a comparative biomarker study of 514 matched cases of male and female breast cancer reveals...

PRECLINICAL STUDY

A comparative biomarker study of 514 matched cases of maleand female breast cancer reveals gender-specific biologicaldifferences

Abeer M. Shaaban • Graham R. Ball • Rebecca A. Brannan • Gabor Cserni •

Anna Di Benedetto • Jo Dent • Laura Fulford • Helen Honarpisheh •

Lee Jordan • J. Louise Jones • Rani Kanthan • Loaie Maraqa • Maria Litwiniuk •

Marcella Mottolese • Steven Pollock • Elena Provenzano • Philip R. Quinlan •

Georgina Reall • Sami Shousha • Mark Stephens • Eldo T. Verghese •

Rosemary A. Walker • Andrew M. Hanby • Valerie Speirs

Received: 3 October 2011 / Accepted: 25 October 2011 / Published online: 18 November 2011

� Springer Science+Business Media, LLC. 2011

Abstract Male breast cancer remains understudied

despite evidence of rising incidence. Using a co-ordinated

multi-centre approach, we present the first large scale

biomarker study to define and compare hormone receptor

profiles and survival between male and female invasive

breast cancer. We defined and compared hormone receptor

profiles and survival between 251 male and 263 female

breast cancers matched for grade, age, and lymph node

status. Tissue microarrays were immunostained for ERa,

ERb1, -2, -5, PR, PRA, PRB and AR, augmented by

HER2, CK5/6, 14, 18 and 19 to assist typing. Hierarchical

clustering determined differential nature of influences

between genders. Luminal A was the most common phe-

notype in both sexes. Luminal B and HER2 were not seen

in males. Basal phenotype was infrequent in both. No

differences in overall survival at 5 or 10 years were

observed between genders. Notably, AR-positive luminal

A male breast cancer had improved overall survival over

female breast cancer at 5 (P = 0.01, HR = 0.39, 95%

CI = 0.26–0.87) but not 10 years (P = 0.29, HR = 0.75,

95% CI = 0.46–1.26) and both 5 (P = 0.04, HR = 0.37,

95% CI = 0.07–0.97) and 10 years (P = 0.04, HR =

0.43, 95% CI = 0.12–0.97) in the unselected group.

Hierarchical clustering revealed common clusters between

genders including total PR–PRA–PRB and ERb1/2 clus-

ters. A striking feature was the occurrence of ERa on

Presented in part at the 32nd Annual San Antonio Breast Cancer

Symposium, 9-13 December, 2009, San Antonio, TX, Breast Cancer

Research 2010, 18 May 2010, London UK and the Pathological

Society of Great Britain and Ireland 2010 Summer Meeting, 30 June-

3 July, St Andrews, UK.

A. M. Shaaban � R. A. Brannan � G. Reall �E. T. Verghese � A. M. Hanby

St James’s Institute of Oncology, St James’s University Hospital,

Leeds, UK

G. R. Ball

Nottingham Trent University, Nottingham, UK

R. A. Brannan � H. Honarpisheh � L. Maraqa � S. Pollock �E. T. Verghese � A. M. Hanby � V. Speirs (&)

Leeds Institute of Molecular Medicine, Wellcome Trust Brenner

Building, University of Leeds, Leeds LS9 7TF, UK

e-mail: [email protected]

G. Cserni

Bacs-Kiskun County Teaching Hospital, Nyiri ut 38,

Kecskemet 6000, Hungary

A. D. Benedetto � M. Mottolese

Regina Elena Cancer Institute, Rome, Italy

J. Dent

Calderdale Hospital, Halifax, UK

L. Fulford

Surrey & Sussex NHS Trust, Redhill, UK

L. Jordan � P. R. Quinlan

University of Dundee/NHS Tayside, Dundee, UK

J. L. Jones

Barts Cancer Institute, Barts and The London School

of Medicine and Dentistry, London, UK

R. Kanthan

University of Saskatchewan, Saskatoon, Canada

M. Litwiniuk

Poznan University of Medical Sciences, Poznan, Poland

123

Breast Cancer Res Treat (2012) 133:949–958

DOI 10.1007/s10549-011-1856-9

distinct clusters between genders. In female breast cancer,

ERa clustered with PR and its isoforms; in male breast

cancer, ERa clustered with ERb isoforms and AR. Our data

supports the hypothesis that breast cancer is biologically

different in males and females suggesting implications for

clinical management. With the incidence of male breast

cancer increasing this provides impetus for further study.

