Biology of the Normal Breast: The Starting Point
As any woman who performs her monthly breast self-examinations knows, the normal breast is a constantly changing organ. The breast's normal changes can obscure the more ominous changes associated with cancer. Researchers have worked hard to determine what constitutes a cancerous change in the breast, but the lack of a thorough understanding of the normal breast makes this work more difficult. Because a relatively small amount of research is being done in this area, the California Breast Cancer Research Program earmarks funds especially for it. In 2001, we funded researchers who are studying the development, structure, hormonal regulation, and genetic control of the normal breast. Our hope is that these studies will provide a strong foundation for distinguishing the difference between benign and malignant breast changes.
A Novel Gene Associated with an Altered Risk of Breast Cancer.
A full-term pregnancy reduces a young woman's lifetime risk of breast cancer by about 30%. Satyabrata Nandi, Ph.D. from the University of California, Berkeley, investigated the hypothesis that pregnancy leads to changes in genes that are responsible for reducing the risk. Dr. Nandi's previous research showed that pregnancy will protect rats from substances used to cause mammary tumors (the rat equivalent of human breast cancers) in laboratory experiments. Using a special cloning method, his lab identified a gene called Rat Mammary Tumor-1 (RMT-1). This gene is present mainly in the mammary gland, at higher levels in breast cancer, but is less evident in older rats and in rats that have completed a full-term pregnancy. A higher level of activity of the gene seems to be associated with sensitivity to mammary cancer induced in the lab with a cancer-causing substance. The presence of RMT-1 appears also to be associated with higher levels of estrogen receptors, proteins within breast cells that bind with the hormone estrogen. This research was published in Cancer Letters 174:45-55 (2001).
Regulation of Breast Epithelial Cell Motility by Proteases
Vito Quaranta, M.D. of The Scripps Research Institute, La Jolla, investigated molecule-level interactions that underlie the ability of cells to move to a new location within the body. In the human breast, epithelial cells—the cells where most cancer arises—are surrounded by a complex structure containing support components and other types of cells. The structural components of the breast include proteins called laminin. To move around and reorganize when the breast goes through normal changes—such as the start of milk production—the epithelial cells need to digest the structural proteins surrounding them. Cancerous cells do the same thing when they migrate to other parts of the body. Dr. Quaranta found that an enzyme produced by epithelial cells, MT1-MMP, digests a laminin protein, Ln-5, by splitting it in two places. The resulting fragments may have a function. Another enzyme, MMP2, may increase the efficiency of cell migration. In mice that genetically lacked MT1-MMP, splitting of the Ln-5 molecule was lacking or greatly reduced. Their tissues also showed abnormal organization of epithelial cells. The results of this study may point to ways to block cell migration as a way to limit or prevent breast cancer cells from spreading to other parts of the body. Publications resulting from this research appeared in Journal of Cell Biology 2000 148(3):615-24, 2000 149(6):1167-70, 2001 153(3):465-78; and Cell Adhesion and Communications, 2001 8:29-44.
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Epithelial Cells |
Hox Genes in Normal Breast Development and Breast Cancer
The loss of control of cell growth and cell specialization is a hallmark of cancer. Understanding genes that control these processes can lead to new candidates for anti-cancer drug development. Carmen Hagios, Ph.D., at the Lawrence Berkeley National Laboratory, studied the role of Hox genes in breast cancer, because these genes regulate cell growth and specialization in the developing embryo. Breast cancer cells also have abnormally high amounts of the proteins these genes produce. Dr. Hagios designed cell cultures that reproduced the conditions in the body, where epithelial cells (the source of most cancers) are surrounded by a collagen-based support structure (the basement membrane). The team found that levels of the Hoxa-1 protein produced by the Hoxa-1 gene were higher in two mouse mammary tumor cell lines (Scg6 and TC-1) compared to normal mouse mammary epithelial cells. The tumor cells formed irregular structures, while the normal cells formed organized spheres. When the team experimentally decreased the level of Hoxa-1 protein in the tumor cells, the cells adopted more organized cell structures, similar to those of normal cells.
