Fibroblast growth factor 19 is correlated with an unfavorable prognosis and promotes progression by activating fibroblast growth factor receptor 4 in advanced-stage serous ovarian cancer

  • Authors:
    • Lingling Hu
    • Lanxiang Cong
  • View Affiliations

  • Published online on: August 20, 2015     https://doi.org/10.3892/or.2015.4212
  • Pages: 2683-2691
Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )
Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )


Abstract

Fibroblast growth factor receptor 4 (FGFR4) has been confirmed to be associated with the progression and prognosis of ovarian cancer, while the underlying mechanism has not been well elucidated and the clinical significance of its ligand, fibroblast growth factor 19 (FGF19), has not been explored. To study the clinical significance of FGF19 in advanced‑stage serous ovarian cancer, we detected the expression of FGF19 and FGFR4 by immunohistochemistry (IHC), evaluated the correlation between FGF19 and clinicopathological factors by Chi-square (χ2) test, and analyzed the association between FGF19, FGFR4 and the overall survival rate using the Kaplan‑Meier method. As a result, we demonstrated that high expression of FGF19 and FGFR4 both predicted unfavorable prognosis (P=0.033 and 0.018, respectively), whereas FGF19-FGFR4 double high expression was a more sensitive prognostic factor of advanced-stage serous ovarian cancer (P<0.001). With experiments in vitro, we demonstrated that both recombinant FGF19 and secreted FGF19 promoted ovarian cancer proliferation and invasion by activating FGFR4 and the subsequent AKT-MAPK signaling pathway, suggesting that FGF19-FGFR4 signaling may auto-activate in a paracrine or autocrine manner. In conclusion, FGF19-FGFR4 double high expression was a more sensitive prognostic factor than FGF19 or FGFR4 alone in advanced-stage serous ovarian cancer. The FGF19-FGFR4 signaling pathway can promote ovarian cancer proliferation and invasion by the AKT-MAPK signaling pathway, indicating that FGF19 could be a potential therapeutic drug target of advanced-stage serous ovarian cancer.

Introduction

Ovarian cancer is one of the most lethal malignancies among all gynecological tumors and results in the second highest number of cancer-related deaths among women worldwide (1). Ovarian cancer is a group of heterogeneous tumors, which comprise several histological subtypes: serous, mucinous, endometrioid, clear cell and undifferentiated carcinomas. High-grade serous carcinoma accounts for ~80% of epithelial ovarian carcinomas and has the highest recurrence rate. Due to advances in surgical management and adjuvant chemotherapy, the survival rate of ovarian cancer patients has been markedly increased, but ovarian cancer still remains the most lethal cancer in gynecology with a 5-year overall survival of only 25–35% and causes more than 15,000 deaths/year (24), which is the focus of ovarian cancer study for decades (5). The high mortality of ovarian cancer is partly due to clinical silence and high recurrence. Most patients are diagnosed at an advanced stage and the majority of patients, up to 75%, eventually suffer recurrence, although they receive debulking surgery and standard chemotherapy (6). In conclusion, effective, predictive or prognostic biomarkers are urgently needed for early diagnosis and survival time prolongation.

In the human, fibroblast growth factors (FGFs) are heparin-binding proteins consisting of a family of 18 members, which are essential for signal transduction through interacting with cell surface-associated heparan sulfate proteoglycans (7,8). FGFs are involved in many cellular processes such as proliferation, differentiation and angiogenesis, and their ectopic overexpression has been discovered in many types of cancers. FGFs function as signaling inducers by interacting with FGFRs. Fibroblast growth factor receptors (FGFRs) comprise five members, including four tyrosine kinase receptors (FGFR1, FGFR2, FGFR3 and FGFR4) and a non-tyrosine kinase receptor (FGFR5, also known as FGFRL1) (9). FGFR4 is demonstrated to bind with FGF1, FGF2, FGF4, FGF6, FGF8, FGF9, FGF16, FGF17, FGF18, FGF19, FGF21 and FGF23, but FGF19 and FGF21 have the highest affinity and specificity to FGFR4. Among the FGFR family, FGFR4 is distinguished in ovarian cancer because previous studies have determined that it is a prognostic biomarker and potential therapeutic target for ovarian cancer (10). However, the underlying mechanism of how FGFR4 leads to an unfavorable prognosis and how FGFR4 is stimulated and triggers the downstream signaling network has not been well elucidated.