Keywords Male breast cancer � Hormone receptors �Androgen receptor � Hierarchical clustering

Introduction

According to figures from Cancer Research UK, there were

45,695 cases of female breast cancer (FBC) and 277 cases

of male breast cancer (MBC) diagnosed in the UK in 2007

[1]. In the US it was estimated that 1,970 men and 207,090

women would be diagnosed with breast cancer in 2010 [2].

Whilst MBC accounts for less than 1% of breast cancer

diagnoses worldwide, the overall improvements in survival

and mortality observed in FBC has not been seen to the

same extent in MBC, as demonstrated in a recent interro-

gation of the Surveillance Epidemiology and End Results

(SEER) database [3]. Moreover, the incidence rate of MBC

is rising steadily [4–7].

The etiology of MBC is poorly understood with most of

our current knowledge regarding its biology, natural his-

tory, and treatment extrapolated from FBC. Retrospective

studies are generally weakened by the small numbers of

cases available from any one centre with studies published

on as few as 15 cases [8], making it hard to draw biolog-

ically meaningful conclusions. It is, therefore, a challenge

to accrue sufficiently large numbers to allow comparative

analysis of possible prognostic or predictive biomarkers.

Many articles imply a general similarity of MBC to FBC

and this has resulted in MBC patients being treated in

exactly the same way as females in the clinic, which may

not be optimal. Survival rates for MBC are generally

assumed to be lower than FBC, probably as a result of later

diagnosis and the assumption that treatments which are

proven in FBC through clinical trials will have the same

impact in men [9].

A 40-year review of records of 759 cases from invasive

MBC from the US Armed Forces Institute of Pathology

database showed that the frequency of histological sub-

types in men was comparable to that of FBC, with the

exception of papillary carcinoma which was twice more

common in MBC [10]. To date, modern molecular sub-

typing has been reported in a single study of MBC where

only luminal A (35/42) and luminal B (7/42) subtypes were

observed [11].

Using a co-ordinated multi-centre approach, the aim of

this study was to conduct the first large scale study to

address and compare the expression profile of hormone

receptors and their effect on survival in FBC and MBC.

Methods

Patient cohorts

Following ethical approval from the Leeds (West) Research

Ethics Committee (06/Q1205/156), 514 formalin-fixed

paraffin-embedded blocks of male (251) and matched

female (263) breast cancers were obtained retrospectively.

The latter were all from Europe and the former from Europe

(n = 196) and Canada (n = 55). Informed consent was not

required as the anonymised material pre-dated September

2006, came from a Tissue Bank approved by the UK Human

Tissue Authority (or equivalent) or were from non-UK

patients. Patients had not received any therapy before sur-

gery. Details on adjuvant therapy were not extensively

available; where available this was predominantly endocrine

therapy (tamoxifen). Patient characteristics are presented in

Table 1. Cases were reviewed by specialised breast con-

sultant histopathologists (AMH, AMS, RAB) to confirm

histology and marked up for assembly into tissue micro-

arrays (TMAs) using 3 9 0.6 mm tissue cores per case

taken from formalin-fixed paraffin-embedded material as

previously described [10].

Immunohistochemistry (IHC)

Antibodies, dilution, and retrieval methods are listed in

Table 1. The antibody panel was focused on hormone

receptors oestrogen receptor (ER)a, ERb isoforms, pro-

gesterone receptor (PR) isoforms and androgen receptor

(AR) and additional biomarkers selected to distinguish

molecular subtypes of breast cancer (CK5/6, 14, 18,

HER2). Each marker was run as a batch with appropriate

positive (tissue known to express the biomarker of interest)

