Mammary Stem Cells

Source: Diana Dragoi

Our research is at the crossroads of basic cell biology, translational cancer research and tissue engineering. As our main experimental system, we develop and continuously improve organoid models based on primary human cells to address fundamental biological questions and stratify patients for precision medicine.

Plasticity, defined as the ability of cells to dynamically change cell state, is crucial for many processes during mammary gland morphogenesis and homeostasis. In breast cancer pathogenesis, dysregulated plasticity causes cellular heterogeneity, thereby promoting tumor progression and complicating therapy.
Our goal is to identify mechanisms of cellular plasticity to determine processes and patterns of breast cancer initiation and progression that guide histopathological and molecular classification.
To address these questions mechanistically, we focus on elucidating the role of cell-cell and cell-matrix adhesion, intra- and extracellular physical forces and redox state in dictating cell-function and –state. Moreover, we dissect dynamic reactivation of developmental pathways in breast cancer metastatic progression. In the mammary gland, proliferation and invasion are delicately balanced during development and homeostasis. We propose that this dynamic balance also determines the capacity of breast cancer cells to generate actively proliferating metastases.

Although overall breast cancer mortality is decreasing, 5-year survival rates of women with advanced disease, i.e. distant metastases, remain below 20%. Plasticity contributes to the enormous inter- and intra-tumor heterogeneity detected at the clinical, histopathological, and molecular level, which greatly impairs patient stratification. Common therapeutic regimens, particularly systemic chemotherapy, cause chronic health problems and a significant increase in secondary malignancies, such as leukemia. Consequently, there is an urgent need to identify patients who will not benefit from systemic therapy. By contrast, breast cancer subtypes with a high risk of progression need to be diagnosed early to be treated aggressively.

1) Primary human mammary epithelial cells (HMEC)
The dynamic nature of cell plasticity complicates classification and treatment of breast cancer. Our goal is to identify mechanisms that govern cell plasticity in the human mammary gland to define patterns of breast cancer initiation and progression. For this purpose, we have developed a human mammary organoid assay that recapitulates ductal architecture, branching morphogenesis and regenerative capacity.
For functional analyses, we have established protocols for conditional overexpression or knock down of transcripts by lentiviral transduction of cDNAs or shRNAs. We currently quantify effects on morphogenesis (length of ducts, number of side-branches, polarization, lumen formation, presence of duct-leading tip cells) and differentiation (expression of multiple lineage markers at correct positions) by confocal immunofluorescence and live-cell imaging.

2) From normal to transformed state
To utilize our organoid assay for breast cancer modeling, we work on establishing new models for immortalized and in vitro transformed HMEC. To this end, we have established lineage-specific immortalization of HMEC using lentiviral transduction of defined genetic elements. Moreover, we have isolated and characterized hundreds of single-cell clones from immortalized HMEC that have been proven highly valuable to delineate signaling-context-dependent responses to EMT-transcription factors such as Twist1, and thereby, acquisition of invasiveness and metastatic competence. Currently, we are in the process to set up CRISPR/Cas9-directed functional genomics, transcriptional repression, activation and epigenetic modification as a screening-compatible tool to engineer primary cells and derive cell lines. We engage in several productive collaborations to complement this experimental approach with patient-derived cancer cells.

We are supported by a “Max Eder” Young Investigator Start-up Grant of the “Deutsche Krebshilfe” (German Cancer Aid) and work in close collaboration with our clinical partners at the Institute of Pathology of the Ludwig-Maximilian University and the Department of Internal Medicine at the Technical University in Munich.