In the present study, we hypothesized that FGFR4 functions in ovarian cancers as part of a signaling network in the cancer microenvironment and thus we detected the expression of FGFR4-specific ligand, FGF19, in advanced-stage serous ovarian cancer with immunohistochemistry (IHC). Furthermore, we evaluated the prognostic value of FGF19 with univariate and multivariate analyses. Using experiments in vitro, we demonstrated that FGF19 can be secreted and promotes ovarian cancer progression such as proliferation and invasion by activating FGFR4, indicating that the FGF19-FGFR4 paracrine pathway can be considered as a potential drug target in ovarian cancer therapy.

Materials and methods

Patients and follow-up

A total of 134 patients were diagnosed with advanced-stage serous ovarian cancer by routine pathology from 2002 to 2012 at Linyi Hospital and Yishui Central Hospital, which was the primary cohort. Advanced stage was identified as stage IIIB-IV in the International Federation of Gynecology and Obstetrics stage system according to a previous study (10). The validation cohort consisting of 74 samples was selected from the primary cohort with criteria as available tissue samples and follow-up of more than 3 months. Samples (tissues and blood samples) were obtained from the Pathology Department with prior consent of the patients or their families and by approval of the Institutional Clinical Ethics Review Board. The diagnoses of the validation cohort were confirmed by two senior pathologists.

Additionally, 15 pairs of fresh frozen tumor tissues and the corresponding adjacent non-tumor tissues were collected immediately after surgery with prior consent of the patients and preserved in liquid nitrogen. Real-time polymerase chain reaction (PCR) with primers of FGF19 and FGFR4 was used to detect the mRNA levels in the tumor and non-tumor tissues.

Cell culture and agents

Ovarian cancer cell lines SK-OV-3, HO8910, HO8910PM and OVCAR3, hepatocellular carcinoma cell line HepG2, breast cancer cell line MCF-7 and colorectal cell line SW480 were purchased from the Chinese Academy of Sciences Cell Bank (Shanghai, China). The cell line SK-OV-3 was cultured in McCoy's 5A medium (Sigma-Aldrich) and HO8910, HO8910PM and OVCAR3 cells were cultured in RPMI-1640 medium. HepG2, MCF-7 and SW480 were cultured in Dulbecco's modified Eagle's medium (DMEM), all of which were supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin and 100 µg/ml streptomycin, and cultured under 5% CO2 in a humidified incubator at 37°C. Recombinant human FGF19 (RFGF19) protein was purchased from Sino Biological, Inc. (Beijing China). Antibodies used in our experiments were as follows: mouse monoclonal anti-FGF19 antibody was from R&D Systems (Minneapolis, MN, USA), anti-FGFR4 (Phospho-Tyr642) antibody was from Signalway Antibody LLC. (College Park, MD, USA), anti-β-actin antibody was from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). The remaining antibodies were all from Cell Signaling Technology, Inc. (Danvers, MA, USA).

IHC and evaluation

IHC was carried out by the streptavidin peroxidase complex method. Followed by deparaffinization with xylene and graded alcohol, endogenous peroxidase was inactivated by incubation in 3% hydrogen peroxide for 10 min. After nonspecific blocking with 5% bovine serum albumin, the slides were incubated in the primary antibody at a dilution of 1:100 at 4°C overnight. After the primary antibody, samples were incubated with the corresponding secondary antibody at 37°C for 1–2 h and then in 3,3′-diaminobenzidine (DAB) solution until satisfactory staining was shown. Every slide was evaluated by two senior pathologists unaware of the clinical information. Cases without concurrence were re-evaluated by a third pathologist.

