The transcription factor
E2F3 is overexpressed in prostate cancer and independently predicts
Authors: Sandra Edwards,
The treatment and management
of prostate cancer needs to be improved. A particular problem
that exists is that it is not possible to predict how early prostate
cancer still localised to the prostate (for example detected by
the PSA test) will behave. Some cases may remain dormant for many
years without progressing while others will progress rapidly to
malignancy. The particular goal of our work is to identify markers
that can be used to distinguish dormant from aggressive early prostate
cancers. This is important because such markers can be used to
identify aggressive cancers early so that they may be treated and
to minimise treatment for dormant tumours.
We have recently demonstrated
that the E2F3 transcription factor gene, which has an
established role in controlling transition through the cell cycle,
is a bladder cancer oncogene that is activated by amplification
and overexpression 1 . We have investigated the role of E2F3
in prostate cancer by immunohistochemistry on prostate tissue
Tissue microarrays (TMAs)
were constructed from TURP and prostatectomy samples that had been
taken from a consecutive series of patients (774 cores from 147
patients) diagnosed with prostate cancer who attended the Royal
Marsden NHS Trust from 1992. Immunohistochemistry (Figure 1) was
carried out using a monoclonal antibody to E2F3 (Upstate, UK).
1. E2F3 expression detected by immunohistochemistry in
formalin fixed tissue in TMAs. (a-c) Primary prostate cancers. Examples
of tumours scored as (a, b) positive and (c) negative are shown.
(d) Example of a hyperplasia scored as positive for E2F3. (e) Example
of morphologically normal epithelia scored positive for E2F3. (f)
Example of cytoplasmic staining observed in morphologically normal
epithelium. Staining: brown, E2F3; blue, haematoxylin counterstain
Analysis of the TMAs demonstrated
intense nuclei staining for E2F3 in 67% (98/147) of prostate cancers
(Figure 1a-c), with the maximum proportion of cells containing nuclear
staining varying from 5 to 90%. Analysis of non-neoplastic epithelium
samples revealed that hyperplastic epithelium (Figure 1d) and morphologically
normal epithelium (Figure 1e) exhibited nuclear E2F3 staining in
respectively 21/124 (19.8%) and 1/43 (2.33%) of cases. Because of
the wide variation in the percentage of nuclei exhibiting staining,
the data were stratified into six bands: negative, up to 20, 21-40,
41-60, 61-80 and 81-100%. In some cases, E2F3 was observed predominantly
in the cytoplasm (Figure 1f). This pattern was not observed in prostate
cancer and was restricted entirely to morphologically normal epithelium
(3/43) and epithelia hyperplasia (15/124).
Studies of clinical correlation
showed a significant association between the presence of nuclear
E2F3 staining and Gleason score (P=0.016), but no association with
age (P=0.327), M stage (P=0.574) or AJCC 2 stage (P=0.613)
Figure 2 shows overall survival
and cause-specific survival according to E2F3 status. These curves
demonstrated significantly increased hazard ratios for patients
whose cancers exhibited E2F3 expression for both overall survival
(log-rank test, df=1, P=0.0022) and cause specific survival (log-rank
test, df=1, P=0.0047) of 1.9290 and 1.8423respectively.
Kaplan-Meier analysis of cause-specific survival in prostate cancer
patients that are positive (red) and negative (black) for E2F3 nuclear
staining. Vertical lines are error bars.
Figure 3 shows overall and
cause-specific survival following stratification of patients according
to the maximum percentage of E2F3-positive nuclei observed in their
prostate cancer. These analyses demonstrated a significant association
between the percentage of nuclear staining for E2F3 and risk of
death for both overall (logrank test, df=5, P=0.0014) and cause-specific
(logrank test, df=5, P=0.004) survival. Importantly, in multivariate
analysis, E2F3 staining was an independent predictor of clinical
Figure 3. Kaplan-Meier analysis of cause specific
survival stratified E2F3 data. Prostate cancer patients were stratified
into six bands according to the maximum percentage nuclear staining
positive for E2F3 in TMA cores: negative (black), up to 20 (red),
21-40 (blue), 41-60 (green), 61-80 (pink) and 81-100% (brown).
Several lines of evidence
now support the view that the pRB-E2F3-EZH2 pathway may represent
a key oncogenic axis that has an important role in determining development
and aggressiveness of
human prostate cancer (Figure
Role of the pRB-E2F3-EZH2 cell cycle control
axis in determining aggressiveness in human prostate cancer. The
INK4A/ARF gene encodes two proteins p16 and p14ARF that are negative
regulators of the pRB-E2F3-EZH2 pathway. Genes overexpressed (red)
or downregulated (green) in human prostate cancer are indicated.
Red stripes indicate that overexpression of p53 measured in studies
of clinical correlation was believed to represent underlying mutations
in the p53 gene.
signifies that the indicated change in gene expression is associated
with adverse clinical outcome
The E2F3 protein has an
established role in controlling cell cycle progression and controls
the expression of EZH2 which has also recently been implicated in
the development of human prostate cancer. Several other alterations
of the expression levels of genes in this pathway have also been
connected with poor clinical outcome (Figure 4).
In conclusion, we demonstrate
aberrant high-level overexpression of E2F3 in 67% of prostate cancer
and that overexpression of this protein is an independent predictor
of poor clinical outcome in prostate cancer 3 . Importantly, the
higher the level of E2F3 protein, the worse the prognosis.
1. Feber A et al.
2. Prostate. In: American
Joint Committee on Cancer: AJCC Cancer Staging Manual.
Philadelphia, Pa: Lippincott-Raven
Publishers, 5th ed., 1997, pp 219-224
3. Foster CS et al.