Transcript (EGF-R) IN HUMAN BREAST CANCER

By.
P.B.TIRUPATHI PICHIAH.
Dept Of Animal Sciences.
BHARATHIDASAN UNIVERSITY
An Introduction with Breast cancer.
BRCA1 and BRCA2 and the genetics of Breast Cancer
The Clinical Significance of Epidermal Growth Factor
Receptor (EGF-R) in Human Breast Cancer.
Causes
Diagnosis
Treatment
Conclusion
INTRODUCTION
In western countries, up to 20% of women diagnosed with breast cancer have at
least one relative who is also affected by the disease.
Part of this familial clustering shows autosomal dominant inheritance with
high penetrance and is due to mutations in the BRCA1 and BRCA2
breast cancer genes.
Germline mutations in the BRCA1 and BRCA2 genes confer a strongly elevated
risk of breast cancer.
Women who inherit a mutated copy of the BRCA1 gene have an estimated lifetime
risk of breast cancer ranging from 56% to 87%, which is comparable to the
lifetime Risk for BRCA2 mutation carriers.
Why mutation in BRCA1 and BRCA2 lead
specifically to Breast and Ovarian cancer ?
BRCA1 AND BRCA2 AND THE GENETICS OF BREAST
CANCER
The function of BRCA1 and BRCA2 suggested that they are involved
in two fundamental cellular processes : DNA damage repair and
transcriptional regulation.
BRCA1 & BRCA2 genes contain high densities of repetitive
elements.
Somatic alteration of BRCA 1 & 2 are common and driven by
rearrangement between repetitive elements (Mary.C.K., et.al 2001)
The breast and ovary are estrogen responsive tissue. Beginning in
puberty, the breast epithelium proliferate rapidly in response to
fluctuating level of estrogen.
BRCA – mediated tumorigenesis are estrogen-mediated proliferation
of breast and ovarian epithelium and distinctive genomic context of
the BRCA genes.
The normal protein products of BRCA1 and BRCA2 are involved in
the fundamental cellular process of maintaining genomic integrity
and transcriptional regulation.
Tumorigenesis in individuals with germline BRCA mutation requires
somatic inactivation of the remaining wild-type allele, suggesting
that the BRCA genes are tumor suppressors. (Smith S.A. et.al 1992
and Collins. et. al 1995)
BRCA1 mutation carrier has a lifetime risk of breast cancer that is
greater than 80%.
BRCA1 & BRCA2 proteins are involved in control of homologous
recombination (HR)

And double-strand break repair in response to DNA
damage.(Hall,J.M. et,al 1990, Miki, Y. et,al 1994, Wooster,R., et,al
1994 Deng,C.X, et, al 2000)

Within minutes of DNA damage, the histone H2A family member
H2AX become extensively phosphorylated and forms foci at break
sites (Paull,T.T., et,al 2000)
BRCT domain of BRCA1 bind double-stranded breaks in DNA
(Yamane, et,al 2000)
BRCA1 & BRCA2 also function as transcriptional co-regulators
through direct interaction with sequence specific transcription
factors with components of the transcriptional machinery.
Chromatin remodeling function have been attributed to both BRCA1
and BRCA2.
BRCA1 is a component of the human SW1/SNF- related chromatin
remodeling complex(Bochar et,al 2000)
BRCA1 has the ability to co-activate endogenous p53-dependent
stimulation of p21and p53 promoter is dependent on its physical
association with the SWI/SNF complex.

It remains to be determine whether the function of BRCA1 in
the SWI/SNF complex is to direct chromatin remodeling to
sites of DNA damage, allowing repair protein to function ,
and/or whether this complex is essential for activation of
genes critical tothe DNA damage response pathway.

Among sporadic breast tumors, 50-70% have lost an allele of
BRCA2 (Futreal, et.al 1994 and Cleton, et.al 1995)
DISTINCTIVE GENOMIC INSTABILITY OF BRCA GENES
The genomic region of BRCA1 and BRCA2 contain very high densities of
repetitive DNA elements that may contribute to genetic instability.
The BRCA1 region consists of 42% Alu sequences and 5% non-Alu
repeats (Smith et.al1996)
The BRCA2 region consists of 47% repetitive DNA, consisting of 20% Alu
sequences and 27% LINE and MER repetitive DNA
[ Alu sequences:- The Alu family is a set of dispersed, related sequences,
each ~300 bp long, in the human genome. The individual members have
Alu cleavage sites at each end (hence the name)]
[LINE elements have two reading frames, one coding for a nucleic acidbinding protein, the other for reverse transcriptase and endonuclease
activity.]
