Investigating the clinical potential for 14-3-3 zeta protein to serve as a biomarker for epithelial ovarian cancer
Journal of Ovarian Research
Investigating the clinical potential for 14-3-3 zeta protein to serve as a biomarker for epithelial ovarian cancer
Ioannis Hatzipetros 0 1
Peter Gocze 1 3
Tamas Koszegi 1 2
Akos Jaray 1 5
Laszlo Szereday 1 4
Beata Polgar 1 5
Nelli Farkas 1 4
Balint Farkas 0 1
0 Department of Obstetrics and Gynecology, University of Pecs, Clinical Center , Edesanyak Str. 17, 7624 Pecs , Hungary
1 Setting: University of Pecs Medical Center Department of Obstetrics and Gynecology/Oncology (Pecs , Hungary)
2 Department of Radiology, University of Pecs , Hungary
3 Institute of Laboratory Medicine, University of Pecs , Hungary
4 Institute of Bioanalysis, University of Pecs , Hungary
5 Department of Medical Microbiology and Immunology, University of Pecs , Hungary
Objective: Recently, 14-3-3 zeta protein was identified as a potential serum biomarker of epithelial ovarian cancer (EOC). The goal of this study was to investigate the clinical potential of 14-3-3 zeta protein for monitoring EOC progression compared with CA-125 and HE4. Design: Prospective follow-up study. Population: Thirteen EOC patients with advanced stage (FIGO IIb-IIIc) epithelial ovarian cancer that underwent radical surgery and received six consecutive cycles of first line chemotherapy (paclitaxel, carboplatin) in 21-day intervals. Methods: Pre- and post-chemotherapy computed tomography (CT) scans were performed. Serum levels of CA-125, HE4, and 14-3-3 zeta protein were detected by enzyme-linked immunosorbent assay (ELISA) and quantitative electrochemiluminescence assay (ECLIA). Main outcome measures: Serum levels of CA-125, HE4, and 14-3-3 zeta protein, as well as lesion size according to pre- and post-chemotherapy CT scans. Results: Serum levels of CA-125 and HE4 were found to significantly decrease following chemotherapy, and this was consistent with the decrease in lesion size detected post-chemotherapy. In contrast, 14-3-3 zeta protein levels did not significantly differ in healthy postmenopausal patients versus EOC patients. Conclusions: Determination of CA-125 and HE4 serum levels for the determination of the risk of ovarian malignancy algorithm (ROMA) represents a useful tool for the prediction of chemotherapy efficacy for EOC patients. However, levels of 14-3-3 zeta protein were not found to vary significantly as a consequence of treatment. Therefore we question if 14-3-3 zeta protein is a reliable biomarker, which correlates with the clinical behavior of EOC.
Epithelial ovarian cancer; 14-3-3 Zeta protein; CA-125; HE-4; Chemotherapy; ROMA
Epithelial ovarian cancer (EOC) is a highly malignant
gynecological neoplasia with an incidence of 12/100 000
women , and this rate has only slightly decreased in
the last 80 years. While women of any age are at risk for
this malignancy, postmenopausal women have a higher
incidence. For example, 90% of women who suffer from
EOC are older than 40 years of age, and the greatest
number are 55 years or older. Moreover, due to the
anatomic position of the ovaries, pelvic malignancies can
remain obscured. In addition, a lack of symptoms until the
advanced stages of tumor development results in an
increased rate of metastasis at the time of diagnosis.
Correspondingly, 80% of the ovarian neoplasias diagnosed
are stage III-IV according to International Federation of
Obstetrics and Gynecology (FIGO) criteria. While FIGO
stage I EOC has a relatively high five-year survival rate
(> 90%), survival rates markedly drop for patients with
stage III-IV EOC (2530%) [2,3]. Therefore, it is crucial
to diagnose EOC as early as possible. Accordingly, the
identification of serum biomarkers to detect EOC would
represent an important and valuable advance for the
monitoring and treatment of EOC progression.
Algorithms and triage protocols designed to evaluate
potential cases of ovarian cancer in their early stages are
currently limited, and rely on pelvic sonography and
CA-125 determination . Moreover, the sensitivity and
specificity of these approaches range from 7080%. .