and negative (no primary antibody) controls. Scoring was

overseen by AMH, AMS, and RAB. Following visualisa-

tion of the signal with 3-30diaminobenzidine chromogen,

TMAs were digitised (Aperio Technologies), and hormone

E. Provenzano

Addenbrookes Hospital, Cambridge, UK

S. Shousha

Imperial College, London, UK

M. Stephens

University Hospital of North Staffordshire, Stoke-on Trent, UK

R. A. Walker

University of Leicester, Leicester, UK

950 Breast Cancer Res Treat (2012) 133:949–958

123

receptor immunoreactivity was scored using the Allred

system with the following cut offs: ERa[ 2, ERb (and

isoforms) [ 3, AR [ 2, PR (and isoforms) [ 2, as vali-

dated in previous studies [12–14].

Hierachical clustering and principal components

analysis (PCA)

For hierarchical cluster analysis, IHC measurements were

used as inputs for all cases in each of the male and female

cohorts. A Euclidian distance measure was employed with

complete linkage of clusters. Clustering was conducted for

the data structure of cases with the immunohistochemical

parameters and for the immunohistochemical parameters as

they were expressed through the population. Cluster

dendrograms were plotted for both of these analyses for

each gender and compared. PCA was applied to the same

dataset. Analysis was based on covariances between

parameters and cases in the data. Variances were computed

based on the sum of squares/n - 1. Plots of the influences

of variables in the factor plane of the first and second and

the second and third principal components for both the

male and female cases were plotted separately. As with the

hierarchical clustering, both the male and female cases

were combined into a single data set. The influence of

variables in the factor plane of the combined cases were

plotted for the first and second and the second and third

principal components. The distributions of cases within the

combined sets were also plotted.

Statistical analysis

Patient and disease characteristics were compared between

male and FBCs using the v2 test (GraphPad). Associations

with disease-free and overall survival (DFS and OS,

respectively) were analysed by Kaplan–Meier plots and log

rank test. P-values were two-sided, and P \ 0.05 was

considered significant.

Results

A total of 514 cases of breast carcinoma were studied,

including 251 males and 263 females. The median age for

the male cohort was 66 years (range 30–94) and 59 years

(range 27–92) for females. Patient characteristics are

shown in Table 2. As this was a matched cohort, no sig-

nificant differences were observed in grade, or lymph node

status between genders. Significant differences were

observed in the distribution of histopathological subtypes

(P \ 0.0001). There was an even distribution of ductal

phenotype whilst lobular carcinomas found in 9% of the

female cohort was only seen in a single male case. Papil-

lary and mucinous phenotypes were restricted to males. A

significantly higher proportion of males expressed ERacompared to females (80 and 68%, respectively), although

no differences in the frequency of PR was observed (71 and

72%, respectively). Follow up data was available on 183

(73%) male and 237 (90%) female cases.

Both cohorts were classified into molecular subtypes by

IHC: luminal A (ERa?, and/or PR?, HER2-), luminal B

(ERa?, and/or PR?, HER2?), HER2 (ERa, PR-, HER2?)

and basal-like (ERa-, PR-, HER2-, CK5/6?) according to

previous studies [15–17]. Representative immunopro-

files for each subgroup are shown in Fig. 1. Significant

Table 1 Patient characteristics of male and female breast cancers

Characteristic Male

(n = 251)

no (%)

Female

(n = 263)

no (%)

P values

Histology \0.0001

Ductal 208 (83) 220 (84)

Lobular 1 (0.4) 23 (9)

Papillary 11 (4) 0

Intraductal papillary 4 (1.5) 0

Micropapillary 1 (0.4) 0

Mucinous 9 (4) 0

Mixed 13 (5) 14 (7)

Unknown 4 (1.6) 0

Grade 0.948

1 25 (10) 29 (11)

2 128 (51) 140 (53)

3 81 (33) 94 (36)

Unknown 17 (6) 0

Lymph node 0.521

? 76 (30) 131 (50)

- 80 (32) 121 (46)

Unknown 95 (48) 11 (4)

ERa \0.0001

? 201 (80) 180 (68)

- 22 (9) 56 (21)

Unknown 28 (11) 27 (10)

PR 0.838

? 177 (71) 190 (72)

- 39 (15) 44 (17)

Unknown 35 (14) 29 (11)

