Biology of the Breast Cell
To understand the origin of breast cancers, more research is needed on the architecture, cell interactions, and molecular pathways in the normal breast. Understanding how cells coordinate migration, maturation, proliferation, and cell death over space and time gives us the foundation from which to learn what it is that makes a tumor cell. The CBCRP funded studies that model normal pre-cancer and tumor breast to learn how cancer develops, and moves to other parts of the body. Important basic science topics represented in CBCRP’s portfolio include: exploring the role of stem cells in normal and tumor breast; cell proliferation control mechanisms through the estrogen receptor and growth factor receptors (e.g., Her-2); alterations in DNA repair processes that permit genetic damage to accumulate in cancer cells; cell cycle changes that permit division under conditions where normal cells would undergo programmed cell death (apoptosis); novel biomarkers to distinguish pre-cancerous and cancerous cells from normal breast epithelium and their validation as potential new detection and therapy targets, and developing methods for accounting for the complexity of the interplay of all of these factors in breast cancer.
Two research topics are presented in this section.
- Biology of the Normal Breast: The Starting Point
- Pathogenesis: Understanding the Disease
Research Conclusions
Normal Mammary Biology of Phosphorylated
Prolactin
The hormone prolactin has two major forms, an
unmodified form that promotes cell proliferation
and a phosphorylated form that inhibits cell proliferation.
Ameae Walker, Ph.D., at the University
of California, Riverside, and colleagues explored
the effect of both types of prolactin on the
breast. Ms. Walker and her team demonstrated
that prolactin turns into the phosphorylated form
that inhibits cell proliferation when the mammary
gland. They also showed that unmodified prolactin
makes changes in the cells that favor proliferation,
whereas phosphorylated prolactin makes
changes that reduce cell proliferation and, under
some circumstances, lead to cell death. These
findings suggest that phosphorylated prolactin is
beneficial to breast health, and may help explain
why breastfeeding reduces breast cancer risk.
While conducting this research, Dr. Walker identified
a new molecule inside breast cells. She also
found that the ratio of this molecule to another
molecule is associated with the absence or presence
of breast cancer, and that breast cells grow
faster when exposed to more of this molecule.
This work could lead to new methods of assessing
breast cancer risk that involve measuring
prolactin levels. It also could lead to the development
of new treatments that use phosphorylated
prolactin (or a molecular mimic of it) to prevent or
treat breast cancer.
Axon Guidance Proteins in Mammary Gland Development
The Slits are a protein family found in many
organs, including the breast. Some studies have
suggested that Slits are a tumor suppressor gene
that can stop cancer cells from growing and
spreading, but others have found that the Slit
gene does not function in breast cancer cells.
Using a mouse model, Lindsay Hinck, Ph.D., at
the University of California, Santa Cruz, and colleagues
showed that the Slit gene stops functioning
in breast cells that have increased levels
of the protein Cxcr4 and a molecule related to
it called Sdfe1. They also discovered that high
levels of Slit are correlated with lower levels of
Cxcr4 and decreased tumor growth, and that
there is a similar inverse correlation between Slit
and Cxcr4 expression in human breast tumor
tissue. These findings support previous research
that has demonstrated that Cxcr4 and Sdf1
play a pivotal role in breast cancer growth and
metastasis. It also suggests that Slit may be a
marker of whether a cancer cell has the potential to become invasive. Dr. Hinck received a grant
from the National Cancer Institute that will allow
her to further investigate how Slit functions. This
work could lead to the development of new treatment
strategies to prevent invasive breast cancer.
Findings from this research were published in Development 2006(133)823 and Cancer Research 2008(68)7919.