Regarding CA-125, its levels are elevated in less than 50%
of EOC cases, and it is undetectable in another 20% of
EOC cases. In addition, high serum levels of CA-125 are
also associated with benign gynecological diseases (e.g.,
cysts, endometriosis, etc.) . In 2008, Moore and
colleagues identified human epididymis protein 4 (HE4) as a
biomarker for ovarian cancer . Based on these findings,
a risk of ovarian malignancy algorithm (ROMA) was
developed, and is currently used to predict the presence of
malignant ovarian cancer using a combination of CA-125
levels, HE4 expression, and menopausal status. In
particular, the combination of HE4 and CA-125 in the ROMA
has been associated with a higher sensitivity than any
single biomarker .
14-3-3 zeta is an important regulatory protein, which
mediates intracellular signaling pathways by interfering
with approximately 100 cellular proteins, including
oncogenes and protooncogenes. Recently, two independent
research groups, Waldemarson et al.  and He et al.
, advocated 14-3-3 zeta as a potential biomarker for
EOC. In addition, Kobayashi et al.  recently
demonstrated that 14-3-3 zeta protein is present in malignant
ascites of patients with EOC, and is secreted by ascetic
monocytes and macrophage. However, while the role of
14-3-3 zeta protein as an intracellular adaptor protein
has been widely investigated, the function of the secreted
protein is unclear. Therefore, the goal of the current
pilot study was to assess the potential for 14-3-3 zeta
protein to serve as a biomarker for monitoring patients
with FIGO stage II-III EOC that undergo chemotherapy.
Materials and methods
Patients and follow-up study design
This prospective study was approved by the University
of Pecs Institutional Ethical Review Board
(#4076.316251/KK15/2011), and written informed consent was
obtained from all enrolled patients.
Peripheral blood samples were collected preoperatively
from 13 patients admitted for six cycles of first line
chemotherapy (paclitaxel/carboplatin; Hungarian OEP
Chemotherapy protocol # 7167). Chemotherapy dosage
was calculated based on body mass (kg), and treatments
were administered in 21 d intervals at the University of
Pecs Medical Center Department of Obstetrics and
Gynecology/Oncology (Pecs, Hungary) in 2012. Blood
samples were collected 12 h before each treatment
into citrate tubes. These tubes were then centrifuged
(5000 rpm for 10 min), and blood plasma samples were
collected and stored at 80C. When needed, samples
were thawed at room temperature (RT), and then were
thoroughly vortexed as indicated by the manufacturers
recommendation. All patients, aged 41 to 73 y (mean,
60 y), underwent radical gynecological surgery for the
removal of both adnexes with or without the uterus.
However, peritoneal or lymph node metastases were not
resected. Computed tomography (CT) scans were
performed prior to and following the completion of
chemotherapy treatment. These images were used to assess
changes in both target and non-target lesions. Each
diagnosis was verified according to histopathology
studies of the original tumors. Histopathological grade and
stage of disease (according to FIGO criteria) were
available for all malignant cases, and included FIGO stage
IIa (n = 2), stage IIIb (n = 2), and stage IIIc (n = 9) cases
Computed tomography (CT) scans
CT scans were performed in the Department of Radiology
(University of Pecs, Hungary). Contiguous 5 mm axial
slices obtained through the abdomen and pelvis. Prior to
examination high-concentration iodinated contrast agent
was administered intravenously (Iomeron 400, Bracco
Diagnostic Imaging). Field-of-view was adjusted to body
habitus (to include the whole body including the skin).
Target and non-target lesions were defined based on
RECIST 1.1 guidelines (www.recist.com). A lesion was
measurable and defined as a target lesion if the tumor
was 10 mm along its longest diameter (LD) on a CT axial
image with 5 mm reconstruction intervals, or if lymph
nodes were 15 mm along their short axis on CT images.
Table 1 Clinicopathological features of the patients
enrolled in this study that underwent six cycles of
paclitaxel/carboplatin-based chemotherapy for treatment
III C 69 High 2 3 4
Adenosquamous II B
Adenosquamous III C
III C 57 41 50
Non-target lesions were considered to be: masses with a
diameter < 10 mm, lymph nodes with a diameter of 10
14 mm along the short axis, ascites, pleural or pericardial
effusion, abdominal masses, or organomegaly identified by
physical exam. Furthermore, these could not be measured
by reproducible imaging techniques. CT scans were
performed 12 weeks after radical surgeries were performed,
12 weeks prior to chemotherapy, and 12 weeks after
the final chemotherapy treatment.