Alu mediated genomic rearrangement within BRCA1 and genomic
rearrangement in BRCA2 in as yet unexplained high risk breast cancer
families will reveal other such complex rearrangements
(T.D. Walsh, Unpublished results)
Chromatin organized into topologically constrained loop that are anchored
to the matrix attachment regions (MARs)
(Getzenberg et.al 1991 , Jack et.al 1992 and Roberge et.al 1992 )
ESTROGEN AND SURVIVAL OF BRCA-DEFICIENT
CELLS
Generally, failure of DNA repair leads to growth arrest or cell death.
BRCA defficient mice die early in embryogenesis as the result of cellular
responces .(Mary.C.K., et.al 2001)
The first paradox of BRCA biology is that BRCA-deficient breast or Ovarian
epithelial cells develop tumours instead of under going cell death.
The longer the women is exposed to estrogen either endogenously or
exogenously the higher her risk of developing breast cancer; both early
onset of menarche and late menopause are associated with increase risk
Human breast tissue begin develop to develop very early, usually during
the sixth week of fetal development. Fetal Breast tissue is responsive to
circulating maternal hormones. After early infancy, no developmental
changes occur in breast until puberty.
INHERITED BREAST CANCER
All breast epithelial cell of a BRCA mutation carrier have one inactivated
allele of BRCA mutation carrier have one inactivated allele of BRCA1 and
BRCA2.
During puberty, in direct response to estrogen surges, these cells rapidly
proliferate.
There is a dramatic increase in the rate of cellular replication strain the
DNA repair capacity of breast epithelia cells.
The BRCA-null cells are deficient in repair, they would sustain DNA damage
at many sites, often including genes essential to cell checkpoint activation.
Mutation of a check point gene would enable a BRCA-null cell to escape
death permanently and to proliferate.For BRCA1-mediated tumorigenesis,
one of the key check-point gene is p53.
Evidence from conditional knock-out mice suggests that loss of BRCA1
in mammary cells leads to incomplete proliferation, apoptosis and tumors
at low frequency (Xu et.al 1999)
Tumors in patients with germline BRCA1 or BRCA2 mutation are
frequently associated with somatic mutation of p53.
If somatic inactivation of wild-type BRCA allele and mutation of critical
check point genes occur during puberty, then breast tumorigenesis would
be an early events in the life of these women.
SPORADIC BREAST CANCER
Analysis of sporadic tumor may suggest that either one or two alleles
can be lost, yielding similar patterns of allelic imbalance.
Repeat mediated loss of chromatin loops formed at different points in the
cell cycle will yield deletion of different sizes. If this process occur in
both copies of the BRCA1 region or BRCA2 region , deletion including
deletion overlap , they may incorrectly appear to define one region of LOH.
Presence of single BRCA1 or BRCA2 allele is sufficient for normal growth
& development, because person heterozygous for BRCA mutations have
normal phenotype apart from their cancer predisposition.
Transcriptional silencing may inactivate BRCA allele by loss of protein
that regulate the expression or by an increase in negative regulatory
protein (Id4 inhibitor of DNA binding 4) (Beger et.al 2001).
Thus far, only one protein has been identified as a regulator of BRCA2
expression. This is NF-ќ B, which upregulates BRCA2 transcription by
binding the BRCA promoter (Wu et.al 2000)
NF-ќ B, regulates expression of a number of genes withcritical role in
apoptosis, tumorigenesis and inflammation. NF-ќ B stimulates cell
cycle progression in estrogen receptor breast cancer.
Methylation of CpG islands also cause transcriptional silencing.
The role of ATM, ATR and Chk2 in phosphorylating BRCA1 in response of
DNA damage.
[The ATR kinase exists in a complex with the another protein (ATRIP).ATRIP is recruited to single-stranded DNA
by the RPA protein complex (a component of the replication fork that binds to free single-stranded DNA).
This localizes ATR at the single-stranded DNA, where it acts on Chk enzymes to trigger the damage response
(Zou and Elledge, 2003).]
The C-terminus of BRCA2 is phosphorylated by hBUBR1, a mitotic
check point gene (Futamura et.al 2000)
THE CLINICAL SIGNIFICANCE OF EPIDERMAL GROWTH
FACTOR RECEPTOR (EGF-R) IN HUMAN BREAST CANCER.
Understand the role of the Epidermal Growth Factor.
