breast
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The Mouse as a Model for Breast Development and Breast Cancer Research
Dr. Tiffany SeagrovesLaboratory of Dr. Johnson
I. BREAST CANCER INTRODUCTION
Statistics
Risk Factors
Hormones
• The lifetime probability of being diagnosed with breast cancer for American women is 1 in 8 (NCI, SEER, 1997)
Breast Cancer Statistics
AGE IS the MOST
IMPORTANT RISK FACTOR
Median Age of Diagnosis is
Between 60-65 (NIH, 2000)
Recent Decrease in UK and USA Breast Cancer Mortality at Ages 50-69 Years
PETO et al. LANCET 355:1822, 2000
• Most common form of cancer in women, excluding skin cancer
Breast Cancer Statistics, cont.(Y-me National Breast Cancer Foundtaion; www.y-me.org)
• Leading overall cause of death between women of age 40-55
• ACS estimates that 192,000 American women will be diagnosed with breast cancer this year and approximately 46,000 women will die
• There are more than 2,000,000 breast cancer survivors in the U.S. today
• Age of menarche, first child, onset menopause
• Diet, level of exercise, obesity, alcohol consumption
• Presence of benign breast disease (DCIS)
• Exposure to radiation
• Family history and genetics (estimated 5% of total cases can be contributed to genetic factors, and 20-30% cases can be linked to a family history of breast cancer)
Factors Associated with an Increased Risk of Breast Cancer [Love et al. 1996]
Increase in Diganosis of Early-Stage Breast TumorsSince the 1980s
70% of breast cancers occur in women who have no identifiable risk factors.
This is why you are supposed to examine
yourself and have yearly exams.
Factors Associated with a Significant Decreased Risk of Breast Cancer (~ 30-50%)
• Completed Pregnancy by Age 20– Exposure to “pregnancy” hormones is protective if
happens early
• Removal of Both Ovaries by Age 35– Over time, exposure to “pregnancy” hormones
increases risk because ~50% of breast tumors are initially hormone-dependent
Paradox of Hormone Function in the Breast:
Why do the same hormones that promote normal
development of the breast (to prepare for lactation) act to
promote breast cancers later in life?
1) ESTROGEN AND PROGESTERONE (E+P): Produced by corpeus luteum of ovary first 6 wks or pregnancy, then taken over by placenta.
Together, E+P stimulates growth and development of secretory units and ducts in the gland. \
2) PROLACTIN (Prl):
Produced by anterior pituitary
Stimulates production of milk
The Role of “Pregnancy Hormones” in Breast Development and Lactation
II. ANATOMY AND HISTOLOGY OF THE
HUMAN BREAST
Whole gland
TDLU unit
Whole mount and H&E staining
Source: http://mammary.nih.gov
Slice of a Whole Human Breast
Brown area= “epithelium”
skin
Fatty tissue
The Structural Units (Terminal Ductal Lobular Units, TDLU) of
the Human Breast
Taken from Cardiif and Wellings, 1999
15-yr female
22 yr nulli-parous 30 yr nulli-parous
55 yr parous in menopause
80 yr parous
Acinar organization
Human TDLU Whole Mount
duct
acini
lobules
extralobular terminal ducts
Cardiff website
TDLU Histology: H&E
III. ANATOMY AND HISTOLOGY OF THE
MOUSE MAMMARY GLAND
Whole Mount
H&E
Comparative Histology with Human
The Terminal End Bud of the Mouse Mammary Gland
TEBs are highlyproliferative unitsthat form the ductal network
Stages of Mouse Mammary Development: Whole mounts
www.mammary.nih.gov
Duct
Alveoli
A-B. 6-P, “early”
C-D. 10-P, “mid”
E-F. 15-P, “mid-to-late”
G. Lactation
H. 4 days regression
Wellings and Cardiff, 1999
Which is mouse, which is human?(row A vs row B?)
A
B
IV. WHY THE MOUSE MAMMARY GLAND IS A
POWERFUL GENETIC TOOL
Reasons
Experiment approaches
How to make a transgenic mouse
to study development/cancer
• Histology is comparable to human
Why Use the Mouse as a Model for Breast Cancer?
• Can use genetics to manipulate the mammary glands
• Have multiple pairs of mammary glands that allow for multiple biopsies
• Can purify epithelial cells from the fat and culture them
Mice have 10 Mammary Glands
How to Biopsy a Mouse Mammary Gland
QuickTime™ and a decompressor
are needed to see this picture.
QuickTime™ and a decompressor
are needed to see this picture.
Remove endogenous epithelium
from stromal fat pad
Transplanted epithelium grows out Into fat pad in 6-8 weeks
Transplant a fragment of tissue Transplant a fragment of tissue Containing epithelium from a Containing epithelium from a donor OR inject purified cellsdonor OR inject purified cells
Mammary Gland Transplantation
How To Make a Mammary-Gland Specific Transgenic Mouse
milk protein gene minimal promoter Your Favorite cDNA
Inject into isolated mouse nucleus
Check mammary gland of female progeny for increased
expression of your gene
usually MMTV or WAP
V. WHAT DO MICE HAVE TO DO WITH BREAST
CANCER?
Differences between breast and mouse mammary tumors
How to get mice to develop tumors
• Most tumors mouse metastasize to the lung. Most human metastasize to the regional lymph nodes.
