2016, Volume 32, Number 1, Page(s) 008-014
The Relation of Parafibromin Expression with Clinicopathological Factors in Invasive Breast Carcinomas
Serap KARAARSLAN1, Berhan GENÇ2, Ahmet NART3, İsmail BÖREKÇİ3, Mehmet Hüsnü BUĞDAYCI1
1Department of Pathology, Şifa University Faculty of Medicine, İZMİR, Turkey
2Department of Radiology Şifa University Faculty of Medicine, İZMİR, Turkey
3Department of General Surgery Şifa University Faculty of Medicine, İZMİR, Turkey
Keywords: Breast carcinoma, Immunohistochemistry, Lymph nodes, Metastasis, Parafibromin
We aimed to find the relationship between parafibromin expression and clinicopathologic variables of breast carcinoma.
Material and Method: Ninety-seven cases of invasive breast carcinoma diagnosed at our department between the years 2010–2013 were included in the study. The parafibromin expression state was compared with the estrogen receptor, progesterone receptor, cerbB2, Ki67 results, and the clinicopathological variables.
Results: Among 97 breast carcinoma cases, 66 (68%) were invasive ductal carcinoma. The average age was 54.3 (min:25, max:100), and the average tumor size was 31.1 mm (min:7, max:120). Lymph node metastasis was detected in 58% of the cases. Eleven were diagnosed with metastasis amongst 77 cases whose distant metastasis data could be reached. Eleven cases were lost due to breast carcinoma. As the tumor grade increased, the possibility of distant metastasis and lymph node metastasis increased as well (p=0.04, p=0.05, respectively). The mean follow-up duration of the cases was 26.6}9.8 (min. 6, max. 53) months, and there was no significant difference in survival between the other variables. Of the cases, 21.6% were negative, 9.3% were (+) positive, 11.3% were (++) positive and 57.7% were (+++) positive for parafibromin. It was found that there was an inverse correlation between the Ki67 proliferation index and lymph node metastasis and the parafibromin expression (p=0.018, p=0.029, respectively).
Conclusion: We suggest that parafibromin may be a possible prognostic and predictive parameter for breast carcinomas. As the data on this matter in the literature is limited, it would be beneficial to investigate the matter and evaluate its relationship with survival in larger series.
Breast cancer (BC) is the most common diagnosed
malignancy in females globally. It ranks second amongst
cancer-related deaths after lung cancer. BCs have a wide
range of morphological and molecular features1
Therefore, the treatment is planned out according to the
patient’s clinical features and the tumor’s pathological
and molecular features. Age, tumor size, histological
grade and lymph node status are the most significant
identifiers that are used to predict the clinical course in
the diagnosis period of BCs. Having a high proliferation
index is accepted as a worse prognostic factor2
. The most
important indicators that shape both pre and post-surgery
treatment are the estrogen receptor (ER), the progesterone
receptor (PR) and the human epidermal growth factor
receptor (HER-2/cerbB2). According to these features,
hormone therapy (tamoxifen and aromatase inhibitors),
trastuzumab, the recombinant human anti-HER2 antidote
(Herceptin®Genentech, California,USA), and lapatinib,
the dual HER1/HER2 tyrosine kinase inhibitor (Tykerb,
GlaxoSmithKline, Philadelphia,USA), can be used3-5
Parafibromin is a new marker that has initially been used
in the definitive diagnosis of parathyroid tumors. CDC73/
Hyperparathyroidism type 2 (HRPT2) is a suppressor
gene (chromosome 1q25-q31) and codes parafibromin
that contains 531 amino acids. Parafibromin is part of the
polymerase-associated factor 1 (PAF1) / RNA polymerase
II complex, which is critical for histone modification.
Inactivity (germ-line mutations / somatic mutations) of
the HRPT2 tumor suppressor gene is associated with the
Hereditary Hyperparathyroidism-jaw syndrome (HHJTS)
and sporadic parathyroid carcinoma. While the expression
decreases and disappears in parathyroid carcinomas,
the expression in sporadic parathyroid adenoma and
hyperplasia is high6-9. Due to the fact that it is a tumor
suppressor gene, it could be predicted that the malignancy
potential and aggressiveness increases as the parafibromin
expression decreases. The first study on this subject in the
literature was published by Selvarajan et al. in the year 200810. No data in the English literature existed before that
In our study, both the features of parafibromin staining and
its relation with the prognostic variables were investigated
and the parafibromin staining features of the peritumoral
normal breast parenchyma were evaluated.