Subtypea 0.0004

Luminal A 199 (98) 197 (90)

Luminal B 0 (0) 14 (6)

Basal 4 (2) 4 (2)

HER2?/ERa- 0 5 (2)

a Subtype classification was not possible in cases from 48 males and

43 females due to core loss from the section for at least one of the

biomarkers, or less commonly, exhaustion of the TMA core

Breast Cancer Res Treat (2012) 133:949–958 951

123

differences were observed between molecular subtypes

(P = 0.0004). Luminal A was seen in 98% of males and 90%

of females. Luminal B or HER2 subgroups were not

observed in males but found in 6 and 2% of females,

respectively. Basal-like tumours (ERa-, PR-, HER2-,

CK5/6?) were infrequent in both cohorts (2% in each).

We then examined the frequencies of expression

between genders of other hormone receptors including AR,

nuclear and cytoplasmic ERb1 and ERb2, nuclear ERb5,

plus the PR isoforms A and B (Table 3). AR immunore-

activity was expressed in 64% of males and 93% females,

respectively, (P \ 0.0001). For ERb1 and -2, both nuclear

and cytoplasmic immunoreactivity was assessed [12].

ERb1 nuclear immunoreactivity was significantly expres-

sed in FBC whilst cytoplasmic ERb1 and ERb2 immuno-

reactivity were associated with MBC. No associations were

observed for ERb5. Of the PR isoforms, only PRA was

significantly expressed in MBC. As the male cohort con-

tained cases of European and Canadian origin, we tested if

there were differences between these; none were found.

In luminal A carcinomas, no differences in overall sur-

vival were observed between genders at either 5 or

10 years (Fig. 2a, b). This was also reflected in the unse-

lected cohorts (data not shown). When hormone receptor

expression was considered, only AR significantly associ-

ated with survival. AR-positive luminal A MBC had sig-

nificantly improved overall survival over the equivalent

FBC at 5 (P = 0.01, HR = 0.39, 95% CI = 0.26–0.87) but

not 10 (P = 0.29, HR = 0.75, 95% CI = 0.46–1.26) years

(Fig. 2c, d). In the unselected group, ERa and AR-positive

MBC had significantly improved overall survival over ERaand AR-negative cases at both 5 (P = 0.04, HR = 0.37,

95% CI = 0.07–0.97) and 10 (P = 0.04, HR = 0.43, 95%

CI = 0.12–0.97) years (Fig. 2e, f) with ERa and AR-

positive MBC also having significantly improved overall

survival over the equivalent FBCs at 5 (P = 0.05,

HR = 0.48, 95% CI = 0.29–1.00) but not 10 (P = 0.37,

HR = 0.79, 95% CI = 0.48–1.32) years (Fig. 2g, h).

Hierarchical clustering based on hormone receptor profiles

classified MBC and FBC into three distinct groups (Fig. 3).

The cytoplasmic ERb cluster was common to both genders. In

FBC an ERa/PR cluster was observed, grouping ERa and PR

isoforms, whilst ERb isoforms clustered with AR (ERb/AR

cluster). In MBC, there were striking changes in the position

of ERa; AR and ERa clustered with ERb (ERa/b AR cluster)

whilst PR isoforms formed an independent cluster (PR clus-

ter). This was also reflected in a PCA-based plot of variable

factor co-ordinates (data not shown).

Discussion

Currently, MBC is treated based on the assumption that it

is essentially the same disease as FBC. In this the largest

comparative study to date, directly comparing the immu-

nohistochemical profile of matched MBCs and FBCs has

revealed that whilst superficially there is similarity between

genders, when probed more deeply, subtle differences are

uncovered.

The histological breakdown of our cohort is in line with

previous reports with the papillary phenotype and variants

thereof, which are twice more common in males, only seen

in the male cohort [10]. Although we did not observe any

mucinous carcinomas in our female cohort the expected

frequency of this phenotype is only 0.9% [18]. We observed

only a single case of lobular carcinoma in males; this is to be

expected given the rarity of this phenotype in men [19].

Thus, our cohort can be regarded as representative.