Enzyme-linked immunosorbent assay (ELISA)
Levels of serum CA-125 (Fujirebio Diagnostics, Malvern,
PA; Catalog #: 40010, Lot. # 29191), HE4 (Fujirebio
Diagnostics, Malvern, PA; Catalog #: 40410, Lot# 28373), and
14-3-3 zeta protein (Cusabio Biotech, Wuhan, China;
Catalog #: CSB-EL026293HU, Lot. #A26174460) were
determined using a quantitative sandwich enzyme
immunoassay according to each manufacturers protocol. Serum
concentrations were calculated using Optima 2.10 R2
built-in data calculator software.
Quantitative electrochemiluminescence assay (ECLIA)
Tumor marker levels were measured using a Roche
electrochemiluminescent fully automated immunoassay system
(ECLIA, Roche Diagnostics, http://www.roche-diagnostics.
com). To determine serum levels of CA-125 (Cat. no.
11776223), and HE4 (Cat. no. 05950929), samples were
processed using a Roche Cobas e411 analyzer. Master
calibration, imprecision, and inaccuracy were checked using
bi-level quality controls prior to the analysis of patient
Risk for ovarian malignancy algorithm (ROMA) index
The ROMA used serum levels of HE4 and CA-125
measured either by ELISA or ECLIA, and was calculated using
an Excel spreadsheet with preset formulas to generate the
predictive index (PI) value for EOC  as follows:
For postmenopausal women: PI = 8.09 + 1.04*LN
[HE4] + 0.732*LN [CA125].
A ROMA value was then calculated as follows: ROMA
value (%) = exp(PI) / [1 + exp(PI)]*100. According to the
manufacturers manual, the detection of HE4 by ECLIA
and CA-125 by ELISA in menopausal women identified
an EOC high-risk index value equal to, or higher than,
Statistical analyses were performed using IBM SPSS
Statistic 20 (IBM Corporation) at the University of Pecs,
Institute of Bioanalysis. The sample size (n) was 13, and
comparisons were made between treatments and between
methods according to the Wilcoxon signed-rank test. To
evaluate trends between the number of treatments and
serum levels of tumor markers, linear regression and
correlation analyses were applied. To examine the
relationship between tumor marker levels and CT scan results,
Spearmans rank correlation coefficient was used. Mean
data are reported standard error of the mean (SEM).
Statistical significance was set at p < 0.05, or p < 0.1.
Radiologic assessment following therapeutic procedures
CT scans were obtained one to two weeks after radical
gynecological surgeries were performed. After an initial
laparotomy, 10/13 (76.92%) patients were found to have
residual tumor present prior to induction of
paclitaxel/carboplatin-based chemotherapy. After six consecutive cycles of
treatment within 21 d intervals, CT scans were repeated.
At this point, residual tumor with a LD value >1 cm was
only detected in 4/13 (30.76%) patients (Table 2). Based on
the detection of 26 non-target lesions in pre-chemotherapy
CT scans, and only 3 non-target lesions in
postchemotherapy scans, the efficacy of chemotherapy for EOC
treatment is demonstrated (Figure 1).
Detection of CA-125, HE4, 14-3-3 zeta protein
Levels of CA-125, HE4, and 14-3-3 zeta protein were
monitored throughout the treatment period by ELISA
and ECLIA. On the first day of chemotherapy, the mean
concentration of CA-125 was 147.87 55.98 U/ml and
648.26 186.52 U/ml, respectively. After completing the
sixth cycle of chemotherapy, CA-125 levels were lower,
with the mean concentrations detected being 58.54
SLD: sum of length diameter; according to RECIST 1.1 guidelines.
*Negative values represent a decrease in lesion size.
Table 2 Evaluation of tumor size before and after
paclitaxel/carboplatin chemotherapy using CT scans
SLD of target lesions
10 (+/ 4) d before
SLD of target lesions
Non10 (+/ 4) d after target
the last cycle of lesion
chemotherapy (mm) changes*
30.89 U/ml and 119.70 22.75 U/ml, respectively.
Similarly, mean serum levels of HE4 detected on the first day
of chemotherapy by ELISA were 455.32 106.39 pM,
and decreased to 120.52 23.76 pM upon completion of
chemotherapy. Using the ECLIA method, serum levels
of HE4 were 1383.49 577.23 pM on the first day of
chemotherapy, and decreased to 70.12 26.44 pM upon
completion of chemotherapy. ROMA index values were
subsequently calculated, and decreased from 58.17
10.05% to 28.95 7.67%, and from 69.62 9.91% to
30.78 7.91% for ELISA and ECLIA, respectively.