|The selection of the patients and tissue:
97 BC’s, diagnosed between the years 2010 – 2013 in the
University Pathology laboratory, were included in the
study. By selecting the blocks with tumor and normal breast
parenchyma, it was aimed to evaluate the parafibromin
expression of the peritumoral breast tissue as well as the
carcinoma regions. Parafibromin, ER, PR, cerbB2 and Ki67
were immunohistochemically (IHC) applied to the slides
of all selected blocks. The clinical features (recurrence,
metastasis, death from disease) of the patients were
acquired from the patient records. Pathological features
(tumor grade, tumor subtype, lymph node status, perinodal
invasion) were re-evaluated according to the standard
Immunohistochemical staining: The 4 micron-thick
sections were taken from the paraffin-embedded blocks of
all the cases and put on the positively-charged slides and
stained with parafibromin (sc-33638, dilution 1/100, Santa
Cruz Bio-technology, USA), ER (IR084, ready to use, Dako,
Denmark), PR (IR068, ready to use, Dako, Denmark),
cerbB2 (A0485, dilution 1/400, Dako, Denmark) and
Ki67 (IR626, ready to use, Dako, Denmark). After the
deparaffinization of the sections, the antigen revelation
was done by PT Link, Dako; staining was performed by
the DAKO Autostainer Link 48. Normal parathyroid tissue
was used as a positive control while the vascular structures
in the breast stroma and the fibroblastic cells were used as
a negative control for parafibromin. The cases that were
evaluated by two pathologists (SK, MHB) as different in
staining ratios, were re-evaluated together and the codecision
used in statistical calculations.
The evaluation variables of the immunohistochemical
staining: The cut-off data from previous research were
used to evaluate the percentage and features of the
parafibromin staining on the tumor cell nuclei, regardless
of staining intensity. In this study, 0–5%, 5–25%, 26–50%,
51-100% were accepted as negative, (+), (++) and (+++)
positive respectively13. The positive rate of ER and PR
was evaluated as the percentage of staining in tumor cells.
At least 1% rate in tumor tissue staining was considered as
positive14,15. For the evaluation of the Ki67 proliferation
index, the surface of the entire section was homogenized by
scanning the stain and three areas were chosen to reflect the overall staining percentage. At least 400 cells were
counted specifically for the percentage of staining in these
areas with high magnification (x400). The 14% value that
has been stated in the article by Cheang et al. was used as a
Statistical data analysis: The SPSS (Statistical Package
for Social Sciences) Windows 18.0 (SPSS Inc., Chicago,
USA) software program was used for statistical analyses. In
addition to descriptive statistical methods, the Chi Square
Test and Fisher’s Exact Test were used in the comparison
of categorical variables. Survival data was obtained with the
Kaplan Meier, Log Rank and Cox Regression analyses. A p
value <0.05 was accepted as statistically significant.
The clinical and pathological features of the cases are
shown in Table I
. Most of the invasive ductal carcinoma
cases were grade 2 or 3. As the tumor grade increased, the
axillary lymph node metastasis (LNM) rate increased as well
(p=0.04). The relation between the increased tumor grade
and distant metastasis was found to be statistically marginsignificant
(p=0.05) (Table II
). Eleven cases were diagnosed
with distant metastasis among 77 cases whose metastasis
data could be reached. The metastasis region was found to
be the bone (n=6), liver (n=2), bone and liver (n=1), thyroid
(n=1) and distant lymph nodes (n=1) in order of frequency.
The mean follow-up time of the cases was 26.6±9.8 (min.
6, max. 53) months. Eleven patients died due to BC related
causes. Mean overall survival time was 48.23±1.5 months.
There was no significant difference in overall survival time
according to age, tumor size, tumor type, tumor grade,
LNM, perinodal invasion and distant metastasis.
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|Table II: The relationship between tumor grade and distant
metastasis, lymph node metastasis
The parafibromin expression was evaluated in the breast
carcinoma field and the fields that included fibrocystic
changes and ordinary breast parenchyma. Parafibromin
was identified to be (+++) positive in the ordinary breast
parenchyma (Figure 1A). (++)/(+++) positivity was
also present with the sclerosing adenosis and fibrocystic
changes as well (Figure 1B,C). Parafibromin (++) and
(+++) positivity was discovered in the areas where the
high-grade ductal carcinoma (n=8) was located (Figure
1D). The details about the parafibromin staining of the
breast carcinomas are shown in the table and pictures
(Figure 2A–E, Table I). The inverse correlation between
the parafibromin expression and Ki67 proliferation index
and LNM was remarkable (p=0.018, p=0.029, respectively)
(Table III). No statistically significant relationship between
parafibromin expression and age, tumor size, tumor type,
tumor grade, perinodal invasion, ER, PR, and CerbB2 was found. There was no significant difference in overall
survival time according to the parafibromin expression, the
Ki67 value (p=0.257, p=0.072, respectively), ER, PR and
Click Here to Zoom
|Figure 1: A) The strong positivity of the parafibromin in the ordinary breast parenchyma (parafibromin; x50). B, C) (+++) parafibromin
positivity in fibrocystic changes and sclerosing adenosis (parafibromin; x100). D) (+++) parafibromin positivity in the high-grade
invasive ductal carcinoma (parafibromin; x200).