A Candidate Marker of Mammary Tumor Initiating
Cells
Researchers have shown that only a small number
of breast cancer cells are able to produce tumors
when they are transplanted into an animal
model. These cells, called cancer stem cells, may
be good targets for drug treatments. However,
no one has yet identified a functional marker on
these cells. Alexey Terskikh, Ph.D., at The Burnham
Institute for Medical Research, La Jolla, and
colleagues investigated whether a newly discovered
gene, called MELK (maternal embryonic leucine-
zipper kinase), might be a functional marker
for breast cancer stem cells. Studies have shown
that MELK is turned on in a number of different
cancer cell lines, but the exact role it plays is not
known. This project allowed Dr. Terskikh and his
team to complete the animal breeding necessary
to develop mice with the proper genetic structure
needed for their experiments. The studies they
conducted with these mice found that MELK
appears to be a marker for breast cancer stem
cells. This work suggests that the small molecule
inhibitors of MELK that Dr. Terskikh’s colleagues
at the Burnham Institute for Medical Research
are developing may make effective breast cancer
treatments.
A New Marker for Mammary Epithelial Stem
Cells?
Scientists believe that it is the breast epithelial
stem cells that give the breast the ability to grow
and start making milk after each pregnancy.
Robert Oshima, Ph.D., at The Burnham Institute
for Medical Research, La Jolla, discovered a new
marker gene, called maternal embryonic leucinezipper
kinase (Melk), on several types of stem
cells. This research project allowed Dr. Oshima
to explore in both cell and animal models whether
Melk is also present in breast epithelial stem cells.
Dr. Oshima and his team found that the dividing
cells that contribute to the interior lining of the
breast ducts are the breast cells that express the
most Melk protein. But even though these cells
increase rapidly, they do not have same ability
that stem cells do to generate a new mammary
gland. Dr. Oshima is continuing to explore the
relationship between Melk-producing cells and
cancer stem cells.
The Role of the ECM in Breast Cancer DNA Damage
Repair
The extracellular matrix (ECM) provides structural
support to cells. It also gives chemical cues
that can stop cells from becoming cancerous.
Albert Davalos, Ph.D., at the Lawrence Berkeley
National Laboratory, used 3-D breast cell and
animal models to investigate the role the ECM’s
basement membrane plays in breast cancer
progression. Dr. Davalos and his team found that
exposing epithelial cells that lack a BRCA1 gene
to drugs that disrupt cell replication causes them
to develop a mutation in a key tumor suppressor
protein called p53. In addition, they grow more
rapidly and fail to die. Their research also showed
that exposing cells that are missing the BRCA1
gene and the p53 protein to drugs that disrupt
cell replication causes them to fail to die and to
divide with numerous DNA breaks. Dr. Davalos
and his team observed the same result when
they turned off the special proteins in cells that
“unwind” double-stranded DNA for replications
and repair processes. These findings suggest that
loss of a “caretaker” and “gatekeeper” protein,
like p53, allows breast epithelial cells to evade
cell death and divide with more DNA damage.
While doing this work, Dr. Davalos and his team
discovered that a protein called HMGB1 is secreted
when other repair proteins are missing. Dr.
Davalos and his team are now exploring whether
HMBG1 is as an early biomarker of genetic instability
in breast cancer. Findings from this research
were published in Cell 2007(128)97.
Stem Cells of Molecularly Diverse ER-negative
Breast Cancers
Cancer stem cells comprise only a small fraction
of a tumor, but they play a critical role in tumor
growth. In fact, 100 cancer stem cells implanted
into a mouse can reproduce a large breast tumor,
whereas 20,000 malignant epithelial cells will not
generate a breast tumor at all. Stefanie Jeffrey,
M.D., at Stanford University, Palo Alto, used a mouse implanted with human tissue to investigate
whether two subtypes of estrogen receptor
negative breast cancer have different cancer
stem cell populations and to explore whether
cancer stem cells are the same as circulating
tumor cells. Dr. Jeffrey and her team found that
circulating tumor cell gene expression could vary
in a single human blood sample. They also found
that, in some instances, the circulating tumor
cells were similar to those seen in the primary tumor,
whereas in other instances they were similar
to the cells in the biopsy taken from the metastases.
These findings advance our understanding
of the cancer stem cells and circulating tumor
cells and could help lead to the development of
treatments targeted at specific types of tumor
cells. Findings from this research were published
in BMC Genomics 2006(7)231, Bioinformatics
2007(23) 957, Breast Cancer Research 9(2007)
R30, Molecular Biology 2007(8)118, Oncogene 2007(26)6269, and Radiology 2008(246)367.
