists and pharmacologists of the University of Liverpool aiming to develop drugs against PDA (as well as pancreatitis).

In the framework of this project, all information and knowledge acquired will be immediately shared with the Pancreatic Research Group and could be used for the informed design of pharmacological approaches for the treatment of PDA. All technical information and cell-based assays performed will also be shared with the Pancreatic Research Group and could be utilised for the development of procedures to test new pharmacological compounds in pancreatic cancer patients. The Pancreatic Research Group has established contacts with pharmaceutical companies interested in developing or testing compounds which could alleviate PDA. 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. multicolour flow cytometry, in vivo imaging techniques, state-of-the-art Aperio Technologies Slide Scanning System for TMA analysis) available in the Pancreas Research Group and the technical expertise accumulated in the group (e.g. vast experience in staining, scoring and interpreting data from TMAs with assistance from pancreas histopathologists, and the isolation and culturing of primary human and murine pancreatic cancer and stellate cells) will complement high throughput instruments and techniques established in the NIHR 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 Cancer Research UK PhD students, as well as for clinical fellows undertaking research projects.

The MRC support will allow me to establish my team among the existing pancreatic research teams here, within the Liverpool Pancreas Research Group and my expertise and interest in the tumour microenvironment of PDA will ideally complement existing research efforts. The postdoctoral research scientist funded by this application and myself, will strongly be involved in the collaboration with the Liverpool Cancer Research UK Centre and the Liverpool NIHR Pancreas Biomedical Research Unit. The MRC support for the salary of an experienced postdoctoral scientist will also allow the establishment and maintenance of technical skills and transfer of expertise in advanced experimentation 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 PDA disease progression, but will also enhance the collaborative efforts and the translational research of the Pancreas Research Group and strengthen the training base for PhD students with an interest in tumour inflammation and immunity, cell biology and in vivo tumour models of PDA.