The score of the IHC staining was based on multiplication of the staining intensity and the positive cell percentage. The staining intensity was scored as negative (0), weak (1), moderate (2) and strong (3), and scores of the stained area were defined as follows: 0, <10%; 1, 10–30%; 2, >30–50%; and 3, >50% positive cells. The cut-off was arbitrarily defined as a score ≥4 was considered as high FGF19 or FGFR4 expression; a score <4 was considered as low FGF19 or FGFR4 expression.

Western blotting

Expression of FGF19 and FGFR4 was detected by western blotting. Briefly, the cells were lysed in modified RIPA buffer (Tris 50 mM, NaCl 150 mM, Triton X-100 1%, SDS 0.1%, deoxycholate 0.5%, sodium orthovanadate 2 mM, sodium pyrophosphate 1 mM, NaF 50 mM, EDTA 5 mM, PMSF 1 mM and 1X protease inhibitor cocktail) on ice for 10 min for complete lysis, and then centrifuged at 10,000 × g for 30 min. Protein concentration of the supernatant was measured by Bradford assay kit (Sangon Biotech Co., Ltd., Shanghai, China). An amount of 40 µg of the extracted protein from each sample was used for electrophoresis in 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and then transferred to an NC membrane (Pall Corporation). The membrane was incubated in 5% defatted milk for 1 h, in diluted primary antibody overnight and in horseradish peroxidase-labeled secondary antibody for 2 h in order. Proteins were finally visualized using enhanced chemiluminescence western blotting detection reagents (Thermo Fisher Scientific, Waltham, MA, USA).

Quantitative real-time polymerase chain reaction (qRT-PCR)

RT-PCR was used to detect the mRNA levels of FGF19 and FGFR4. Primers were designed and purchased from Sigma-Aldrich as follows: FGF19 forward, 5′-GCACAGTTTGCTGGAGATCA-3′ and reverse, 5′-ATCTCCTCCTCGAAAGCACA-3′; FGFR4 forward, 5′-AGCACCCTACTGGACACACC-3′ and reverse, 5′-ACGCTCTCCATCACGAGACT-3′. Briefly, total RNA of ovarian cancer cells was first extracted from the cell lines with the Rneasy kit (Qiagen) and the concentration of total RNA was tested. Then qRT-PCR was achieved and relative gene expression was measured by the StepOnePlus Real-Time PCR system (Applied Biosystems) using the SYBR-Green method according to the manual. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was applied as an internal control.

Small interfering RNA and transfection

Knockdown of FGF19 or FGFR4 was accomplished by small interfering RNA (siRNA) purchased from Invitrogen (Carlsbad, CA, USA). The siRNA sequences of FGF19 and FGFR4 were obtained from Santa Cruz biotechnology, Inc. Transfection of siRNA was realized with Lipofectamine 2000 (Invitrogen) according to the user guide. Results of the knockdown were validated by qRT-PCR and western blotting.

Cell proliferation assays

Cell proliferation was assessed by the MTT kit (Sangon Biotech Co., Ltd.). Briefly, an equal number of cells was planted into a 96-well plate and then starved for 6 h before stimulation. After stimulation with recombinant FGF19 or conditioned medium for the expected time, the proliferation was terminated by addition of 10 mg/ml MTT and then incubated at 37°C for 4–6 h. After removing the superior carefully, the crystals at the bottom were resolved by 100 µl DMSO and the absorbance was read at 490 nm. The optical density (OD) at 490 nm was set as the baseline, and the proliferation index of the other groups was calculated by the OD490 ratio to the baseline. Every group had at least eight parallel wells, and analyzed data were from at least three independent experiments.