Table 2 Details of antibodies used for immunohistochemical analysis

Antibody Clone Supplier Dilution Secondary detection Pre-treatment

ERa 1D5 DAKO 1:100 DAKO EnVision High pressure heat retrieval

in Vector low pH antigen

unmasking solutionERb1 PPG5/10 Serotec 1:20

ERb2 57/3 Serotec 1:20

ERb5 5/25 Serotec 1:50

Total PR PgR636 DAKO 1:200

PRA 16 Novacastra RTU

PRB San27 Novacastra RTU

AR AR441 DAKO 1:200

CK5/6 D5/16 B4 DAKO 1:100

CK14 LL002 Serotec 1:50

CK18 CY-90 Sigma 1:500

CK19 RCK108 DAKO 1:150

RTU ready to use


123

We used IHC to classify our breast tumours into

molecular subtypes. Whilst gene array analysis is still

considered the ‘gold standard’, we were unable to apply

this to our 514-case cohort due to high cost. Nevertheless,

molecular profiling of breast cancer based on immunohis-

tochemical typing has now gained widespread acceptance

as a surrogate method and is arguably more robust as it

overcomes the limitations of gene array in that it directly

identifies the cells expressing the marker of interest [15–17,

20]. When applied to TMAs, a limitation is core loss,

which we experienced in this study and which may have

impacted on the higher than anticipated levels of luminal A

phenotype in the female population. Nevertheless, luminal

A phenotype was the most common in both sexes with

basal-like tumours infrequent in both. Sporadic expression

of basal cytokeratins has been previously reported in MBC

[21]. Interestingly, the luminal B phenotype was not seen

in males. This contrasts a recent study of 42 MBCs where

luminal B was seen in 17% of cases [11]. The lack of

luminal B carcinomas in our male cohort reflects the

absence of HER2 expression, which has been variably

reported in MBC ranging from 0 to 95% [22]. It is notable

that some of the earlier studies on HER2 relied solely on

IHC to determine positivity, considering any degree of

membrane immunoreactivity positive [8, 22–28]. The

validity of studies relying on HER2 IHC without recogni-

tion of gene expression is questionable. According to

ASCO/ACP and NEQAS guidelines only those scoring 3?

or above are considered HER2 positive. Equivocal cases

are scored 2? and go forward for FISH analysis and only

those with HER2 gene amplification are considered posi-

tive. In studies using both IHC and FISH to detect HER2,

protein expression was always higher than gene amplifi-

cation [29–31]. Although we relied on IHC to detect

HER2, we are confident our data is robust; we are a

regional HER2 testing centre and our HER2 IHC was

conducted via this service using two different antibodies

and running the test according to clinical standards. Whilst

ERα PR HER2 CK5/6

Not

applicable

Not

applicable

Not

applicable

a

b

c

d

Fig. 1 Semi-serial sections from male (a, c) and female (b, d) breast

carcinoma TMAs showing immunoprofiles for each molecular

subgroup. a luminal A (ERa?, PR?, HER2-), b luminal B

(ERa?, PR?, HER2?), c basal (ERa-, PR-, HER2-, CK5/6?),

d HER2 (ERa-, PR-, HER2?). Original magnification = 910

(TMA core) and 940 (insets)


123

we did not observe any cases which scored [2? in our

MBC cohort, we observed scores of 1? in 22/251 cases

(8%). These would be considered negative in clinical

practice. Nevertheless, this provides confidence that the

high frequency of HER2 negativity we observed was not

simply due to antigen degradation in archival material.