Relationship with other prognostic factors
EGF can stimulate the growth of normal mammary epithelium and human breast
cancer cells in vitro . Receptors for EGF have been demonstrated on several breast
cancer cell lines, especially on estrogen receptor (ER)-negative tumor cells, and in
human primary tumors and metastases (4, 13, 14).
ER-positive rates ranging from 34-82%. In general, EGF-R positivity was observed
in 29-91% (mean: 59%) of ER-negative and in 4-51% (mean: 29%) of ER-positive
tumors reported by 21 different laboratories .
Relationship with age or menopausal status
With respect to menopausal status, Sainsbury et al reported 45% and 30%
EGF-R positivity in premenopausal and postmenopausal patients,
respectively. Six other groups found no relationship between menopausal
status and EGF-R levels (20,35, 39, 41, 44, 66).
Relationship with tumor size.
Nicholson (25) reported a significant positive correlation between EGF-R and
increasing tumor size. In addition, Spitzer et al. (56) found higher
EGF-R l
Based on 40 separate studies comprising 5232 patients, the mean
percentage of EGF-R positivity reported in breast cancer is 45%
(range 14-91%). (J. G. M. KLIJN,1992)
A few studies indicate a positive correlation between EGF-R and higher
rates of breast cancer proliferation
CAUSES
1. Getting Older
Like most cancers, breast cancer becomes more common the older
women are
2. A Previous Breast Cancer
Awoman who has had breast cancer has an increased risk of getting
another in the other breast.
3 .Significant Family History
The National Institute for Clinical Excellence have issued guidelines in
2004 that set out the criteria for a family history that means she may
have an increased risk of breast cancer .
A women may have an increased risk of breast cancer if she have ONE of
the following in her family:
•
Mother or sister diagnosed with breast cancer before the age of 40 .
2 Close relatives from the same side of the family diagnosed with breast
cancer - at least one must be a mother, sister or daughter
3. Close relatives diagnosed with breast cancer at any age
4. Father or brother diagnosed with breast cancer at any age
5. Mother or sister with breast cancer in both breasts the first cancer diagnosed before the age of 50
6. One close relative with ovarian cancer and 1 with breast cancer,
diagnosed at any age - at least one must be a mother, sister or daughter
4. Breast Cancer Genes
BRCA1 and BRCA2. Your lifetime risk of getting breast cancer if
she carry either of these breast cancer gene faults could be as much
as 85 %.
5. Benign Breast Disease
If a women have a history of benign breast lumps then she have a
slightly increased risk of breast cancer
6. Hormone Replacement Therapy
Cancer Research UK looked specifically at HRT and risk of breast cancer.
They have shown, for the first time, that combined HRT
(oestrogen and progesterone) is more likely to cause breast cancer
than oestrogen only HRT.
7. Having Dense Breasts
This really relates to how your breast tissue shows up on a mammogram.
Women with dense breast tissue have less fat and more breast cells and
connective tissue in their breasts. Because they have a greater proportion
of breast cells, the risk of breast cancer is higher.
8. Alcohol
Alcohol intake has been linked to breast cancer. A large study looked at
about 80% of the worldwide research on alcohol. They found there was an
increase in risk of breast cancer with the amount of alcohol that women
regularly drink. With each additional daily measure - a glass of wine for
example - there is an increase in risk of breast cancer of about 7%.
9. Body Weight
This is a bit confusing. If a women is overweight before she had her menopause,
her risk of breast cancer is lower than average. But if she is overweight after
menopause, her risk of breast cancer is higher than average.This is because
women who are overweight ovulate less than average. So their breast cells
may be exposed to lower levels of oestrogen. But once she is post menopausal,
her oestrogen levels are linked to the amount of body fat she have.
The more fat, the higher your oestrogen levels are likely to be.
10. Being Tall
Taller women may have more breast tissue, which may mean a higher
risk of breast cancer.
11. Lobular Carcinoma In Situ (LCIS)
LCIS is not cancer. It means there are some changes to the cells in
a women’s breast, but the cells are not yet cancer cells. Having LCIS does
increase the risk of getting breast cancer. But, even so, most women
with LCIS will not have cancer.
12. Radiation And Breast Cancer
13. Past Treatment For Hodgkin's Lymphoma
If a women had radiotherapy to her chest for Hodgkin's lymphoma in the past,
you could be at increased risk of getting breast cancer. This applies to all those
women treated as children or in their 20s.
Features of Breast.
Each breast has 15 to 20 sections called lobes, with many smaller
sections called lobules.
Within each lobe are many smaller lobules.
Lobules end in dozens of tiny bulbs that can produce milk.
The lobes, lobules, and bulbs are all linked by thin tubes called ducts.