Identification of Novel Id-1 Regulated Genes in Breast Cells
Id-1 is a gene that plays a role in the normal growth and development of breast epithelial cells, the cells where most cancer arises. The gene also helps epithelial cells continue to grow, which is one of the characteristics of cancer cells, and triggers the process where cells migrate to other parts of the body. Jarnail Singh, Ph.D., at the California Pacific Medical Center, San Francisco, investigated genes involved in the normal growth and functioning of the breast that Id-1 turns on or off. The research team found that Id-1 causes another gene to produce more of a protein called clusterin. Clusterin tends to be high in cells that are producing milk in a laboratory culture and in mice. Clusterin may play a role in the process epithelial cells go through to produce milk. The team also found five more genes that Id-1 turns on or off. One is involved in cell death and another is turned on in breast cancer cells that have the ability to spread to other body parts.
Research in Progress
Breast Development
Hormonal Regulation of TGF-beta1 During Mammary Development
The reproductive organs of female mice go through the estrus cycle, where fertile periods, when pregnancy is possible, alternate with infertile periods, when it isn't. Levels of the hormones estrogen and progesterone rise and fall in a pattern during the mouse estrus cycle, as they do during the human menstrual cycle. Mary Helen Barcellos-Hoff, Ph.D., at the Lawrence Berkeley National Laboratory, is investigating how changes in hormone levels interact with a protein, transforming growth factor beta1 (TGFbeta1) found in a small proportion of breast cells. Dr. Barcellos-Hoff's hypothesis is that the hormones estrogen and progesterone cause some cells to produce TGF-beta1, and that TGF-beta1, in turn, plays a role in the growth of new cells and replacement of worn-out cells during normal breast development. The team has shown that when mouse mammary cells (the equivalent of breast cells in humans) produce more TGFbeta1, they multiply more slowly. However, during puberty, the fertile periods, and early pregnancy, cells that produce more TGF-beta1 multiply more quickly. TGF-beta1 also stimulates the normal process of cell death, but only during the mouse fertile period. During the coming year, the team intends to identify more characteristics of TGF-beta1. Discovering the function of TGF-beta1 in the normal breast can lead to defining its role in breast cancer.
Other Processes in Breast Biology
Method for Measuring Breast Epithelial Turnover in Humans
Epithelial cells in the breast produce milk and deliver it to the nipple, and are also the source of most breast cancers. Breast cancer cells divide more rapidly than normal cells. Each time a normal cell divides, the chance of a genetic mutation goes up, and so does the risk that the mutation will lead to cancer. Therefore, it is important to have a reliable way to measure the division rate of cells in the breast. Marc Hellerstein, M.D., Ph.D., at the University of California, Berkeley, is investigating further a technique developed in his laboratory to measure cell division rates directly, without using radioactivity or toxic substances. His team has used the technique successfully to measure the cell division rate in women using breast tissues from core biopsies. They have also found that genistein, a substance found in soybeans, decreases the cell division rate in rats. They are working on establishing normal rates of breast cell division and factors that might be associated with variations of this epithelial turnover rate in women, such as age, weight, ethnicity, and diet.
Genetic Changes in Normal Epithelium of the Cancerous Breast
Shanaz Dairkee, Ph.D., of the California Pacific Medical Center Research Institute, San Francisco, is investigating genetic changes that occur in normal-appearing breast cells. The goal is to identify changes that indicate a propensity to become breast cancer. The team is looking at four genes-p53, ATM, BRCA1, and BRCA2. The normal version of all these genes suppresses tumors. Women can inherit a mutation on each of these genes that makes them more likely to get breast cancer. Many researchers believe that mutations in these genes play a role in non-inherited breast cancer, and that something other than an inherited mutation turns off these genes. One common way genes get turned off is by losing an actual part of the gene when the cell reproduces itself. This process is called loss of heterozygosity (LOH). Dr. Dairkee's team analyzed 40 tissue samples of ductal carcinoma in situ (DCIS, a pre-cancerous breast condition that can develop into cancer). The team found LOH at p53 in more than 50% of the samples. For 11 of these samples, they examined normallooking adjacent breast tissue and found LOH in 2. This suggests that mutations in p53, which are commonly found in later stages (III and IV) of non-inherited breast cancer, begin with partial loss of the gene at a much earlier stage. The team found LOH at the BRCA1 gene in 29% of the samples and at the ATM gene in 11%. They could not detect any LOH at these two genes in the nearby normal-appearing tissue. During the coming year, the team will analyze a spectrum of breast cells ranging from normal to invasively cancerous to establish at what point during the course of breast cancer these four genes get inactivated.