It has been suggested that separation of luminal A and

luminal B breast tumours should be based on the expression

of proliferation markers such as Ki67, not on HER2

expression as is currently the case [32]. However, this has

yet to gain widespread acceptance and we believe there are

several issues that still need to be standardised before this

can be implemented. These include choice of Ki67 antibody,

e.g. MIB1 or SP6 [33] and how to optimally distinguish

between low and high proliferation scores. Once these issues

are resolved it will be interesting to determine if the differ-

ence between MBC and FBC in this series is purely the result

of lack of HER2 expression, or if MBCs have a lower pro-

liferative index as well. Contrary to the general impression,

one of the most significant findings from this, the most

authoritative study to date, was the observation of no dif-

ferences in overall survival at 5 or 10 years between genders

in either our unselected or the luminal A cohorts. Whilst

there have been a number of case–control and population-

based studies addressing survival in MBC using data from

cancer registries [3, 4, 34, 35], direct comparative studies

between genders are scarce. A Chinese study of 35 MBC and

70 matched FBC showed the latter had significantly better

overall survival at 5 and 10 years, but when the comparison

was restricted to female postmenopausal, outcomes were

similar [36]. Despite a small number of cases in the male arm

and unbalanced cohort size, a Japanese study of 14 MBC and

140 FBC showed no difference in overall survival [37]. This

was also reflected in a UK study comparing outcome in 41

MBC and 123 FBC which showed that when matched for

key prognostic factors (size, grade and lymph node status),

outcome was similar between genders [38], agreeing with

our study. Of note was the observation that when MBC was

compared with an unselected FBC group, males had worse

outcome [38], which may explain some of the earlier studies

inferring a worse prognosis in men [39, 40]. Gender com-

parative information obtained from 1988 to 2003 SEER data

showed worse breast cancer-specific survival in males

diagnosed with stage I disease; however, the authors

attributed this to in-stage migration rather than being of

clinical relevance [41].

A common finding in MBC is the higher frequency of

hormone receptor expression in particular ERa (reviewed

in [42]), which was also reflected in this study. We have

explored this further using hierarchical clustering where

one of the striking features was the occurrence of ERa on

distinct clusters in males and females. In FBC, ERa clus-

tered with PR and its isoforms; as PR is oestrogen-regu-

lated [43], this is unsurprising. In MBC, the position of

ERa changed, clustering instead with ERb isoforms and

AR. The potential role of ERb in breast cancer has been the

subject of much debate and recent work in FBC by us and

others shows this depends on the cell location and the type

of isoforms expressed [9, 44]. However, ERb is subject to

complex regulation involving 30UTRs [45] and microRNAs

[46], and we need to further understand its biology before

speculating on any role it may play in MBC.

Regarding the potential relationship of AR with ERa, at

a functional level, AR transfection into ERa-positive breast

cancer cells inhibited ERa transactivation and oestrogen-

stimulated growth through interaction with oestrogen

response elements [47]. Other work indicates that oestro-

gen activation via ERa and ERb can mediate AR signalling

[48]. Given the recognised pro-proliferative effects of ERaand PR and the anti-proliferative effects of ERb and AR

[49] this suggests the coordinated expression of these

receptors could influence survival. This was demonstrated

Table 3 Expression of AR, ERb, and PR isoforms in male and

female luminal A breast carcinoma

Biomarker Negative

(%)

Positive

(%)

P value OR (95% CI)

AR

Male 86 113 \0.0001 0.12 (0.06–0.208)

Female 16 181

PRA

Male 47 152 \0.0001 3.47 (2.25–5.34)

Female 102 95

PRB

Male 79 120 0.486 1.18 (0.80–1.76)

Female 86 111

ERb1 (nuc)

Male 77 122 \0.0001 0.14 (0.08–0.25)

Female 16 181

ERb1 (cyto)a

Male 26 157 \0.0001 22.34 (12.99–38.43)

Female 148 40

ERb2 (nuc)

Male 38 161 0.36 1.29 (0.79–2.09)

Female 46 151

ERb2 (cyto)b

Male 63 117 \0.0001 2.50 (1.64–3.80)

Female 109 81

ERb5 (nuc)

Male 49 150 0.20 0.71 (0.44–1.15)