Matrigel invasion assays

Matrigel invasion assays were performed to evaluate the effect of FGF19 on cell invasion using 8-µm pore size BD BioCoat Matrigel invasion chambers (Becton-Dickinson, Franklin Lakes, NJ, USA). After being planted into the upper chambers and starved for 6 h, the cells were incubated in 10 ng/ml recombinant FGF19 protein of conditioned medium for 24 h and then fixed and stained with Diff-Quik stain. Non-invading cells in the upper chambers were wiped off while the cells in the lower chamber were counted using at least 8 random fields. The invaded number of cells without stimulation was counted and set as the baseline and the invasion index of the other groups was calculated as the ratio to the baseline. Analyzed data were from at least three independent experiments.

Enzyme-linked immunosorbent assay (ELISA) and conditioned medium

Serum FGF19 levels were measured with sandwich ELISA by the FGF19 Quantikine ELISA kit (R&D Systems), following the manufacturer's instructions and previous studies (11,12). FGF19 concentration in the cell medium was tested after treatment with 50 µM chenodeoxycholic acid (CDCA) for 48 h, and was then detected by an ELISA kit according to previous studies (13,14). All samples from the cell culture media or serum were detected in duplicate.

To obtain efficient conditioned medium (CM), OVCAR3 cells were cultured in normal medium with 10% FBS for the expected time. Then CM was collected and filtered with a low protein-binding filter (0.22 µm) (Millipore, Billerica, MA, USA). Amicon Ultra 15 ml filters were used to concentrate the CM at 4,000 g when necessary. The same procedures were performed using SK-OV-3 cells as a control group. The CM was used immediately, stored at 4°C for a week or frozen at −80°C.

Statistical analysis

All data were analyzed by SPSS 17.0 software (SPSS, Inc., Chicago, IL, USA). Correlations between FGF19 and clinicopathological features were analyzed by χ2 test, and correlations between FGF19, FGFR4 and overall survival rates were calculated by the Kaplan-Meier method, and the difference between the high-expression and low-expression group was analyzed by log-rank test. In experiments in vitro, differences between the control group and tested group were analyzed by the Student's unpaired t-test. P<0.05 was considered to indicate a statistically significant result.

Results

Expression of FGF19 and FGFR4 in ovarian cancer

Expression of FGF19, the known specific ligand of FGFR4, was first detected in ovarian cancer samples with IHC. FGF19 expression was mainly observed in the cytoplasm, where it may be secreted out and function as a growth factor (Fig. 1A and B). FGFR4 expression was observed in both the membrane and cytoplasm (Fig. 1C and D). According to the criteria described in Materials and methods, expression of FGF19 and FGFR4 was divided into a high expression and low expression group. The percentage of samples with high-FGF19 and high-FGFR4 was 41.89 and 39.19%, respectively (Table I). Moreover, to evaluate the significance of FGF19 and FGFR4 co-expression, we further defined FGF19-FGFR4 double high expression as cases having high expression for both FGF19 and FGFR4 expression, with the rest of the cases defined as the control group. The percentage of cases with FGF19-FGFR4 double high expression was 21.62% in our cohort.

Table I

Characteristics of the patients with advanced-stage serous ovarian cancer.

Table I

Characteristics of the patients with advanced-stage serous ovarian cancer.

CharacteristicsNo. of patients%
Age (years)
 <501722.97
 ≥505777.03
Lymph node metastasis
 No3750.00
 Yes3750.00
Histological grade
 I1114.86
 II2432.43
 III3952.70
FGF19
 Low4358.11
 High3141.89
FGFR4
 Low4560.81
 High2939.19
FGF19+FGFR4
 Low5878.38
 High1621.62

Moreover, FGF19 and FGFR4 mRNA expression levels in tumor tissues and adjacent non-tumor tissues were detected and compared (Fig. 1E). The mRNA levels of FGF19 and FGFR4 in tumor tissues were demonstrated to be much higher than levels in the non-tumor tissues.

Clinical significance of FGF19

The correlation between FGF19 and clinicopathologic factors and the 5-year overall survival rates were analyzed to evaluate the clinical significance of FGF19 in ovarian cancer. Based on the χ2 test, we demonstrated that high expression of FGF19 was significantly associated with positive lymph node metastasis (P=0.033), indicating that FGF19 may play an important role in tumor invasion and metastasis. High FGF19 expression tended to be correlated with high FGFR4 expression, but the tendency was not statistically significant (P=0.063) (Table II).