According to Wilcoxon statistical analyses, the differences
in the values determined at the start of treatment versus
upon completion of treatment were significant (p < 0.05),
thus further demonstrating the effectiveness of
chemotherapy for EOC (Figure 2/A-F, Table 3).
For 14-3-3 zeta protein levels detected in patients prior
to chemotherapy by ELISA, the mean concentration was
1.93 0.57 ng/ml. In contrast, the mean concentration of
14-3-3 zeta protein for healthy postmenopausal women
(mean age, 58 y), was 0.39 0.11 ng/ml. Subsequently, at
the start of chemotherapy, the mean serum level of 14-3-3
zeta protein in EOC patients was 2.38 1.44 pg/ml, and
2.17 1.71 pg/ml after the final treatment. Neither the
difference in levels detected for EOC patients and healthy
Figure 1 Axial CT slices of patient # 3 after contrast material was intravenously administered. A target lesion with a SLD of 84 mm is
localized near the minor pelvis before (A) and after (C) six cycles of first-line chemotherapy. (B) The arrow represents a mesenteric peritoneal
carcinosis (non-target lesion) in the same patient that is level with the lower edge of the liver prior to chemotherapy. A significant amount of
ascites associated with the non-target lesion is also observed (B). (D) Both non-target lesions are absent after the completion of chemotherapy.
Figure 2 Mean levels of serum biomarkers (CA-125, HE4 and 14-3-3 zeta protein) measured with ELISA and ECLIA methods, and the
ROMA index values. Average values of CA-125 (U/mL) +/ SEM determined by ECLIA (A) and by ELISA (D) are shown according to the
chemotherapy cycles (consistently on each x-axis from 1 to 6). Mean concentrations of HE4 (p/M), measured by ECLIA (B) and by ELISA (E) are
also shown. Furthermore average values determined by ELISA (G) of 14-3-3 zeta protein (ng/ml) are depicted. Based on the above-mentioned
data, we calculated the postmenopausal ROMA index values (%) for ECLIA (C) and for ELISA (F) techniques. We represent the mean levels of
14-3-3 zeta protein (ng/mL) for each of the treatment days. Wilcoxon signed-rank test analysis was performed for each diagram, and statistical
significance represented with asterisk (*) was set at p < 0.05, and (**) when p < 0.01.
Table 3 Levels of CA-125, HE4, and 14-3-3 Z protein determined by ELISA and ECLIA methods (Continued)
8 1 475.23 259.74 90.07
2 500 90.83 75.92
3 189.18 67.99 53.34
4 500 904 97.18
5 172.63 57.11 47.22
6 402.03 62.74 64.64
9 1 21.16 904 77.35
2 29.53 170.69 43.41
3 4.97 97.08 10.38
4 8.13 96.13 14.09
5 5.90 105.57 12.56
6 8.57 84.25 12.96
10 1 500 904 97.18
2 189.91 62.04 51.08
3 500 441.44 94.22
4 500 149.23 84.07
5 84 127.48 54.97
6 29.43 90.79 28.37
11 1 7.85 904 62.12
2 9.58 201.95 28.59
3 0.93 131.04 4.42
4 3.75 105.53 9.34
5 2.65 107.64 7.54
6 NA NA NA
12 1 126.02 397.04 84.2
2 500 365.72 93.07
3 192.79 273.67 83.2
4 178.79 223.37 79.09
5 121.86 168.58 68.17
6 NA NA NA
13 1 500 904 97.18
2 NA NA NA
3 500 904 97.18
4 500 904 97.18
5 120.65 355.03 82.16
6 NA NA NA
ROMA, risk of malignancy algorithm.
NA, not available - when serum samples were too hemolyzed and measurements could not be made.
postmenopausal patients, nor at the beginning and end of
chemotherapy, was found to be significant (Figure 2/G).