• Mouse tumors have much less fibrosis and inflammation
• *Half of human breast cancers are hormone-dependent. Most mouse tumors are hormone independent*
General Differences Between Human Breast and Mouse Mammary Tumors
• Molecular lesions causing breast cancer in human have proven to cause breast cancer in mice
• Similar morphological patterns of lesions appear in both species
• Development of cancer consistent with multi-hit kinetics
• Breast cancers in both species are metastatic
• Breast cancers may be hormone- independent
SIMILARITIES DIFFERENCES
Taken from Thompson and Cardiff
• Treat young mice (time most susceptible) with a chemical carcinogen
• Make a transgenic mouse that overexpresses a gene product that regulates growth
• Make a “knockout” mouse that deletes a gene that is a tumor suppressor
• Breed them several times and watch for spontaneous tumors (rare in mice, more common in rats)
How to Cause a Mammary Tumor in a Mouse
VI. HUMAN BREAST PATHOLOGY
Examples of Benign Diseases
Tumor Grades/Types
Hyperplasia vs Carcinoma
Benign vs. hyperplastic vs. carcinoma
Well- vs. poorly-differentiated
Nuclear morphology- uniform or not
Degree of proliferation
Ductal or lobular in origin?
How do pathologists classify and grade breast tumors?
Wheel of Prognosis
• HISTOLOGIC GRADE-higher the grade, less chance for survival
What factors predict outcome of treatment?
• SIZE-the larger the size, fewer patients survive
If <2 cm, 11% lymph-node negative patients will have recurrence in 5 yr; >5cm, 25-30% patients will relapse
• ER STATUS-loss of estrogen receptor tends to be negatively associated with outcome.In particular women with ER-negative tumors are no longer responsive to tamoxifen, a widely used adjuvant therapy
• PROLIFERATIVE RATE -low rate proliferation, increased chances of survival. This factor is also independent of other factors.
• Amplification of certain growth factors or receptors or loss of certain tumor suppressors (p53)-lead to decreased survival
Benign Breast Disease
1) Fibroadenoma-overgrowth of stromaMost common benign tumor of the breast; typically occurs in the 20s-30s
2) Cysts-fluid filled epithelium
may make breasts feel “lumpy”
Human Breast HyperplasiasHyperplasia: Any increase in cell number without cytologic
changes in cellular morphology
Atypical Hyperplasia: Any increase in cell number WITH cytologic changes
in cellular morphology, especially nuclear morphology
low power
MILD
low power
SEVERE
highpower
note different staining intensity of nuclei BUTcells still attached to each other
fine needle aspiration
DCIS (ductal carcinoma in situ): the “precursor” to breast cancer
It is in situ or “in place” because the cells are still bound by the extracellular matrix
Solid
Papillary
Cribiform
Breast Carcinomasthe tumor is invading the breast, it has broken through the matrix
Well-differentiated
still see glandularstructures resembling acini
Poorly-differentiated
mass of cells, no resemblance to acini
arrows point to nuclei withdifferent morphology. Note also cells no longer attached to each
other
Abonormal nuclei fromfine needle aspiration of carcinoma
• Human tumors tend to spread to regional lymph node first (why lymph nodes under arm also biopsied with tumor)
• Then tumors spread to small capillaries of the vascular network
• Breast tumors tend to metastasize to lung, liver, brain, bone
• It is more rare for rodent tumor models to exhibit metastasis, but when observed, are usually restricted to lung
Metastasis
Liver mets, (white spots)
VII. MOUSE MAMMARY TUMOR PATHOLOGY
Benign Disease
Hyperplastic Alveolar Nodules
Carcinomas
Comparative with Human Tumors
Mouse hyperplasia: the HAN
HANs and Atypical Hyperplasias
2) From GEM, usually transgenic mice
In contrast to non-GEM, several GEM transgenic mouse models develop tumors with similar pathologies to human breast tumors and more are described every year.
examples: TGF, neu, c-src, myc transgenic mice
Classification of Mouse Mammary Carcinomas:
1) From non-Genetically Engineered Mice (GEM)A. spontaneous tumors
B. tumors as a result of infection of with mouse mammary tumor virus (MMTV)
C. tumors from chemical carcinogen treatment
Classification by Cardiff, 2000
Examples of Mouse Carcinomas
ras myc neudifferent nuclear morphologiesfrom different genes
neu (erb-2)
ras
myc
ret-1
more solid more glandular
left; papillary carcinomaright; a protein kinase
transgene
both MMTV-inducedmouse tumors
left; DCIS, solid formright; neu transgene
left; schirrhous carcinomaright; src transgene
IX. BACK TO HORMONES:
HOW DO WE KNOW PREGNANCY CAN BE PROTECTIVE AGAINST BREAST CANCER?
A combination of E+P treatment reduces chemical carcinogen-induced tumorigenesis
in rodents (Nandi et al. 1995, 1999)
http://mammary.nih.govhttp://pathology.ucdavis.edu/tgmice/firststop.html http://www.y-me.orghttp://www.komen.orghttp://www.nci.nih.gov/THE JOURNAL OF MAMMARY GLAND BIOLOGY AND NEOPLASIA
REFERENCES