Identifying disease promoting macrophages and tissue-identity in endometriosis

Endometriosis affects 176 million women worldwide and is associated with debilitating pelvic pain and/or infertility. Endometriosis is defined by the presence of tissue similar to womb lining (endometrium) outside the womb (lesions), most commonly on the wall of the pelvic cavity. Endometriosis is currently treated by invasive surgery or with drugs that suppress sex hormones. Sadly, in many women symptoms recur after surgery and available medical treatments have undesirable side-effects and are contraceptive. New treatments for endometriosis are desperately needed.

Why some, but not all, women with endometriosis develop debilitating symptoms is unknown. What we do know is that endometriosis lesions contain nerves and have high numbers of immune cells called 'macrophages' within them. Macrophages are known to adapt their function depending upon the tissues in which they exist and several types have been identified. Long-lived 'tissue-resident' macrophages maintain normal tissue function. In contrast, during inflammation or following injury immature 'macrophages' called monocytes infiltrate tissues where they mature into macrophages (monocyte-derived macrophages) and play roles vital in clearing cell debris and microbes as well as stimulating the immune system.

In endometriosis, macrophages play an essential role in controlling the growth of lesions and regulating the infiltration of blood vessels. Using an experimental mouse model of endometriosis, we have shown that macrophages can regulate the infiltration of nerves into lesions and play an important role in generating a pain response. Currently, we do not know if different 'types' of macrophages exist in endometriosis lesions or if a specific type promotes disease. If we can identify the 'type' of macrophage that drives the disease we could target it without affecting 'good' macrophages elsewhere in the body and develop new treatments for endometriosis that do not involve surgery or hormonal manipulation and associated side-effects.

Our preliminary data shows that macrophages in lesions have different origins: the endometrium shed from the uterus, the fluid lubricating the pelvic cavity, and others are monocyte-derived (from blood). Our latest findings suggest that amongst these there are multiple different 'types' of macrophages and we believe these different macrophages play distinct roles in endometriosis. In this project, we will identify which macrophages drive the disease. We also aim to define what it is about the lesion that can cause macrophages to change into a type of macrophage that supports and increases the severity of the disease.

We will initially use our laboratory mouse model of endometriosis and extend our findings using biopsies from women with endometriosis. Our objectives are:
1. To determine how many different 'types' of endometriosis macrophages exist and determine the genetic 'signature' of each population
2. To identify where macrophages in endometriosis lesions come from and what happens to them (do they multiply, die or change 'type')
3. To determine which 'type' of macrophage promotes endometriosis
4. Identify what it is about endometriosis lesions that create a pro-disease 'type' of macrophage.

Our world-class research environment, clinical resources and experienced team of investigators means we are uniquely placed to exploit new methods that will allow us to identify the disease causing 'type' of macrophages in this debilitating disorder and develop new therpeutic interventions.

Endometriosis is a chronic inflammatory condition associated with debilitating pelvic pain and/or infertility that affects 176 million women worldwide. It is defined by the presence of endometrial-like tissue ('lesions') outside the uterus, commonly in the pelvic cavity. Treatments for endometriosis are limited and recurrence rates are high. There is an unmet need for new therapies.

Macrophages are present at high densities in lesions and play a vital role in the pathophysiology of endometriosis by regulating lesion growth, vascularisation, neurogenesis and pain generation. In cancers, macrophages with different origins exist side-by-side. 'Tissue-resident' macrophages become educated by the tumour and promote disease whilst infiltrating 'monocyte-derived' macrophages act as innocent bystanders. We do not know if different subsets of macrophages exist in endometriosis, or if a subset promotes disease. Our preliminary data indicates that lesion-resident macrophages have multiple origins e.g. endometrium, peritoneum and circulating monocytes. We also have evidence that significant phenotype heterogeneity exists amongst lesion-resident macrophages.

We aim to identify which macrophages drive endometriosis and how the lesion educates 'pro-disease' macrophages using our in-house endometriosis mouse model with validation in patient biopsies. The identification of 'pro-disease' macrophages in endometriosis could inform the design of a therapy targeting pathogenic macrophages whilst leaving those critical for normal tissue function intact.

Our objectives are to:
1. Determine phenotype diversity amongst endometriosis macrophages using scRNA-Seq
2. Use lineage tracing to determine ontogeny and fate of lesion-resident macrophages
3. Use targeted depletion strategies combined with imaging of lesions to determine which population drives disease
4. Identify lesion 'tissue-identity' signals required for macrophage recruitment/ differentiation using antibody arrays

To be read in conjunction with Pathways to Impact.

Patients
Endometriosis is a chronic inflammatory disorder associated with debilitating pelvic pain and infertility. In the UK, ~1.5 million women suffer from endometriosis. Current treatments include surgical excision of lesions or drugs that cause ovarian suppression. National patient organisations report that women want to avoid surgery and to have access to better medical treatments with fewer side effects.
We aim to identify the disease promoting macrophage population in endometriosis and the factors responsible for recruitment / differentiation of this population. Our research is intended to inform the design of a targeted immunotherapy. We envisage that our studies will lead to a clinical trial which if successful will increase the treatment options for women allowing them to avoid surgery and the side-effects of current hormone therapies. A timescale of 10 years is expected for this to be realised.

Biomedical research community
We will generate and make publically available, single cell omics data which will impact researchers in the fields of endometriosis, macrophage biology and other inflammatory conditions. Endometriosis is associated with a number of co-morbidities e.g. IBS and CV disease, the impact of our research may also extend to researchers working in these fields. We are using a novel in-house mouse model that will be adapted and humanized to answer particular questions. These preclinical platforms will be available for the biomed community to use. In the past, we have trained students and postdocs to implement the model. The platform has already been taken up by others and we expect the same will occur with new adaptations. We will collect biopsies from endometriosis patients, under internationally-recognised EPHect guidelines, which will be used for our own related research locally, as well as for collaboratve research with other EPHect registered academic centres worldwide.

Policy makers
AWH and PTKS were members of the Steering Group for the Endometriosis Priority Setting Partnership managed under the auspices of the James Lind Alliance run by NIHR. The results were published in the Lancet, highlighted in a BMJ editorial and covered in the national press. The top two priorities stemming from the project were 1) Can a cure be developed for endometriosis? 2) What causes endometriosis? In the current and follow-up proposals, we will be addressing both these research priorities.

Commercial sector
The research will inform the design of a future immunotherapy for endometriosis. IP generated during the project will be protected and we will seek to attract collaborative input from Industry for the development of a therapy. We will collaborate with nanostringTM through their Technology Access Program to implement digital spatial profiling on human biopsies.

Training and capacity building
The research will allow the postdoc to gain experience in using a mouse model, lineage tracing, scRNA-Seq and bioinformatics. The technician will learn skills in animal handling, surgery and histological analysis. The postdoc will gain experience in presenting / defending their research at national / international conferences and contributing to public engagement. Students will have the opportunity to work on the project; the junior academics will gain hands on lab experience and mentoring and the postdoc will develop supervisory and teaching skills.

Public and wider society
We will engage the public at all stages of the research. This will enhance our ability in communicating our research and may lead to new ideas. By publically sharing our research we will generate informed citizens that understand endometriosis, increasing awareness. Engagement with the public will foster transparency for the science that the UK tax payers fund. We also hope to inspire the next generation of scientists.