These ducts lead to the nipple in the center of a dark area of skin called the areola.
Fat surrounds the lobules and ducts.
There are no muscles in the breast, but muscles lie under each breast and cover
the ribs.
Types of Breast Cancer.
The most common type of breast cancer is the one that affects these tubes.
called ductal cancer, it is found in the cells of the ducts.
Cancer that begins in the lobes or lobules is called lobular carcinoma.
Lobular carcinoma is more often found in both breasts than are other types of
breast cancer
The swelling of the breast is an uncommon type of breast cancer. This rare type
of breast cancer is called inflammatory breast cancer which symptoms
include an increase in the skin temperature, redness, and swelling of the breast.
The skin may show signs of ridges and welts or it may also have a pitted
appearance. This type of cancer tends to spread quickly.
Pictures of Types of Breast Cancer
Ductal Carcinoma in situ (DCIS)
Lobular Carcinoma in situ (LCIS)
Invasive Ductal Carcinoma (IDC)
Invasive Lobular Carcinoma (ILC)
Diagnosis.
1.Self Examination.
2.Ultra Sound guided Breast biopsy.
Si RNA induced silencing of defective BRCA genes.
My personal hypothesis.
2.Ultra Sound guided Breast biopsy
Ultrasound is an excellent way to evaluate breast
abnormalities detected by mammography.
Ultrasound-guided breast biopsy is a highly accurate way to evaluate suspicious
masses within the breast that are visible on ultrasound, whether or not they can
be felt on breast self-examination or clinical examination.
The procedure prevents the need to remove tissue surgically, and also
eliminates the radiation exposure that comes from using x-rays to locate a mass.
After placing an ultrasound probe over the site of the breast lump and
using local anesthesia, the radiologist guides a biopsy needle directly
into the mass.
Tissue specimens are then taken using either an automatic spring-loaded
or vacuum assisted device (VAD).
Stages in Breast Cancer.
Tumor Size.
STAGE I - The cancer is no wider than 2 centimeters (about 1 inch) and has not
spread outside the breast.
Stage II - The tumor is more than 2 cm but less than 5 cm in the greatest
dimension.
Stage III - Tumor is more than 5 cm in the greatest dimension.
Stage IV- Tumor of any size with growth extending to the chest wall or skin.
Breast Cancer Treatment Methods
1. Surgery
2. Radiation therapy
3. Chemotherapy
4. Hormonal therapy
5. Biological therapy
Surgery
An operation to remove the cancer but not the breast is called
breast-sparing surgery or breast-conserving surgery .
Lumpectomy and segmental mastectomy (also called partial mastectomy)
are types of breast-sparing surgery. After breast-sparing surgery,
most women receive radiation therapy to destroy cancer cells that remain in the
area.
An operation to remove the breast (or as much of the breast as possible) is
a mastectomy.
Breast reconstruction is often an option at the same time as the mastectomy,
or later on.
In lumpectomy, the surgeon removes the breast cancer and some normal
tissue around it. Often, some of the lymph nodes under the arm are removed.
Radiation therapy
Front view
Side view
Cross–sectional view
During radiation therapy, patients may become extremely tired, especially
after several treatments.
Resting is important, but doctors usually advise their patients to try to
stay reasonably active
Chemotherapy
chemotherapy affects normal as well as cancer cells .
In general, anticancer drugs affect rapidly dividing cells .
These include blood cells, which fight infection, help the blood to clot, and
carry oxygen to all parts of the body.
When blood cells are affected, patients are more likely to get infections, may
bruise or bleed easily, and may feel unusually weak and very tired.
Rapidly dividing cells in hair roots and cells that line the digestive tract may
also be affected.
Some anticancer drugs can damage the ovaries. If the ovaries fail to produce
hormones, the woman may have symptoms of menopause She may not be able
to become pregnant
Hormonal Therapy
Estrogen Receptors
Tamoxifen Blocking Estrogen Receptors
Tamoxifen is the most common hormonal treatment
This drug blocks the cancer cells' use of estrogen but does not stop estrogen
production.
 It can cause blood clots in the veins, especially in the legs and in the lungs.
tamoxifen can cause cancer of the lining of the uterus
Biological Therapy
Is a treatment designed to enhance the body's natural defenses against cancer.
For example
Herceptin® (trastuzumab) is a monoclonal antibody that targets breast cancer
cells that have too much of a protein known as
human epidermal growth factor receptor-2 (HER-2).
By blocking HER-2, Herceptin slows or stops the growth of these cells.
Herceptin may be given by itself or along with chemotherapy.