A Novel Epithelial-Stromal Model of Metastatic
Breast Cancer
Identifying the genes that directly regulate cell
physiology and architecture in the breast can help
researchers understand how breast cancer tumors
spread to other organs (metastasize). Richard
Neve, Ph.D., at the Lawrence Berkeley National
Laboratory, and colleagues used an animal
model to study the role a receptor, called EPHA2,
and its protein, EFNA1, play in breast biology.
They were interested in EPHA2 because it is seen
in a subset of breast cancers that scientists have
learned are predisposed to metastasis. Dr. Neve
and his team found that reducing the EPHA2
protein keeps the cancer cells in triple-negative
breast tumors from becoming invasive. (They are
called triple negative because they are estrogen
receptor, progesterone receptor, and Her-2 negative.)
They also found that a malignant cell will
not become invasive when it is adjacent to a cell
with EFNA1 on its surface. This suggests that
stromal cells (connective tissue cells) with EFNA1
on their surface may be able to stop breast tumors
from becoming invasive. Dr. Neve and his
team developed a screening system that mimics
the stromal cells to study cell-to-cell interactions
of EPHA2 and EFNA1 in a variety of breast tumor
cell lines. These experiments indicated that this
interaction has the potential to slow the growth
of cancer cells. These findings provide evidence
that EPHA2 plays a role in breast cancer metastasis
and could lead to the development of new
treatments for metastatic breast cancer.
MYC and CSN5 in the Breast Cancer “Wound
Signature” Profile
In normal wound healing, as in cancer growth,
there is rapid cell proliferation, cell migration, and
new blood vessel development. For this reason,
cancer is sometimes referred to as “wounds
that do not heal.” Adam Adler, B.A., at Stanford
University, Palo Alto, and colleagues previously
found that when two genes, called CSN5
and MYC, are turned on, they induce a genetic
process referred to as a “wound signature.”
Furthermore, when this “wound signature” is
present, a breast cancer is more likely to become
invasive. To investigate these findings, Mr. Adler
and his team developed human and mouse cell
models that would allow them to explore the role
of CSN5 and MYC in promoting breast cancer
progression. Mr. Adler and this team found that
when CSN5 or MYC is turned off in this model,
cancer does not progress. This means that both
genes are necessary for cancer to develop. Additional
animal model studies confirmed that
CSN5 is required for MYC-induced breast cancer
growth. These findings show that MYC and
CSN5 play a critical role in regulating breast
cancer progression, and they could lead to the
development of new breast cancer treatments
that target CSN5. Results of this research were
published in PLoS Genetics 2007(3)91e and in
Cancer Research 2008(68)369 and 506.
Role of Cell Division Asymmetry in Breast Cancer
Stem Cells
Breast cancers contain a small population of
breast cancer stem cells that appear to be more
resistant to existing treatments than other tumor
cells. Claudia Petritsch, Ph.D., at the University
of California, San Francisco, attempted to discover
the very first changes normal stem cells undergo
when they turn into breast cancer stem cells.
Dr. Petritsch and her team began by developing a
cell-based test to analyze the rate and nature of
asymmetric cell division in mouse mammary stem
cells. This test allowed them to show that normal
breast stem cells undergoing asymmetric cell division
generate another stem cell and a differentiating
cell. They also showed that breast stem cells that do not perform this asymmetric cell division
properly generate too many breast cancer stem
cells. Dr. Petritsch is now exploring what occurs
when she disrupts asymmetric cell division by
taking out the gene Lgl-1, which regulates asymmetric
cell division in the developing breast. She
is studying Lgl-1 because the human equivalent
of this gene, called Hugl-1, is missing in 76% of
breast cancers. Dr. Petritsch and her team also
intend to investigate how Lgl-1 prevents cancer
from developing by preserving normal asymmetric
cell divisions. This work could lead to new treatments
that more specifically target breast cancer
stem cells and could lead to the development of
tools for the early diagnosis of breast cancer.