Female 37 160

Nuc nuclear expression, cyto cytoplasmic expressiona 16 male and 9 female and b 19 male and 7 female cases were

unscoreable due to core loss/exhaustion


123

0 10 20 30 40 50 600

25

50

75

100

P = 0.09

MonthsP

erce

nt

surv

ival

0 20 40 60 80 100 1200

25

50

75

100

P = 0.42

Months

Per

cen

t su

rviv

al

0 10 20 30 40 50 600

25

50

75

100

P = 0.01

Months

Per

cen

t su

rviv

al

0 20 40 60 80 100 1200

25

50

75

100

P = 0.29

Months

Per

cen

t su

rviv

al

0 10 20 30 40 50 600

25

50

75

100

P = 0.04

Months

Per

cen

t su

rviv

al

0 20 40 60 80 100 1200

25

50

75

100

P = 0.04

Months

Per

cen

t su

rviv

al

0 10 20 30 40 50 600

25

50

75

100

P = 0.05

Months

Per

cen

t su

rviv

al

0 20 40 60 80 100 1200

25

50

75

100

P = 0.37

Months

Per

cen

t su

rviv

al

a b

c

e

g

d

f

h

15/119

47/185

25/119

69/177

7/79

46/178

16/77

67/170

7/75

44/182

17/80

69/193

5/57

6/27

17/80

8/22

Fig. 2 Kaplan–Meier survival

curves comparing survival

between genders according to

different tumour classifications.

No gender-related differences

were observed comparing OS of

MBC (blue line) and FBC (pinkline) at 5 (a) and 10 (b) years.

Comparison of OS of Luminal

A AR-positive MBC (blue line)

and FBC (pink line) showed

MBC had significantly

improved OS at 5 (c) but not 10

(d) years. In unselected ERa-

positive MBC those which were

AR positive (black line) had

significantly improved over

those who were AR negative

(orange line) at both 5 (e) and

10 (f) years. Comparison of OS

of unselected ERa and AR-

positive carcinomas showed

MBC (blue line) had

significantly improved survival

over FBC (pink line) at 5 (g) but

not 10 (h) years


123

in the current study where AR-positive luminal A MBC

had improved overall survival than the equivalent FBC and

was also borne out in the unselected group. This supports

other recent work showing AR is an important prognostic

factor in ERa-positive FBC [12]. Previous studies exam-

ining the impact of AR in MBC have been compounded by

the high expression frequency of AR in some studies [22],

the low number of cases in others [50], or combinations of

both [51, 52] precluding meaningful survival analysis.

Studies examining the effect of AR on survival in MBC are

contradictory, possibly as a result of limited numbers

available from single centre studies. In a series of 43 MBC

Kwiatowska et al. [53] showed AR expression correlated

with reduced survival. A similar-sized study (n = 47)

showed no effects of AR expression in MBC on survival,

however, it is noteworthy that in the study, MIB-1

expression, which detects proliferating cells, was higher in

AR negative compared to AR-positive cases [26]. The

association of AR positivity with better outcome in MBC

in this study indicates this is a potentially important

prognostic factor, paralleling observations in FBC where

the potential prognostic role for AR in FBC is receiving

increased attention [54] following the observation that AR

is an independent prognostic marker in a large series of

FBCs (n = 953; [12]). Moreover, AR expression in MBC

could turn out to have both prognostic and predictive value;

its presence suggests that anti-androgen therapy could be

explored as a therapeutic approach. Androgen blockade is

commonly used in prostate cancer treatment but so far

remains inadequately tested in breast cancer [55]. The

complete absence of cell line models derived from MBCs

presents a challenge in being able to model this in vitro.

To our knowledge, this is the largest retrospective bio-

marker study directly comparing matched male and female

breast carcinoma; however, we acknowledge there are

some limitations. Despite our best efforts, these include

lack of availability of follow up data and missing/

incomplete clinicopathological data. Of note is the absence

of data on germline mutations, particularly BRCA2 which

is involved in MBC development and associated with

reduced survival [53, 56, 57]. Nevertheless, our study has

confirmed that whilst superficially similar to FBC, when

studied more rigorously MBC is biologically different,

echoing a hypothesis proposed by the Multidisciplinary

Meeting on MBC [58] and supporting recent work at the

transcriptional [59], microRNA [60, 61] and genomic [62]

levels. With the incidence of MBC rising [4–7], collec-

tively these studies provide a strong impetus for further

study of this rare cancer type.

Acknowledgments Thanks to the Tayside Tissue Bank for kindly

providing some of the MBC cases. This study was supported by the

Breast Cancer Campaign (UK Charity no. 05074725).

Conflict of interest None.

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a comparative biomarker study of 514 matched cases of male and female breast cancer reveals...

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