Triple-negative

Breast Cancer
Dr Rebecca Dent
Sunnybrook Odette
Cancer Center, Canada
Basal-Like
The Triple Negative Breast Cancer
Estrogen Receptor (ER) negative
Progesterone receptor (PR) negative
Her2neu (HER2) negative
ER/PR/HER2 -
ER/PR/HER2 -
Background
Increasing evidence that breast cancer is not
one disease
Development of therapies targeting steroid hormone
receptors
led to recognition of estrogen receptor-positive and negative
breast cancers
Trastuzumab (Herceptin) therapy
highlighted the importance of reliably identifying tumors amplified
or over expressed HER2
Baselga and Norton, 2002
Evolution in Breast Cancer Classification
Classical Diagnosis
Ductal infiltrating
carcinoma of breast
with high grade of
nuclear atypia
Protein Expression
ErbB2 over expressing
breast tumour
Gene Expression
Profiling
Partial two dimensional
cluster analysis of
17 breast tumours
Morphological
Diagnosis
Immunohistochemical
assessment
DNA microarray
analysis
Gene expression patterns of breast
carcinomas distinguish tumour subclasses
with clinical implications
Therese Srlie
a,b,c
, Charles M. Perou
a,d
, Robert Tibshirani
e
, Turid Aas
f
, Stephanie Geisler
g
, Hilde Johnsen
b
, Trevor
Hastie
e
, Michael B. Eisen
h
, Matt van de Rijn
i
, Stefanie S, Jeffrey
j
, Thor Thorsen
k
, Hanne Quist
l
, John C. Matese
c
,
Patrick O. Brown
m
, David Botstein
c
, Per Eystein Lnning
g
, and Anne-Lise Brresen-Dale
b,n
Basal-like
Subgroup
= E
HER2
Subgroup
= D
Normal
breast
like
Luminal
Subtype
C
Luminal
Subtype
B
Luminal
Subtype
A
ER
Gene
expression
E
D
C
B
A
O.S.
months
Adapted from Sorlie et al. PNAS, 2001
Gene Expression Patterns of Breast
Carcinomas Predict Survival
ER
Gene
expression
Molecular classes are predictive of outcome
1
0.8
0.6
0.4
0.2
0
0 24 48 72 96
RFS
p<0.01
P
r
o
b
a
b
i
l
i
t
y
1
0.8
0.6
0.4
0.2
0
0 24 48 72 96
Survival Months
p<0.01
P
r
o
b
a
b
i
l
i
t
y
Lum A Lum B&C Censored NorB-like Basal ERBB2+
Overall survival:
A. 5 tumor subtypes
(based on figure 1)
Relapse-free survival:
B. 5 tumor subtypes
(based on figure 1)
Why is this group called
Basal-Like ?
Breast Cancers can be broadly divided
Expression of
luminal keratins
(ie. simple epithelial
type keratins)
CK 7, 8, 18 and 19
High levels of
expression of genes
that are characteristic
of basal epithelial
cells of the normal
mammary gland
(ie. the stratified
epithelial cytokeratins)
CK 5, 6, 14, 15 and 17
How do we identify
Basal-Like Breast Cancers
in the clinical setting?
Basal-Like Breast Cancers
Fadare O, and Yeh IT
Pathology Case Reviews
July/August 2007
Immunohistochemical Characterization of the
Basal-Like Cancers
Took 21 tumors defined as basal-like by microarray
and performed IHC analyses with six distinguishing
markers (ER, CK5/6, EGFR, HER2, cKIT)
Concluded that an IHC surrogate for gene array
experiments to identify basal-like breast cancers is:
ER
HER2
CK5/6 + and/or EGFR +
16 out of 21 tumors expressing the basal gene
signature would have identified 76% of the BLC with
a specificity of 100%
Nielsen et al. 2004
Limitations of Nielsen Definition
(ER -, HER2 -, CK5/6 + and/or EGFR +)
The extent and intensity of staining for CK5/6 is quite
variable in carcinomas
Hannermann et al. found that only one-third of cases showed positive
staining in more than 50% of tumor cells
Livasy et al. attempted to replicate using same
definition as Nielsen
Calculated a sensitivity of 55% and specificity of 100%
Definition of Basal-Like Breast Cancers
Multiple immunohistochemical markers have been
used to identify Basal-Like differentiation
No universally agreed upon criteria or precise set of
basal markers
Nielsen et al. ER/PR/HER2 negativity in addition to
either EGFR + and/or CK5/6 +
Smith et al. high molecular weight cytokeratins CK5,
14 and 17
Carey et al. ER/PR/HER2 negativity
80 to 90% of these will be
Basal-Like Breast Cancers
100 Patients with
Triple Negative Breast Cancer
ER/PR/HER2 -
ER/PR/HER2 -
Triple Negative Breast Cancers
Comprise approximately 15% of all invasive cancers
More common in:
Younger patients
African Americans
BRCA1 mutation carriers
Unique Morphologic Attributes
Pushing border
high grade
central scarring/acellular zone
Stromal/peritumoral lymphocytic infiltrate
Ethnicity and the BLC
Carolina Breast Study (N=500, 1993-96)
ALL women = 20%
Caucasians
Pre and Postmenopausal = 16%
African American Women
Premenopausal = 39%
Postmenopausal = 14%
*Nigerian data up to 50-60% of women with invasive breast
cancer with the BLC (mean age of dx 44)
Olopade et al. AACR April 2005
Carey et al, JAMA 2006
BRCA1 and BLC phenotype
BRCA1 is a tumor suppressor gene that functions in
DNA repair
These tumors are often ER-, HER2- (up to 80% of BRCA1
associated tumors have been shown to be basal-like)
Thoughmost basal-like tumors have normal BRCA1
Hereditary BRCA1 breast tumors and basal-like
sporadic tumors have a similar phenotype and gene
expression signature
Suggesting involvement of BRCA1 in the pathogenesis of
sporadic basal-like cancer
BRCA1/BLC
BRCA1 mutations are rarely found in sporadic breast
cancer, but BRCA1 expression is often reduced,
especially in basal-like breast cancers
Why?
BRCA1 promotor methylation has been demonstrated in
7 to 31% of sporadic cancers (Catteau et al)
LOH of the BRCA1 locus which occurs in 15 to 45% of
sporadic breast cancers (Rio et al)
However, studies looking at the relationship between BRCA1
methylation, BRCA1 LOH and clinical features of breast
tumors have been inconsistent
ID4, a negative regulator of BRCA1, was found to be
expressed at a 9-10 times higher in BLC (Turner et al)
Shared Features with BRCA-1 Cancers
BRCA1
associated
breast cancer
Basal-Like
Breast Cancer

