Grant: CIHR-ICR / Cancer Research Society Partnership - Res. Gr. Cancer Prev.

Endometriosis-associated ovarian carcinomas (EAOC) and endometriosis impact genetically and biologically female individuals, primarily those with an intact uterus, including cisgender women, transgender men, and non-binary people. Up to 1 million individuals in Canada have endometriosis and are potentially in need of a way to reduce or mitigate ovarian cancer risk. While other types of ovarian cancer are plateaued EAOCs are becoming more common, particularly in Asian and Hispanic populations. EAOC prevention-focused research will help mitigate future racial and ethnic disparities in ovarian cancer incidence and mortality. EAOCs, including clear cell and endometrioid ovarian carcinoma, arise from endometriosis and represent >30% of ovarian carcinomas. EAOCs are thought to arise from stepwise accumulation of cell intrinsic factors (somatic alterations) and the development of a permissive microenvironment, potentially including dysbiosis, that together create a cancer-prone state. Functional contributions of cancer-driver genes are relatively well studied, though interactions with the microbiome are not. Recent studies have highlighted the presence of microbial pathogens, in particular Fusobacterium nucleatum, within endometriosis lesions, cancers, and specifically linked their presence to DNA damage. My team has pioneered genomic analyses of EAOCs, its origins from endometriosis, and normal-appearing uterine tissue. In each we have identified cancer driver mutations affecting genes such as KRAS, ARID1A and PIK3CA. We have also used these mutations and other features for prognostic molecular classifications of EAOCs. Herein, we propose to investigate the prevalence and diversity of the intralesional microbiome in EAOCs and their precursor endometriosis, the microbiome relationship with DNA damage, onset of disease, and molecular subtypes. Our core hypothesis is that endometriosis lesions destined for malignant progression are more likely to harbor DNA damage associated microbial pathogens, such as F.nucleatum. Pathogens will be associated with evidence of genotoxic stress and DNA damage and could be further enriched in specific EAOC subtypes. Aim 1: will identify microbial presence in both cancer and precursor lesions. Here we will leverage genomic and biomarker data being collected from a cohort of ~60 cases of endometriosis that later developed EAOC (as well as resulting cancers, and controls that did not develop cancer). • In Aim 1A we will apply marker-gene sequencing for bacterial and fungal taxa to interrogate endometriosis from patients that later developed EAOC, endometriosis from patients that did not develop cancer, and molecularly defined clear cell and endometrioid ovarian carcinomas. • In Aim 1B we will use florescence in-situ hybridization to validate the presence of pathogens and establish their spatial position. Aim 2: will investigate DNA damage response signatures in endometriosis cohorts (as above) using multi-colour immunohistochemistry surrogates such as staining for gamma-H2AX, cytosolic DNA, and replication fork stalls (R-loop) indicating DNA damage and/or oncogene-induced stress responses. Central to this proposal is identifying potential high-risk features in precursors of EAOC. Identifying and acting on tumor precursor lesions is an established mechanism to successfully prevent cancers: programs to detect and remove colon polyps, cervical intraepithelial neoplasia, or early skin cancers have prevented hundreds of thousands of cancer deaths. Often, preventive procedures are targeted to a high-risk population, such as risk-reducing salpingo-oophorectomy for carriers of germline mutations in