Abstract
Breast cancer has one of the highest mortality rates among cancers afflicting women. Despite six decades of cancer treatment using chemotherapeutic drugs and hormonal and radiotherapy, many breast cancer patients ultimately develop drug resistance and succumb to their disease. Breast cancer recurrence is attributed to the ability of cancer cells to persist, maintain, and repopulate themselves, effectively surviving therapies designed to target rapidly-dividing cells. One population of cancer cells in particular, termed cancer stem cells (CSCs) for their stem cell-like ability of self-renewal, has emerged as the crux of disease. CSCs can simultaneously proliferate the CSC population as well as the non-CSC population that constitutes the rest of the tumor. Standard therapies have the ability to target actively dividing cells, but if more curative treatments are to be established, the proliferative state of CSCs must be determined. Due to poor prognoses for patients who do not have the benefit of early detection, it is paramount that primary prevention is addressed to avoid relapse and drug resistance. A chronological investigation of three different cell types: non-cancerous, pre-cancerous, and cancerous cells will make establishing dormancy and tracking growth in vivo possible. The origin of the CSC is unclear as it is not understood whether a non-cancerous stem cell is altered in some way or whether a differentiated cell takes on the characteristics of a CSC. Of importance, is the analyses of a diverse set of cancer cell lines characterized for breast CSC markers, CD44+/CD24-/low. By characterizing cell lines with differential percentages of stem-like cells, a methodology that can identify CSC populations in both well-characterized cell lines as well as those characterized to a lesser extent can be developed to be utilized for complex tumor samples. Relying on the dual CD44 and CD24 phenotypic expression of CSCs, they can be isolated using fluorescence cell-activated sorting (FACS) of cultured cells. Cell sorting using antibodies allows for recognition of cells exhibiting the CD44+/CD24-/low phenotype. The goal of the project was to compare the non- and CSC populations phenotypically, in terms of marker expression in order to establish more effective therapies by targeting both types of populations. The first objective involved optimizing methods for the identification of persistent stem-cell like populations across two cell lines using cell surface markers. Our data questions the reliance upon purportedly effective antibody markers, namely CD44. By performing FACS analysis on two established human breast cancer cell lines, our data did not support the use of CD44 for isolation of stem cell populations. The very foundation of the project was more closely examined – the use of widely accepted antibody markers. The results obtained from this project have provided useful insight into the varied efficacy of antibody markers and the need for the discovery and study of more efficient and reproducibly reliable breast cancer stem cell markers.