Table II

Correlation between FGF19 and clinicopathological features.

Table II

Correlation between FGF19 and clinicopathological features.

CharacteristicsFGF19
P-valuea
LowHigh
Age (years)
 <501070.946
 ≥503324
Lymph node metastasis
 No26110.033
 Yes1720
Histological grade
 I830.122
 II1014
 III2514
FGFR4
 Low30150.063
 High1316

a Chi-square test.

Kaplan-Meier method was performed to further evaluate the prognostic value of detected clinicopathological parameters (Table III). High FGFR4 expression was demonstrated to be significantly associated with a poorer overall survival rate (P= 0.018), which was consistent with a previous study (Fig. 2A) (10). In addition, we first proved that high FGF19 expression was closely correlated with unfavorable prognosis (P=0.033), suggesting that FGF19 could be considered as a prognostic biomarker and potential drug target (Fig. 2b). Notably, double high expression of FGF19 and FGFR4 was a better biomarker and predicted prognosis more reliably than FGF19 or FGFR4 alone (P<0.001) (Fig. 2C). FGF19-FGFR4 double high expression can predict poorer prognosis more accurately and sensitively than FGF19 or FGFR4 alone, indicating the promising and potential clinical significance of considering FGF19-FGFR4 double positivity as a biomarker. Moreover, the association between FGF19-FGFR4 and poor prognosis indicated that the FGF19-FGFR4 signaling pathway may promote cancer progression by a self-sufficient autocrine or paracrine pathway.

Table III

Correlation between clinicopathological features and the overall survival rate.

Table III

Correlation between clinicopathological features and the overall survival rate.

Characteristics5-year survival rate (%)P-valuea
Age (years)
 <50390.967
 ≥5031.7
Lymph node metastasis
 No54.30.002
 Yes7.8
Histological grade
 I38.10.401
 II37
 III31.4
FGF19
 Low41.60.033
 High20.6
FGFR4
 Low41.80.018
 High0
FGF19+FGFR4
 Low37.9<0.001
 High13.4

a Log-rank test.

FGF19 and FGFR4 in ovarian cancer cells, medium and blood

Using experiments in vitro, we performed a series of functional assays to explore the mechanism explaining why high FGF19 and FGFR4 expression imply a much poorer prognosis. Expression levels of FGF19 and FGFR4 in ovarian cancers were first detected by western blotting. As shown in Fig. 3A, FGF19 and FGFR4 were widely expressed in ovarian cancer cell lines. SK-OV-3 had the lowest FGFR4 expression and HO8910 had the lowest FGF19 expression. OVCAR3 had relatively high FGF19 and FGFR4 expression, suggesting OVCAR3 as a suitable model for a knockdown assay. To evaluate the role of FGF19 and FGFR4 in ovarian cancer progression, we regulated FGF19 and FGFR4 expression in OVCAR3 cells by siRNA transfection and validated the successful knockdown of FGF19 (Fig. 3B and C) and FGFR4 (Fig. 3D and E) by qRT-PCR and western blotting. As a secreted growth factor, FGF19 was suspected to be secreted out from ovarian cancer cells. Thus medium of the SK-OV-3 and OVCAR3 cells, which had the lowest and highest FGF19 expression respectively, was used to detect the FGF19 concentration. After treatment with 50 µM CDCA for 72 h, FGF19 in both the SK-OV-3 and OVCAR3 medium was detectable (Fig. 3F). The FGF19 concentration in the OVCAR3 medium was markedly higher than that in the SK-OV-3 medium, which was consistent with the intracellular FGF19 content. After FGF19 was knocked down by siRNA, the concentration of FGF19 in the medium was significantly decreased compared to the control group, suggesting that its concentration in medium was directly related to the intracellular content. Based on the finding that the FGF19 is associated with prognosis and is secreted out from ovarian cancer, we hypothesized that FGF19 concentration in blood may predict earlier ovarian cancer progression and poorer prognosis. Thus, we detected the FGF19 concentration in 12 healthy individuals and 12 pre-operational patients with advanced-stage ovarian cancer by ELISA method. However, the FGF19 concentration between the healthy individuals and the ovarian cancer patients had no significant difference (P=0.492) (Fig. 3G).

