Using a murine bioengineering model to study cancer cell biology: the effects of mammographic density on breast cancer progression

© 2017 Dr. Cecilia Wanchen HuoBreast cancer (BC) remains a leading cause of cancer-related morbidity and mortality for women worldwide. Mammographic density (MD) has been well recognized as a strong risk factor for BC, independent of its masking effect for small tumours on a mammogram. Due to the common presentation of high MD (HMD) in the community, especially in pre-menopausal women, MD is arguably the most important risk factor for BC when taking into account of its high population-attributable risk. However, much remains to be learned about the biological mechanism underlying MD-associated BC risk. My team previously utilised a murine engineering biochamber model to show that human breast tissues of high and low MD not only remained viable, but also maintained their radiological and histological features in relation to their MD status for at least 6 weeks. Extending from the prior findings, this PhD study aimed to further explore the biological mechanisms behind MD-associated BC risk, and is divided into four main sections: (i) I first attempted to develop the murine biochamber model further by implanting collagenase digested and flow cytometry sorted single cells to pave the way for manipulations of specific cell types that might be responsible for HMD. I found that collagenase digested and flow cytometry sorted single cells of high or low MD breast tissue reconstituted glandular organoids in murine chambers, albeit in limited numbers; (ii) second, based on a collection of human high and low MD breast tissues from prophylactic mastectomy procedures over a period of 5 years, I evaluated the histological differences between within-individual high and low MD mammary specimens of all participants, and found that the HMD tissue microenvironment was significantly altered compared with that of LMD -- HMD was characterised by increased levels of collagen organisation and quantity, aromatase immunoreactivity and immune cell infiltration of various subtypes; (iii) The serendipitous finding of increased immune cell presence in HMD led to my subsequent examination of the potential differences in immune cell representation between patient-matched high and low MD tissues, the immune infiltrates of both innate and adaptive system, and cytokines such as IL-6 and IL-4, and I found that immune cells of various subtypes were significantly raised in HMD tissue compared with LMD; and (iv) parallel to the human breast tissue studies, I also tested whether high and low MD human tissue had any direct effect on cancer cell growth and dissemination; using our murine biochamber model, I showed that compared to co-inoculation with LMD tissue, HMD tissue stimulated the progression of MCF10DCIS.com cells that represented cells of ductal carcinoma in situ (DCIS), to lesions resembling invasive ductal carcinomas, as well as their metastases in the mice hosting the murine biochambers. Over the past twenty years, the body of evidence on various aspects of MD and its associated BC risk has been expanding, however, to the best of my knowledge, (i) my study contained the largest cohort of high-risk women in Australia to characterise immunohistochemical and immune cell differences between high and low MD, and (ii) the work presented in this thesis is the first to utilise the murine biochamber xenograft model to test human breast organoids formation from single cells and to evaluate the direct effect of MD on human breast cancer cell progression in vivo. Collectively my work has defined that HMD is characterised by an increase in stromal cells, extracellular matrix and inflammation. I have shown that HMD stimulated the progression of early stage BC cells, which highlights the importance of MD being considered for BC diagnoses, treatments and surveillances