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|Figure 2: A) The (+++) positivity of the parafibromin in invasive
ductal carcinoma morphology in tumor regions (parafibromin;
x100). B) The (+++) positivity of the parafibromin in invasive
lobular carcinoma regions and the (+++) parafibromin positivity in
the surrounding ordinary breast ducts (parafibromin; x50). C) The
(++) parafibromin positivity in breast carcinoma (parafibromin;
x100). D) The (+) parafibromin positivity in invasive ductal
carcinoma regions (parafibromin; x50). E) The parafibromin
negativity in invasive ductal carcinoma regions (parafibromin;
Click Here to Zoom
|Table III: The relation between parafibromin expression and Ki67, lymph node metastasis
Parafibromin was initially used in the differential diagnoses
of the parathyroid tumors. Parafibromin was negative
with HPT-JT related tumors and sporadic parathyroid
carcinomas, but expression in hyperplasia and sporadic
adenoma was encountered6,8
Studies on parafibromin expression in other organ
malignancies were then performed13. The only article in
the literature about the features of parafibromin staining
in breast carcinomas is the study performed by Selvarajan
et al. in 200810. Parafibromin expression was evaluated
with the micro array method in 163 breast carcinomas.
Parafibromin staining features were compared with tumor
size, tumor grade, lymphovascular invasion and cerbB2
expression. An inverse correlation between tumor size
and parafibromin expression was observed (p=0.05). In
addition, parafibromin was reported to be negative in the
tumors with aggressive histopathological features (high
grade, lymphovascular invasion and cerbB2 overexpression)10. In our study, although no relationship was found
between the parafibromin expression and tumor size,
tumor grade, ER, PR and cerbB2 expression, a significant
relationship between parafibromin expression and the Ki67
proliferation index and LNM was detected. In cases with
increased parafibromin expression, the Ki67 proliferation
index was lower and the number of LNM was less. These
features suggested that there may be a relationship between
low parafibromin expression and tumor aggressiveness.
LNM is a parameter used to predict the clinical course in the
breast carcinomas17. In a study on stomach carcinoma by
Zheng et al., it was reported that parafibromin expression
decreases incrementally from the normal stomach mucosa to carcinoma. In our study, parafibromin was positive
in normal breast parenchyma, fibrocystic regions and
sclerosing adenosis but sufficient data to compare it with
the carcinoma could not be retrieved. In the same study, a relationship between lymphatic invasion, invasion
depth, LNM and tumor stage were detected. As a result,
it was reported that it could predict an aggressive course
in stomach carcinoma13. Since a relationship between
LNM and parafibromin expression loss was detected in
our study, it could be suggested that loss of parafibromin
expression could be related to tumor aggressiveness. Our
study detected a significant relationship with decreasing
parafibromin expression and LNM. In the cases that
contained LNM (n=56), 33 perinodal invasions had been
seen, and in the cases that showed perinodal invasion, seven
distant metastasis (n=11) had been detected. In addition,
eight of the 11 distant metastatic cases contained LNM.
Even though there is no statistically significant difference
between LNM and other parameters, the parallelism of
the data may provide support for the aggressive behavior
Ki67 proliferation index is a criterion used in the grading
systems and it should be correlated with the other variables.
It was reported that cases with high Ki67 proliferation
index got worse than all the other groups, except for those
who had not received treatment and the lymph nodenegative
group5. A statistically significant relationship
was detected between parafibromin expression loss and
increasing Ki67 proliferation index in our study. The Ki67
value was above 14% in the 8 cases among 11 cases who
died. Thus, it was considered that parafibromin expression
loss might predict a worse clinical course.
In conclusion, we encountered relationships with different
variables compared to the first study on parafibromin
expression in breast cancer. In some cases it is possible to
make a prediction even with the morphological criteria,
but various markers can be used other than the standard
markers in some BC where a decision cannot be made. In
our review, although no relationship was found between
parafibromin expression and the tumor size, tumor
grade, ER, PR and cerbB2 expression, a significant inverse
relationship between the Ki67 proliferation index and LNM
was detected. In the cases with increased parafibromin
expression, the Ki67 proliferation index was lower and the
number of LNM was less. These features suggest that there
may be a relationship between decreased parafibromin
expression and tumor aggressiveness, and supports further
evaluation with larger samples.
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