Role of Integrins in Lymphangiogenesis During
Breast Cancer
Breast cancer spreads predominantly through
lymphatic vessels and lymph nodes. The lymphatic
vessels that surround breast tissue consist of a
single layer of cells, called lymphatic endothelium.
Barbara Susini, Ph.D., at the University of California,
San Diego, previously found that the number
of lymphatic vessels in breast tissue increases
dramatically during breast tumor development, a
process called lymphangiogenesis. Now, she is
exploring the mechanisms that drive this increase
in lymphatic endothelial cells or promote breast
tumor cell invasion of the lymphatic vessels. Dr.
Susini and her team found that growing lymphatic
vessels and cells express only one protein;
it is called alpha4/beta1, and it works with a
molecule called VCAM in the metastatic process.
They also found that the CCL21 protein and its
receptor, called CCR7, help tumor cells get to the
lymph nodes by making it easier for the tumor
cells to attach to the lymphatic endothelium.
Furthermore, they were able to identify which
molecules interact with alpha4/beta1 to induce
lymphatic endothelial cell migration. In addition,
they discovered that tumor cells cause lymphatic
vessels to grow both in the tumor and in the
lymph nodes. This research advances our understanding
of breast cancer metastasis and could
lead to the development of new breast cancer
treatments. Three papers were published on this
research, including a summary in Nature Reviews
Cancer 2008(8)604-17.
Imaging RhoC-induced Breast Cancer Invasion
and Angiogenesis
Metastasis—the spread of cancer cells to other
parts of the body—is the major cause of death
in breast cancer patients. Metastasis is a highly
dynamic process that occurs in several distinct
steps. Konstantin Stoletov, Ph.D., at the Scripps
Research Institute, La Jolla, and colleagues grew
human cancer cells that contained a metastatic
gene, called RhoC, in optically clear Zebrafish
so that they could directly observe how tumors
grow, invade, and develop new blood vessels, a
process called angiogenesis. Dr. Stoletov and
his team found that a gene, called RhoC, causes
the tumor cell to develop specific features that
allow it to penetrate the blood vessel. They also
found that tumor cells only penetrate the blood
vessels in places where new vessels are currently
developing, and that continuous secretion of the
growth factor called VEGF is necessary to create
an opening in the blood vessel for the cancer cell
to pass through. These findings could lead to the
development of new drug treatments that target
these processes. Dr. Stoletov and his team are
continuing to investigate how tumor cells and
blood vessels interact during metastasis. Three
papers were published on this research, including
a summary in Oncogene 2008(8)604-17.
Identifying Metastatic Breast Cells from Peripheral
Blood
Surgeons examine the lymph nodes of breast
cancer patients to assess whether metastases
has occurred. But this method is not perfect, and
new approaches are needed. Studies have shown
that tumors shed cancer cells into the blood
when they become invasive. Kristen Kulp, Ph.D.,
at the Lawrence Livermore National Laboratory,
and colleagues are attempting to develop a blood
test that could determine whether circulating
tumor cells are present in the blood and, in turn,
whether metastases has occurred. Dr. Kulp and
her team identified a way to prepare cells for this
type of analysis. However, the methods currently
available to isolate circulating tumor cells
are not able to detect as few as 10 cells in 15
milliliters of human blood, which is what would
be necessary to identify metastases. As a result,
they were not able to implement this new technique.
Dr. Kulp and her team intend to monitor
the development of new methods for cell isolation and will continue to attempt to develop a blood
test for breast cancer metastasis. Three publications
resulted from this funding, including Analytical
Chemistry 2006(78)3651-8 and Journal
of the American Society of Mass Spectrometry 2008(19)1230-6.