Correlation between radiological findings and serum
Neither ELISA nor ECLIA measurements of CA-125 and
HE4 serum biomarkers provided significant linear
regression correlations. However, the ROMA index values
that were calculated based on these values did provide a
strong significant regression correlation (r = 0.840, p =
0.036 and r = 0.920, p = 0.009, respectively) (Figure 3/C
and F). Moreover, with a p-value margin of 0.01, a
significant linear correlation was found for all ECLIA
measurements. Linear regression analysis of 14-3-3 zeta protein
Figure 3 Changes in CA-125, HE4, and 14-3-3 zeta protein serum levels, and ROMA index values during the six cycles of paclitaxel/
carboplatin-based chemotherapy that were performed. Mean concentrations of CA-125 determined by ELISA (A) and by ECLIA (D) are shown
for each of the treatment days. Mean levels of HE4 determined by ELISA (B) and ECLIA (E) are also shown for each of the treatment days. Mean
concentrations of 14-3-3 zeta protein were determined by ELISA (G) are represented at each chemotherapy days. Postmenopausal ROMA index
values were calculated based on ELISA (C) and ECLIA (F) data. Linear regression analysis was performed for each diagram (see r value), and
statistical significance was set at p < 0.05.
levels at each treatment day, showed no significant
correlation between the mean serum values and the
chemotherapy cycles (r = 0.073; p = 0.089) (Figure 3/G).
Spearmans correlation analysis further identified a
significant correlation between CA-125 serum levels
determined by ELISA and the largest tumor diameter measured
by CT scans obtained following chemotherapy (p = 0.011).
Levels of HE4 detected by ECLIA were also found to
significantly correlate with tumor diameter (p = 0.04), while
levels of 14-3-3 zeta protein did not significantly correlate
with any of the examined parameters.
Several studies have demonstrated the limitations
associated with depending on any single tumor marker for the
detection of EOC. Initially, CA-125 was widely used.
However, other malignant and benign diseases also
express CA-125, thereby limiting its reliability as a tumor
marker. In particular, CA-125 has a high false-positive
rate among women with benign gynecological conditions
such as endometriosis , and a low sensitivity in
identifying patients with early-stage ovarian cancer .
Accordingly, when EOC is diagnosed, 80% of cases are in
an advanced stage of disease (e.g., FIGO III-IV) . To
improve the specificity and sensitivity of an ovarian
cancer diagnosis, additional tumor markers have been
investigated. One novel tumor marker is HE4, which contains
two whey acid protein (WAP) domains and eight
cysteine residues that constitute a four-disulphide bond core
. HE4 localizes to human chromosome 20q12-13.1
and its expression significantly increases during
malignant transformation. However, HE4 is expressed in
normal tissues as well, and therefore, is not tumor specific.
Correspondingly, it has been hypothesized that the
function of HE4 is related to both spermiotelcosis (a protease
inhibitor involved in sperm maturation) and natural
immunity, although the mechanistic details of HE4
functions remain to be clarified . As a tumor marker for
the early detection of ovarian cancer, Moore et al.
reported a sensitivity of 72.9% and a specificity of 95% for
HE4 . Moreover, when both HE4 and CA-125 were
detected, the sensitivity increased to 76.4%. Therefore,
the detection of more than one biomarker resulted in a
33.1% increase in the sensitivity of CA-125, and a 3.5%
increase in HE4 sensitivity .
In the present study, ROMA values provided a PI based
on the pre- or postmenopausal status of a patient, and the
presence and levels of biomarkers CA-125 and HE4. As
such, this PI relies on an accurate determination of serum
levels of HE4 and CA-125. Moreover, in a recent study,
the ROMA was found to be more effective in predicting
ovarian cancer than the widely used risk of malignancy
index (RMI), which employs ultrasound findings, CA-125
concentrations, and menopausal status . Furthermore,
when the specificity was set to 75%, the RMI had a
sensitivity of 84.6%. For the same specificity, the sensitivity of
the ROMA was significantly higher (94.3%). Although
biomarker concentrations can be assayed by various methods
(e.g., ELISA, chemiluminescent microparticle
immunoassay), a recent study conducted by Ruggeri et al.
demonstrated that chemiluminescent immunoassays are more
adequate and more reproducible than commercially
available ELISA kits that are characterized by interassay
imprecision percentages (CV%) ranging from 6.8-10.3%,
compared to < 4% for ECLIA . The results of the
present study are consistent with these findings, and they
further support the use of the ECLIA method for routine
determinations of CA-125 and HE4 levels. Furthermore,
the deviation in accuracy for ELISA versus ECLIA can be
attributed to the fully automated format of ECLIA, while
ELISAs are manual assays that also require testing samples
14-3-3 zeta protein plays an important role in several
different biological mechanisms. For example, it has
been reported to be an adaptor protein for intracellular
signaling since it contains tandem repeats of
phosphoserine motifs that have the capacity to bind upstream
and downstream signaling molecules [21-24]. 14-3-3 zeta
protein also facilitate cell migration by forming a ternary
complex with integrin alpha-4 and paxillin . However,
14-3-3 zeta also has potential roles in cancerogenesis,
based on its ability to bind NF-kappa B, beta-catenin, and
Bcl-2, and to augment cancer cell proliferation .