High grade

Cytogenic abnormalities

Estrogen receptor Negative

HER2 negative
Express Cytokeratin 5/6
EGFR overexpression and mutation
p53 mutation
Turner et al. 2006
Typical Histological Features of Triple
Negative Breast CA
Grade 3, IDC,
with central fibrosis
Positive Staining
for EGFR
Positive Staining
for CK 5/6
Immunohistochemical Features:
Other Breast CA
n (%)
Basal-like
n (%)
p value
3/21 (14%)
< 0.001
C-KIT 67/605 (11%) 6/21(30%) < 0.001
< 0.001
P53 protein 27/124 (22%) 32/63 (51%) < 0.006
0.0001
0/21 (0)
13/21 (62%)
12/21(57%)
9/11 (82%)
17/18 (94%)
517/665 (78%)
23/87 (26%)
105/761 (14%)
14/521 (8%)
13%
2/28 (7%)
ER expression
HER2
Cytokeratin 5/6
EGFR
P53 mutation
Vimentin
Livasy et al, Sorlie et al, Foulkes et al, Nielsen et al
Triple-Negative Breast Cancer: Clinical
Features and Patterns of Recurrence
HBBC database (1987-1997)
1601 (80%) of patients had details on hormone
receptors/HER2 and were eligible for the study
180 (12%) of the 1601 patients were defined as
triple negative breast cancers
Mean follow up was 8.1 years
Dent, R. et al. Clin Cancer Res 2007
Characteristics of Triple Negative vs. Other Breast Cancers
Characteristic
Other
(N=1421)
number (percent)
Triple Negative
(N=180)
number (percent)
Significance
p value *
Mean Age at Diagnosis (yrs) 57.7 53 p < 0.0001
Mean Tumor Size 2.1 cm 3.0 cm p < 0.0001
Tumor Size
(62.7) (36.5)
(55.6)
(7.9)
(54.4)
(44.6)
(9.8)
(24.2)
(66.0)
(32.8)
(4.6)
(45.6)
(54.4)
(19.9)
(51.8)
(28.3)
T1 ( 2 cm) 880 65 p < 0.0001
T2 (>2cm to 5cm) 461 99
T3 (>5cm) 64 14
Missing 16 2
Lymph Node Status
Positive 510 87 p = 0.02
Negative 609 70
Missing or Not Tested 302 23
III 336 101
Tumor Grade
I 237 15 p < 0.0001
II 616 37
Missing
* p values were calculated with the use of the chi-square test
Dent, R. et al. Clin Cancer Res 2007
Tumor Size by Nodal Status according to Basal-Like Group
Non Basal-like Group
(N=1421)
Basal-like Group
(N=180)
Tumour Size
Lymph Node Positive
Number (percent)
Lymph Node Positive
Number (percent)
<1.0 cm 38 (19.3) 5 (55.6)
(39.3) (55.6)
(48.9)
(92.3)
p=0.042
(59.5)
(91.4)
p<0.0001
1 - 2 cm 180 25
2.1- 5 cm 238 43
>5.1 cm 53 12
* p values were calculated with the use of the chi-square test
Dent, R. et al. Clin Cancer Res 2007
Distant Recurrence
Dent, R. et al. Clin Cancer Res 2007;13:4429-4434
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1.0
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0.0
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18
Years after diagnosis
p<0.0001 (log-Rank test)
Other (290 of 1421) Triple-negative (61 of 180)
Overall Survival
Dent, R. et al. Clin Cancer Res 2007;13:4429-4434
P
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1.0
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0.0
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18
Years after diagnosis
p<0.0001 (log-Rank test)
Other (261 of 1420) Triple-negative (62 of 180)
Hazard Rate of Distant Recurrence
Dent, R. et al. Clin Cancer Res 2007;13:4429-4434
H
a
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a
r
d