The Role of Serine and Metallo-hydrolases in
Breast Cancer
Extracellular and cell-surface enzymes (a type
of protein made by cells) from the serine and
metallo-hydrolase family are believed to play a
role in breast cancer metastases. Sherry Niessen,
M.S., at Scripps Research Institute, La Jolla, and
colleagues used the most advanced techniques
available to identify and characterize novel serine
and metallo-hydrolase enzymes that play a role in
breast cancer biology. Ms. Niessen and her team
found that a serine hydrolase called KIAA1363
was increased in tumors and aggressive cell
lines. Additional studies showed that KIAA1363
regulated levels of a family of lipids known as
monoalkylglycerol ethers (MAGEs); had an impact
on a larger lipid signaling network that included
lysophosphatidylcholine (alkyl¬LPC) and lysophosphatidic
acid (alkyl-LPA); and suggested that
KIAA1363 has an effect on these lipids. Ms. Niessen
and her team were able to define an aggressive
gene signature regulated by KIAA1363. This
signature included a protein called Fra-1, which
they demonstrated is regulated by both alkyl-
LPC and alkyl-LPA. These findings indicate that
KIAA1363 is an important molecule in human
cancer biology, and contribute to our understanding
of the role enzymes play in breast cancer
progression.
Twist Activation in Breast Cancer Metastasis
Metastasis occurs when tumor cells spread from
a primary site to distant organs and establish secondary
tumors. During metastasis, tumor cells obtain
the ability to break away from their neighbor
cells and migrate. Jing Yang, Ph.D., at the University
of California, San Diego, previously showed
that tumor cells activate a gene called Twist to
begin this process. She is now using a mouse
model to investigate how Twist gets breast tumor
cells to spread to distant organs. Dr. Yang found
that turning the Twist gene “on” alters the form
and structure of the breast. She also found that
turning on Twist is sufficient to get certain human
breast cancer cells to spread to distant organs,
such as the lung. Using human tumor cells, Dr.
Yang and her team demonstrated that Twist appears
to facilitate metastasis. However, continued
Twist expression appears to inhibit proliferation
at metastatic sites, like the liver and lung. Dr.
Yang and her team have generated new mouse
models that will allow them to learn more about
the impact Twist has on breast tissue. This work
could establish Twist as an important prognostic
marker. It could also lead to the development
of new drug treatments for metastatic breast
cancers.
Identification of Metastasis Competent Breast
Cancer Cells
It currently is not possible to diagnose the
earliest stages of metastasis. As a result, many
women undergo chemotherapy and radiation to
kill metastatic cells, even though it’s not known
whether they are present. These post surgical
treatments undoubtedly save lives, but they have
no medical benefit if the cancer has not spread.
Barbara Mueller, Ph.D., at the La Jolla Institute
for Molecular Medicine, is developing tools that
can measure a cancer cell’s ability to cause
metastasis before metastasis actually occurs. Dr.
Mueller and her team have identified four specific
molecules that, when present, appear to indicate
that a breast cancer cell has the capability to metastasize.
Dr. Mueller is currently seeking funding
from the National Institutes of Health to conduct
the additional studies necessary to validate these
findings. The ability to identify cells with metastatic
potential could result in more effective use
of existing treatment options. It could also lead
to the development of new treatments for early
stage metastatic disease.
Modeling, Targeting Acetyl-CoA Metabolism in
Breast Cancer
Cancer cells differ from normal cells in that they
grow uncontrollably, require increased energy,
and withstand low pH and low oxygen conditions.
In addition, cancer cells use glucose as an energy
source in ways that normal cells do not. Chen
Yang, Ph.D., at The Burnham Institute for Medical
Research, La Jolla, studied how breast cancer
cells metabolize glucose in an attempt to develop
an anticancer drug that would interrupt this
process. By comparing normal and breast cancer cells, Dr. Yang was able to pinpoint tumor-specific
metabolism and characterize the metabolic
changes that occur during cancer development.
He was also able to select a set of prospective
drug targets. Dr. Yang is continuing to study
the genetic patterns in breast cell metabolism to
determine the best ways to target this process.
The research was published in Breast Cancer
Research and Treatment 2008(100)297-307 and
Metabolomics 2008(4)13-29.
The Role of Estrogen-Related Receptors in Breast
Cancer
The small family of estrogen-related receptors
consists of three proteins that control the expression
of many genes important in maintaining
normal cell growth. The three estrogen-related
receptors are similar to the estrogen receptors,
but they are not activated by natural estrogens.