Furthermore, 14-3-3 zeta protein has been shown to block
activation of p38 mitogen-activated protein kinase (MAPK),
thereby mediating an anti-apoptotic mechanism .
Numerous investigations have also suggested that
14-33 zeta protein is a key molecule in the malignant
pathological processes of several malignancies, including oral,
esophageal, lung, and breast cancers, as well as B cell
lymphoma. Recently, He et al. reported that 14-3-3 zeta
protein represents a candidate biomarker and a
metastasis-promoting factor in ovarian cancer based on
a serum proteomic analysis of a nude mouse xenograft
model containing SKOV-3 cells and a mass
spectrometry [liquid chromatography-tandem mass spectrometry
(LC-MS/MS)] analysis to identify metastasis-related
serum proteins . Significantly higher expression of
143-3 zeta was detected in EOC patients than in patients
with benign gynecological diseases. Furthermore,
compared to CA-125, serum levels of 14-3-3 zeta protein was
significantly upregulated when microscopic peritoneal
metastasis was present, or when bilateral ovaries were
involved. Accordingly, the authors suggested that 14-3-3
zeta protein may be a useful tool in differentiating FIGO
stage Ib and Ic ovarian cancers from stage Ia ovarian
cancers in the clinic . However, the results of the present
study are not consistent with these findings. For example,
significant differences in the serum levels of 14-3-3 zeta
protein was not detected in healthy menopausal women
versus patients with advanced stage EOC. Furthermore,
significant changes in serum levels of 14-3-3 zeta protein
was not detected during the six consecutive cycles of
chemotherapy treatment that were administered (Figure 2/G),
although CT scans and CA-125 and HE4 levels
unambiguously indicated the efficacy of the treatment. A possible
explanation for these results is the insufficient number of
patients enrolled in the current study. Thus, future studies
should include a larger cohort in order to identify statistically
significant changes. It is also possible that serum proteins
may undergo degradation, even when stored at 80C. In
particular, it may be that 14-3-3 zeta is an unstable protein
that needs to be assayed shortly after collection.
Furthermore, an intriguing possibility is that 14-3-3 zeta may bind
proteins activated by chemotherapeutic agents, or present as
a result of chemotherapy, thereby obscuring detection of
143-3 zeta protein in serum. In the future s large-scale clinical
investigation is necessary to evaluate the efficacy of 14-3-3
zeta protein, and to determine the sensitivity and the
specificity of this biomarker comparing it to CA-125 and to HE4.
In conclusion, determination of CA-125 and HE4
serum levels for the ROMA represents a useful tool for
the prediction of chemotherapy efficacy for EOC
patients. However, based on our current findings, levels of
14-3-3 zeta protein were not found to reliably correlate
with the clinical behavior of EOC, and therefore we
question if it would be a useful biomarker for this
EOC: Epithelial ovarian cancer; FIGO: International federation of obstetrics
and gynecology; HE4: Human epididymis protein 4; ROMA: Risk of ovarian
malignancy algorithm; CT: Computed tomography; LD: Longest diameter;
ELISA: Enzyme-linked immunosorbent assay; ECLIA: Quantitative
electrochemiluminescence assay; PI: Predictive index; SEM: Standard error of
the mean; RMI: Risk of malignancy index; WAP: Whey acid protein;
MAPK: Mitogen-activated protein kinase; LC-MS/MS: Liquid
chromatographytandem mass spectrometry.
The authors have no conflict of interest with the present study to report.
IH has made substantial contributions to the conception and the design of
this study, he also contributed significantly by collecting ovarian cancer
samples. Furthermore he is responsible for all corrections and finalizations
made to the manuscript.PG participated in the study design and
coordination.TK along with LS and BP performed all
the necessary immunoassays and biomarker measurements. AK performed
the radiological assesments of all patients and is responsible for the
interpretation within the manuscript of the radiological findings.NF
performed all the statistical analysis necessary. BF participated in the design
of the study, collection of samples and in addition,he has also been involved
in drafting the manuscript. All authors read and approved the final
manuscript and have given final approval of the version to be published.
This study was funded by OTKA K104960 (Szereday, L). We thank the
laboratory assistants in the Department of Medical Microbiology and
Immunology and the Institute of Laboratory Medicine for technical
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