R
a
t
e
0.35
0.00
0 1 2 3 4 5 6 7 8 9 10
Years after first surgery
Other (290 of 1421) Triple-negative (61 of 180)
0.30
0.25
0.20
0.15
0.10
0.05
Median Time from Distant Relapse to Death
0 5 10 15 20 25
22 months
9 months
Triple Negative
Breast CA
Other Breast
CA
Dent R, Trudeau M, Pritchard K, Hana W, Narod S. et al. Clinical Cancer Res 2007
Hazard Ratios (HR) for local and distant recurrence & death
Basal-like vs.
Non-Basal
a
Univariate Analysis
HR (95% CI)
Significance
p value
Multivariate Analysis
HR (95% CI)
Significance
p value
Recurrence Risk of Local
Whole FUP 1.12 (0.7 1.7) p = 0.6 0.82 (0.5 1.3) p = 0.38
Up to 5yrs 1.54 (0.9 2.5) p = 0.08 1.01 (0.6 1.7) p = 0.98
p = 0.12
p < 0.0001
p < 0.0001
p = 0.09
p < 0.0001
p < 0.0001
p = 0.84
(0.1 1.3) (0.1 1.2)
(0.8 1.5)
(1.1 2.0)
(0.1 0.8)
(1.0 1.6)
(1.3 2.4)
(0.5 1.2)
(1.4 2.5)
(2.0 3.5)
(0.2 1.1)
(1.3 2.2)
(1.9 3.6)
(0.6 1.5)
5 yrs to End 0.40 0.34 p = 0.08
1.9 1.2 p = 0.35
Risk of Distant
Recurrence
2.6 1.5 p = 0.02
0.5 0.3 p = 0.02
1.71 1.25 p = 0.009
2.63 1.74 p = 0.0007
0.96 0.77 P = 0.22
Risk of Death
Adjusted by age at diagnoses, BR grade (1, 2, 3), tumor size, lymph node status, Tam. therapy (yes/no),
and chemo therapy (yes/no).
Patterns of Metastatic Spread
Patterns of Metastatic Spread
More likely to spread to brain, lung and possibly liver
and less likely to spread to bone and soft tissues
Tsuda et al. 2000 Am J of Surgical Pathology
Rodriguez-Pinilla et al. Clinical Cancer Research 2006
Fulford et al. Breast Cancer Research and Treatment 2007
Hicks et al. 2006 Am J of Surgical Pathology
More likely to present with visceral metastases
versus bone metastases as first site of metastases
70% vs 37%, p < 0.001 (Dent et al. SABCS 2007)
Basal-like breast cancers tend to be large
and spread to lung, brain and liver
Why? Faster growing or not diagnosed
Faster growing tumors and more likely to be found
between annual mammograms
The mode of spread of these cancers may be
different and early hematogenous spread is favoured
possibly due to different protein expression profiles
Angiogenic Switch and VEGF dependency
Adapted from Bergers G, et al. Nature 2002;3:40110
Small tumour (12mm)
Avascular
Dormant
Larger tumour
Vascular
Metastatic potential
Angiogenic
switch
Over-expression of pro-angiogenic signals, such as VEGF
Triple Negative Breast Cancer Summary
Triple Negative breast cancers are a distinct category of
breast cancer
Mean age of diagnosis is younger
Tumors tend to be larger and higher grade
Most rapidly fatal breast cancer
Rapid rise in risk of recurrence following diagnosis
Peak risk of recurrence at 1-3 y
Frequent vascular metastases to viscera
Likely early angiogenic switch and thus ideal group to
evaluate the role of anti-VEGFR therapies such as
Bevacizumab early in the course of their treatment