This similarity has led researchers to hypothesize
that estrogen-related receptors, like estrogen receptors,
play a role in breast cancer development
or growth. Anastasia Kralli, Ph.D., at the Scripps
Research Institute, La Jolla, used human breast
cancer cells to study estrogen-related receptors
and the role they play in breast cancer growth,
metastasis, and response to drugs. Dr. Kralli and
her team found that cells with higher levels of
estrogen-related receptor activity responded as
expected to chemotherapy drugs in cell culture
studies. However, these cells were not able to
grow and develop when transplanted into the
breast area of mice. These findings demonstrate
that certain changes in estrogen-related receptor
activity appear to keep breast cancer tumors
from growing in animal models. This work could
lead to the development of new treatments that
use estrogen-related receptor molecules to slow
breast cancer growth.
The Role of LMO4 in Breast Cancer
Cancer cells have acquired genetic mutations that
give them the ability to grow and divide uncontrollably.
Zhengquan Yu, Ph.D., at the University
of California, Irvine, and colleagues investigated
whether a protein called LMO4, which is found
in breast epithelial cells (the cells in which breast
cancer begins), helps to regulate cell proliferation
and cell death. Dr. Yu and his team also explored
whether cells that have too much of this protein
begin to grow and divide uncontrollably. Using
a mouse model, Dr. Yu and his team showed
that mammary epithelial cells that lack an LMO4
gene are less likely to divide and more likely to
die. While conducting these studies, the research
team found that another gene, called BMP7, is
regulated by LMO4 in breast cancer cells. Dr. Yu
and his team intend to continue to study the
role of BMP7 in mammary gland development
and breast cancer. This work could advance our
understanding of how breast cancer develops. Results
from this research were published in Oncogene 2007(26)6431-41.
Grants in Progress: 2008 Breast Cancer Studies in a 3-D Cell Culture System Breast Tumor Responses to Novel TGF-beta Inhibitors Competition for ADA2 and 3 to Inhibit p53 in
Breast Cancer Cytoskeletal Regulation of Invading Breast Cells Defining Mammary Cancer Origins in a Mouse
Model of DCIS Determination of Stromal Gene Expression in
Breast Cancer Functional Analysis of BORIS, A Novel DNA-binding
Protein Indole (I3C) Control of Breast Cancer by ER
Downregulation Inflammation Alters Transcription by ER in Breast
Cancer Lipid Raft Composition in Deregulated ERBB2
Signaling Mechanisms of Daxx-mediated Apoptosis in
Breast Cancer A New Mouse Model of PI3-kinase Induced
Breast Cancer Novel Approach to Analyze Estrogen Action in
Breast Cancer Novel Regulation of the Rb Pathway in Breast
Epithelium Profiling Drug Metabolism (P450) Proteins in
Breast Cancer Reactivation of the Inactive X Chromosome and
Breast Cancer Regulation of Mammary Epithelial Invasion by
MMPs and FGFs |
The Relationship of BRCA1 and HMGA2 in Breast
Cancer The Role Chk1 in Breast Cancer DNA Damage
Repair The Role of Podosomes in Breast Cancer Metastasis Stem Cells in Breast Cancer Metastasis Structural Analysis of Cancer-relevant BCRA2
Mutations Targeting Tissue Factor in Breast Cancer Telomerase, Mammary Stem Cells, and Breast
Cancer Trask, a Candidate Breast Cancer Metastasis
Protein Research Initiated in 2008 Chemokine Receptor Signaling in Breast Cancer Dietary Metabolite Inhibition of Breast Cancer Cell
Survival Dissecting the Role of Twist in Breast Cancer
Metastasis Global Analysis of Protein Ubiquitination in Breast
Cancer Maternal Embryonic Leucine Zipper Kinase in
Mammary Tumors Nanolipoproteins to Study Breast Cancer Growth
Receptors Regulation of Breast Stem-progenitor Cell Chromatin
by Pygo2 Role of Estrogen-modulated Protein AGR2 in
Breast Cancer Tumor Suppressor 14-3-3sigma in Breast Cancer
Progression |
