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Analysis Of Telomere Dysfunction For Early Breast Cancer Detection
Investigator Initiated Research
Rationale: The cancer stem cell theory proposes that cancers arise from malignant transformation of normal stem or progenitor cells. The inherent properties of normal stem/progenitor cells may impart their transformed counterparts with the ability to form invasive tumors, evade traditional anti-tumor therapies and establish metastasis. Telomere dysfunction is likely a key driving force behind the genomic instability observed in early cancer lesions. Since genomic instability is one of the earliest neoplastic changes known to occur in malignant transformation, determining its cause(s) in cancer stem/progenitor cells is critical for understanding the etiology of cancer. Several recent reports support the concept that defects in telomere maintenance initiate genomic instability eventually resulting in the development of breast cancer and other cancers. While the general concept of defective telomere maintenance initiating genomic instability has been acknowledged, there remains a critical gap in the understanding of the extent of telomere dysfunction in human cancers due to limitations in detecting telomere dysfunction within tissue. This proposal addresses this critical problem by focusing on an innovative method for detecting and analyzing telomere dysfunction in order to elucidate mechanisms responsible for the origin of genomic instability in breast cancer stem/progenitor populations leading to cancer.
Hypothesis: The central hypothesis of this research is that telomere dysfunction is one of the key driving forces behind the genomic instability observed in early cancer lesions. The hypothesis of this proposal is based on several reports and on our recent findings that: 1) telomere dysfunction is present in over 90% of breast tumors, but absent in normal tissue and 2) telomere fusion junctions contain relatively short regions of non-telomeric DNA from known highly recombinagenic regions such as fragile sites and subtelomeric regions. In light of our findings and the findings of others, we propose the following mechanism for the destabilizing effect of telomere dysfunction: loss of telomere function occurs initially in a breast cancer stem cell(s) resulting in end-to-end chromosome fusions, causing genomic instability via breakage-fusion-bridge cycles during subsequent cell cycles. Consequently, loss of genomic integrity leads to misregulation of genes involved in growth control and other critical functions, such as regulation of telomerase activity, ultimately resulting in tumorigenesis . These telomere fusions are thus early indicators of tumorigenesis, and we have developed an innovative PCR-based assay to detect and analyze these fusions in human cell lines and tissue.
The objective of this proposal is to study the role of telomere maintenance and the extent of telomere dysfunction in cancer stem/progenitor cells during tumorigenesis. Although the basic theory of a stem cell origin for breast cancer is supported by several lines of evidence, this basic concept does not account for the following seemingly paradoxical observations of telomere maintenance and telomerase reactivation during tumorigenesis: 1) stem and progenitor cells generally have relatively long telomere lengths though tumors have relatively short telomere lengths; 2) telomerase is moderately active in stem and progenitor cells but telomerase is highly active in tumors. These questions will also be directly addressed by experiments in this proposal. These experiments are expected to help resolve a longstanding question of the tumor stem-cell theory: whether tumor precursor cells are derived from transformed stem cells themselves or from more differentiated progeny that then attain stem-cell like properties during carcinogenesis .
Specific Aim: Determine the extent of telomere dysfunction in breast cancer stem/progenitor cells.
Study Design: We have developed a PCR-based method to detect and analyze chromosome fusions from isolated genomic DNA. Additionally, we have successfully isolated several subpopulations of primary breast cancer cells including cancer stem cells by flow cytometry and have used them for several biochemical assays. Stem/progenitor cell populations from both normal and tumor tissue will be compared in regard to telomere maintenance parameters such as the extent of telomere dysfunction, telomere length, and telomerase activity. Combining innovative developments between our two labs will allow us to determine the extent of telomere dysfunction in breast cancer stem/progenitor cells. This is a valuable collaboration between Dr. David Gilley with expertise in telomere maintenance and Dr. Harikrishna Nakshatri with expertise in breast cancer stem/progenitor cells.
Impact: Given the major role of cancer stem/progenitor cells in the development of cancer, it is important to increase our understanding of telomere structure, function, and maintenance in this critical cell population. This proposed research is therefore highly significant not only because of its impact on the fields of both telomere and stem/progenitor cell cancer biology, but also because of its translational applications for early detection and treatment of cancer.
Background: One of the earliest changes occurring in the development of breast cancer is a disturbance or rearrangement of the DNA in a small subset of normal breast cells. This rearrangement of the genetic material in these cells causes normal cell growth to become uncontrolled potentially resulting in development of a cancerous tumor. A key component necessary for maintaining stability of the DNA and thus preventing uncontrolled cell growth is a structure called the telomere found at the very ends of chromosomes. These telomeres serve as protective caps preventing damaging events such as fusion of the end of one chromosome with the end of another. Recently there has been evidence demonstrating that when the normally protective telomeres become dysfunctional, a series of events takes place that leads to development of breast cancer. Our theory is that loss of function of these protective telomeres results in fusions of the ends of chromosomes and may be a key driving force in the development of cancer. We already have evidence that breast tumors do contain these telomere fusions and will now do further research on their precise role in development of breast tumors by studies involving cancer stem/progenitor cells. The cancer stem cell theory proposes that cancers arise from malignant transformation of normal stem or progenitor cells. The inherent properties of normal stem/progenitor cells may impart their transformed counterparts with the ability to form invasive tumors, evade traditional anti-tumor therapies, and establish metastasis. By doing this research, we hope to gain a deeper understanding of how this process of tumor formation in breast tissue begins in breast cancer stem/progenitor cells so that we can apply this knowledge to early detection of and treatment strategies for breast cancer.
Hypothesis: Our objective is to determine how breast cancer begins. Loss of stability in the DNA of the cell occurs very early in development of cancer. We now want to investigate what is responsible for these early adverse changes and rearrangements. Specifically, we are testing our hypothesis that the telomeres, which normally serve as protective caps at the ends of the chromosomes in the cell, become dysfunctional, leaving the ends of the chromosomes unprotected and eventually leading to breast cancer development. When a chromosome end is left uncapped in this way, it can then fuse with other uncapped or broken chromosomes. This may result in breakage and further damage of chromosomes or errors in cell divisions producing new cells with the wrong number of chromosomes. These changes lead to instability of the genetic material in the cell and ultimately to the development of breast cancer.
Study Design: We have developed an innovative method to detect and analyze these telomere fusions. Using this method, we have recently gained information that provides critical clues about the mechanism involved in formation of the telomere fusions and have also confirmed that they are found in the cells of breast tumors. We will now be able to find out exactly how early these fusions occur in the development of breast cancer by testing breast cancer stem/progenitor cells. This is a collaboration between Dr. David Gilley, with expertise in telomere maintenance, and Dr. Harikrishna Nakshatri, with expertise in breast cancer stem/progenitor cells.
Potential Outcomes and Benefits of the Research: Based on our preliminary findings, we expect that dysfunction of the telomeres and the resulting fusion of chromosome ends is a very early event in development of breast cancer. Development of a method that allows identification of these fusions is therefore anticipated to be a valuable means of detecting very early forms of breast cancer.
We anticipate that as a result of this research it will be possible within the next decade to provide a simple, accessible clinical test for very early detection of breast cancer. The ability to use this new test to detect breast cancer at a significantly earlier stage than is currently possible using the previously available diagnostic tools, would prove immensely valuable as a means of identifying pre-symptomatic at-risk individuals and thus especially useful for prevention and treatment strategies for breast cancer.