Report
Teaser, summary, work performed and final results
Periodic Reporting for period 1 - Tolltum (Role, regulation and targeting of nucleic acid-sensing TLRs within the tumour microenvironment)
Teaser
The early detection of tumour cells by the immune system is crucial for stalling carcinogenesis and for tumour regression. Immunotherapy is gaining momentum as a primary treatment option owing to its success in treating tumours of different origins. However, the mechanisms...
Summary
The early detection of tumour cells by the immune system is crucial for stalling carcinogenesis and for tumour regression. Immunotherapy is gaining momentum as a primary treatment option owing to its success in treating tumours of different origins. However, the mechanisms leading to failure of the immune system to control tumour formation in the absence of therapy are poorly understood and requires more attention. Insights into these mechanisms would provide therapeutic targets to enhance the efficacy of immunotherapy and boost the endogenous immune response. The primary goal of this study is to identify mechanisms by which tumour cells prevent the activation and function of the immune system thereby leading to an inefficient anti-tumour response.
Breast cancer ranks as the first in cancer deaths and the primary cause of death is tumour metastasis. Studies in mice and humans have shown that metastatic tumour cells develop several evasion mechanisms that suppress their detection by the immune system. The present project aims to shed light on these suppressive mechanisms that would provide novel therapeutic targets to deal with metastatic disease.
The overall objective of this project is to use primary human tumours and established metastatic cancer cell lines to study cellular and molecular interactions within the tumour microenvironment that suppress the immune response. Our main focus is on inflammation and the recruitment of key immune cells within the tumour both of which are crucial to induce an efficient immune response.
Work performed
During the two years of the Marie Curie fellowship I have worked on two complementary projects.
I. Effect of metastatic breast cancer cells on the inflammatory response
An efficient immune response requires the sensing of danger by the immune system. Within tumours, this danger signal is the presence of extracellular nucleic acids released upon chemo or radiotherapy and in DNA-rich neutrophil extracellular traps (NETs) released by neutrophils under some circumstances. The sensing of nucleic acid by cells leads to the production of type I interferons (IFN-I), a family of cytokines that has been shown to have strong anti-tumour and anti-metastatic properties.
My work has revealed that human metastatic breast cancer cells induce the formation of DNA-rich NETs by neutrophils. NETs have previously been shown to favor the metastatic potential of cancer cells. Interestingly, the recognition of nucleic acids in NETs by key immune cells, dendritic cells (DCs), is suppressed by metastatic cancer cells. The absence of IFN-I production leads to the failure of an anti-tumour immune response and potentiates metastasis. Thus tumour cells use a double-pronged approach to propagate, by inducing metastasis-favorable NETs on one hand while suppressing their detection by immune cells on the other hand.
I have further characterized that the suppression of nucleic acid recognition is mediated by soluble proteins secreted by tumour cells that bind to free nucleic acid shielding them from sensing by the myriad receptors present in DCs. The identification of these suppressive proteins reveal novel therapeutic targets to increase nucleic acid-sensing.
Identification of target nucleic acid-binding proteins will be the basis of a patent application providing us with the opportunity to collaborate with pharmaceutical companies to generate specific drugs.
II. Effect of tumour-associated fibroblasts on disease burden and immune cell infiltration
Fibroblasts infiltrate tumours and form the supporting stroma. Cancer-associated fibroblasts (CAFs) have been linked to the progression of disease and metastasis. CAFs keep cytotoxic T cells out of the tumour while their selective ablation leads to tumour regression. Few studies have focused on the role of CAFs on other cells of the immune system. Particularly, the impact on the infiltration and function of DCs remains unknown. The present work attempts to shed light on the effect of CAFs on DC subsets.
In primary human clear cell Renal Cell carcinomas (ccRCC), we found that a sub-population of fibroblasts where enriched within and around the tumour. The presence of these CAFs co-related with the severity of the disease. Furthermore, systematic multi-panel flow cytometry analysis of ccRCC’s revealed that a key DC subset, CD141+ DCs, failed to enter the tumour. Absence of CD141+ cells co-related to the presence of the CAF subset around the tumour suggesting that CAFs may be responsible for the block in entry. At present, we are performing deep sequencing analysis of sorted fibroblasts to ascertain factors that could be responsible for this blockade. Targeting these factors would enable entry of DCs into the tumour where they could efficiently sample tumour antigens and mount a specific immune response.
Results from both projects are part of grant applications for further funding to continue the research. Publication of the results are underway in scientific journals of high impact. Both projects reveal novel pathways of immune suppression; the secretion of nucleic acid-binding soluble proteins and the impact of fibroblasts on the infiltration of DCs into tumours. The publication of these results will open paths for further investigation into similar mechanisms not only during carcinogenesis but also in infectious diseases and autoimmunity.
Final results
In Europe in 2012, there were an estimated 3.5 million new cancer cases and 1.9 million cancer deaths, just over one quarter (26.0Â %) of the total number of deaths. Breast cancer is the most common cancer in women and is the third most common cause of cancer death. Although advances in the early detection and improved therapies has decreased mortality rates, metastasis remains the main cause of death. Combating metastatic disease is a challenge in all age groups of women and few treatment options are available. Immunotherapy has shown success in a small percentage of patients while the reasons for failure in most patients is unknown. New therapeutic targets are in dire need to be able to extend the efficacy of immunotherapy.
Mis-localisation of nucleic acid-binding proteins to the extracellular space, reveals a novel strategy exerted by malignant cells to hide prototypical danger signals, nucleic acids, from the detection by the immune system. Targeting such soluble proteins would potentiate inflammation that is pre-requisite to anti-tumour immunity. The present study has thus opened doors to study a novel pathway of immune suppression and provides druggable targets.
The tumour microenvironment and particularly the stroma provides an essential backdrop for the growth of cancer cells. Understanding the interaction between cells within this environment is crucial to understand the progression of the tumour. Several groups are studying the cellular composition of various human tumours using multipanel flow and mass cytometry approaches. Our study combines flow cytometry and deep sequencing of sorted cells to provide functional evidence to the phenotypic co-relation we find within primary human ccRCCs. We hope to find targetable pathways that are a specific to the CAF population that we have identified. This would provide us with tools to manipulate the tumour microenvironment.