A Vascular Restriction of Mammary Tumor Progression
The mouse mammary gland is equivalent to the breast in humans. Robert G. Oshima, Ph.D., at The Burnham Institute, La Jolla, is developing mice with genes altered so they are prone to mammary gland tumors and also so they have higher levels of two substances in the mammary gland cells where most tumors arise. The two substances are VEGF and Ang1, proteins called growth factors that are normally found in cells. VEGF and Ang1 normally stimulate the growth of blood vessels in situations such as a wound. Dr. Oshima will investigate whether mice with higher levels of these two growth factors in their mammary glands produce new tumor blood vessels more quickly, and have a faster rate of tumor formation. The team hopes to discover whether the process of forming new blood vessels is the crucial step that determines how big a tumor can grow.
Research Initiated in 2001
Breast Development
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Telomere Dynamics During Breast Development. Telomeres are composed of DNA and proteins located at the end of human chromosomes. They play a role in normal breast development and are also linked to both aging and cancer. Sahn-Ho Kim, Ph.D., of the Lawrence Berkeley National Laboratory, is investigating the role of TIN2, a protein associated with telomeres, in normal breast development.
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Analysis of a Protease Involved in Mammary Development. Rana Zahedi, Ph.D., at the Lawrence Berkeley National Laboratory, is hunting for a protease (a type of enzyme) involved in the changes in the breast during pregnancy that lead to milk production. The goal is to shed light on cellular specialization and reorganization in the normal breast, and how these processes may go wrong in cancer.
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Genetic Aspects of Physiological Response During Lactation. Scientists believe that when tissue grows too large for its existing blood vessel network, the level of oxygen in the tissue drops. In response, a protein, HIF-1alpha, increases and activates genes that control new blood vessel growth. Randall S. Johnson, Ph.D., at the University of California, San Diego, is investigating whether the HIF-1alpha response to lowered oxygen levels contributes to mammary gland development and the production of milk in mice. The goal is to generate information that will lead to better therapies to regions of tumors with lower oxygen levels, which are often resistant to treatment.
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Role of IKKa in Mammary Gland Development. Michael Karin, Ph.D., at the University of California, San Diego, is investigating an enzyme called IKKa. Inactivating this enzyme prevents the extensive growth in breast cells that normally occurs during pregnancy. There is some evidence that the enzyme may also be involved in stimulating the growth of breast cancer cells and in protecting them from being killed by anti-cancer drugs. Inhibiting this enzyme doesn't affect other tissues or organs, so it could eventually be a target for a treatment that would have few side effects.
Other Processes in Breast Biology
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Role of Chromatin Regulator in Breast Cell Growth. To grow, normal cells and cancer cells must synthesize a large number of different proteins using information carried on their DNA. DNA in human cells is organized into a structure called chromatin. The flow of information from DNA to make cellular proteins is highly controlled at the chromatin level by a group of proteins. Hongwu Chen, Ph.D., of the University of California, San Francisco, has found that one of these proteins can increase the growth of breast cancer cells and prolong the life of normal breast cells. It is also present in elevated amounts in breast tumors. Dr. Chen will investigate how this protein controls normal breast cell life span and spurs breast tumor cell growth.
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Telomere Clustering is Lost in Mammary Epithelial Tumors. Telomeres—structures of DNA and proteins that cap the end of chromosomes—play a role in normal breast development, aging, and cancer. Paul Kaminker, Ph.D., of Lawrence Berkeley National Laboratory, is investigating whether disrupting the structure of telomeres in cells leads to the formation of breast cell tumors.
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Coactivators in Mammary Development and Tumorigenesis. Zhiyong Wang, Ph.D., at The Salk Institute for Biological Studies, La Jolla, is studying a protein, p/CIP, to see whether and how it causes cancer. The team will compare mammary gland and tumor development in mice with no p/CIP and mice with excessive p/CIP in the mammary glands.
