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mammary gland development and breast cancer
mammary gland development and breast cancer
Mammary gland development requires complex interactions between mammary epithelial cells and their surrounding environment. The overarching research focus of the Schwertfeger laboratory is to define the mechanisms that contribute to normal mammary gland development and function and to understand how aberrant regulation of these mechanisms drives breast cancer initiation, growth and malignant progression. The long-term goal of our research program is to use this knowledge to enhance the development of effective therapeutic and diagnostic strategies for breast cancer patients. The following examples highlight the approaches the laboratory has taken to 1) understand the complex mechanisms that drive epithelial-stromal interactions in the context of normal mammary gland development and 2) use this information to gain novel insights into the mechanisms through which the tumor microenvironment impacts breast cancer progression and response to therapy.
A key focus of research in the Schwertfeger laboratory is on understanding the biological importance of tissue resident macrophages. These are innate immune cells that reside within distinct organ sites and contribute to immune surveillance, among numerous other functions. Depending on their sites of localization, these macrophages can exhibit additional tissue-specific behaviors, which are driven in response to factors within the local tissue microenvironment. While tissue resident macrophages are present in the normal mammary gland, their tissue-specific functions and the mechanisms by which they respond to the local tissue environment have not been extensively characterized. In pathologies such as breast cancer, macrophages in the tumor-associated stroma can both inhibit and promote malignant progression. As described below, our laboratory has recently characterized distinct populations of tissue resident macrophages in the mammary gland with the goal of identifying their localization and function in the context of mammary gland development.
The Schwertfeger lab is also actively engaged in studies to identify tissue-specific factors and intracellular signaling pathways that drive resident macrophage function within the mammary gland. These studies led to the identification of a specific signaling pathway, the JAK/STAT pathway, which is critical for driving resident macrophage function during mammary gland development (Brady et al, 2017). Based on our understanding of how this pathway impacts macrophages in the normal mammary gland, we have also determined that this pathway is critical for driving macrophage functions in the tumor-associated stroma. As described below, this earlier work has led to a novel potentially clinically relevant area of investigation in which we have found that inhibition of JAK/STAT signaling in tumor associated macrophages leads to resistance of breast cancer cells to JAK-targeted therapies. Based on these findings, we identified a novel combinatorial approach to enhance the efficacy of JAK inhibitors in breast cancer. These findings provide an example of the complexities by which host cells within the tumor microenvironment can interfere with tumor cell targeted therapies. A major goals moving forward is to expand this line of investigation to identify key targetable pathways in macrophages that impact on therapy. Exploitation of these macrophage pathways in combination with tumor cell-based therapeutics could effectively limit progression and ultimately enhance patient outcome.
Identification of a distinct macrophage subpopulation in the mammary gland --- Lyve-1+ macrophages
Tissue resident macrophages in the mammary gland are found in close association with epithelial structures and within the adipose stroma, and are important for mammary gland development and tissue homeostasis. While epithelial-associated macrophages have been linked to ductal development, the contributions of stromal macrophages to mammary gland homeostasis remain unknown. Using transcriptional profiling and single cell RNA sequencing approaches, we identify a distinct resident stromal macrophage subpopulation that is characterized by expression of Lyve-1, a receptor for the extracellular matrix component hyaluronan. This subpopulation is enriched in genes associated with extracellular matrix remodeling and is found to be specifically associated with hyaluronan-rich regions within the mammary stroma. Furthermore, macrophage depletion leads to increased accumulation of hyaluronan-associated extracellular matrix in the mammary stroma. These results demonstrate the presence of a distinct subpopulation of macrophages and provide insights into the functional contributions of these macrophages to stromal homeostasis in the mammary gland. Next step, we are aiming to investigate the mechanisms by which this subpopulation modulate the stroma in the mammary gland and how does this population function in the context of breast tumor and its role in breast cancer progression.
FGFR signaling pathway in breast cancer progression and metastasis
We utilize the inducible fibroblast growth factor receptor (iFGFR) transgenic mouse model to study the macrophage populations and functions in the context of breast cancer. This is an inducible model of mammary tumorigenesis in which oncogene activation is induced following treatment of mice with the synthetic dimerizer B/B. This is a well characterized model in which mammary gland development is normal until iFGFR1 is activated using B/B, which leads to formation of estrogen receptor positive pre-neoplastic lesions eventually leading to tumor formation. However, the low tumor take rate and low metastasis rate is potential downside to overcome. Hence, we developed a derivative iFGFR tumor model based on the parental iFGFR model. The new derivative tumor cell line shows higher tumor take rate and extensive lung metastasis, which will enable us to exploit this cell line in other projects.
Tumor Cell Associated Hyaluronan-CD44 Signaling Promotes Pro-Tumor Inflammation in Breast Cancer
Cancer has been conceptualized as a chronic wound with a predominance of tumor promoting inflammation. Given the accumulating evidence that the microenvironment supports tumor growth, we investigated hyaluronan (HA)-CD44 interactions within breast cancer cells, to determine whether this axis directly impacts the formation of an inflammatory microenvironment. Our results demonstrate that breast cancer cells synthesize and fragment HA and express CD44 on the cell surface. Using RNA sequencing approaches, we found that loss of CD44 in breast cancer cells altered the expression of cytokine-related genes. Specifically, we found that production of the chemokine CCL2 by breast cancer cells was significantly decreased after depletion of either CD44 or HA. In vivo, we found that CD44 deletion in breast cancer cells resulted in a delay in tumor formation and localized progression. This finding was accompanied by a decrease in infiltrating CD206+ macrophages, which are typically associated with tumor promoting functions. Importantly, our laboratory results were supported by human breast cancer patient data, where increased HAS2 expression was significantly associated with a tumor promoting inflammatory gene signature. Because high levels of HA deposition within many tumor types yields a poorer prognosis, our results emphasize that HA-CD44 interactions potentially have broad implications across multiple cancers.
JAK/STAT inhibition in macrophages promotes therapeutic resistance by inducing expression of protumorigenic factors
In the breast cancer field, the development of small molecule and other tumor cell-targeted therapies has been instrumental in enhancing patient survival. However, there are still patient populations, including those with triple negative and metastatic breast cancer, for which tumor cell targeted therapies are not effective. Many of the targeted therapies were developed to inhibit oncogenic signaling pathways in tumor cells. While these inhibitors work well in cell culture models and pre-clinical studies, most of them show low levels of efficacy in patients. As a result of our earlier work on the importance of macrophage JAK/STAT activation in regulating mammary gland development, we initiated studies to address the potential impact of JAK/STAT inhibitors on tumor associated macrophages. Our results demonstrated that JAK/STAT inhibitors promote a protumorigenic phenotype in tumor associated macrophages, which compromises the ability of JAK/STAT inhibitors to limit tumor progression in our animal models. We extended these findings to identify an alternative combination therapy to inhibit the macrophage mediated resistance mechanism to enhance the efficacy of JAK/STAT inhibitors on mammary tumors (Irey and Lassiter et al. 2019). We are currently working to identify the pathways activated in tumor associated macrophages by JAK/STAT inhibitors, which lead to the observed pro-tumor response. We also aim to identify combination therapies including JAK/STAT inhibitors that inhibit primary tumor growth and metastasis.
Investigating the role of LYVE-1+ macrophages in breast tumor invasion and metastasis
The LYVE-1+ macrophage subpopulation has been identified in the normal mammary gland as well as mammary tumors. Interestingly, LYVE-1+ macrophages present in mammary tumors have been found specifically at the tumor margin (left, Wang and Chaffee, eLife, 2020). LYVE-1 itself is a transmembrane protein commonly associated with the lymphatic endothelium and has been more recently described as a macrophage subpopulation marker. LYVE-1 is also known to bind hyaluronic acid (HA). When in high molecular weight form, HA is considered homeostatic and tumor promoting. When fragmented to low molecular weight HA, it is tumor promoting. Further, the LYVE-1/HA binding event is thought to trigger cellular internalization of both proteins, resulting in fragmented HA. Therefore, due to the unique localization and associated with HA fragmentation, the LYVE-1+ macrophage population is theorized support tumor invasion in mammary tumors. The Schwertfeger Lab continues to study LYVE-1+ macrophages in mammary tumors and investigate their contributions to invasion and metastasis.
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