Additionally, we detected the expression and function of FGF19 and FGFR4 in control cell lines MCF7, SW480 and HepG2 to determine the exclusive effect of FGF19 on FGFR4. HepG2 had the highest FGFR4 expression and SW480 exhibited the most abundant FGF19 expression (Fig. 4A). Under 10 ng/ml recombinant FGF19 stimulation for 48 h, these three cell lines exhibited different proliferative rates (HepG2 had the highest and MCF7 had the lowest rate) (Fig. 4b). Moreover, this variation to FGF19 stimulation faded away after FGFR4 was knocked down in these cell lines (Fig. 4C), indicating that FGF19 accelerated the proliferation dependent on FGFR4 existence.

FGF19 promotes ovarian cancer progression in a paracrine manner

To explore the role of FGF19 in ovarian cancer progression, we further examined the effect of exogenous FGF19 on the AKT-MAPK signaling pathway in OVCAR3 cells. Proteins in the AKT-MAPK signaling pathway were detected after 10 ng/ml RFGF19 stimulation for 0–30 min (Fig. 5A). The phosphorylation level of FGFR4, AKT, Erk and p38 increased notably along with the FGF19 stimulation, indicating that FGF19 promoted FGFR4 phosphorylation and activated the AKT-MAPK signaling pathway. Although considered as a specific ligand of FGFR4, FGF19 was also proven to interact with FGFR1, FGFR2 and FGFR3 in the presence of β-klotho (1517). To evaluate whether FGFR4 was essential in the FGF19 activation process, FGFR4 was knocked down in OVCAR3 cells and the AKT-MAPK signaling pathway with recombinant FGF19 stimulation was subsequently detected (Fig. 5b). After FGFR4 knockdown, the phosphorylation levels of AKT, Erk and p38 were significantly reduced due to the decrease in FGFR4 phosphorylation, which demonstrated that FGFR4 was essentially required in the FGF19-induced AKT-MAPK activation.

To explore FGF19-FGFR4 signaling in tumor progression, 10 ng/ml FGF19 was used to stimulate OVCAR3 cells for 0–72 h after FGFR4 knockdown, and MTT assay was performed to evaluate cell proliferation (Fig. 5C). Cells with FGF19 stimulation had an obviously higher proliferation rate than the control group, and FGFR4 knockdown reversed this FGF19-induced acceleration of proliferation, indicating that FGF19 promoted ovarian cancer cell proliferation by activating FGFR4. To estimate FGF19 influence on invasion, OVCAR3 cells were cultured in 10 ng/ml RFGF19 for 24 h, at which time the proliferation did not differ, and then the Matrigel assay was carried out to detect cell invasion (Fig. 5D). In our experiment, FGF19 markedly promoted cell invasion and FGFR4 knockdown impaired this tendency, which confirmed that FGFR4 was also required in the FGF19-induced invasion. Based on our findings that FGF19 was secreted by ovarian cancer cells, we suspected that the FGF19-FGFR4 signaling pathway may be stimulated ectopically by an autocrine or paracrine pathway. Thus, we used the conditioned medium of OVCAR3, the cell line with high FGF19 expression, to stimulate SK-OV-3 cells to confirm our hypothesis. In our experiments, the conditioned medium of OVCAR3 cells notably accelerated the proliferation of SK-OV-3 cells at 72 h and the invasion of SK-OV-3 at 24 h (Fig. 5E and F), demonstrating that the FGF19 concentration in the ovarian cancer microenvironment was high enough to activate FGFR4 and downstream signaling pathways.

Discussion

In our experiments, we detected the expression of FGF19, the FGFR4 specific ligand, in advanced-stage ovarian cancer tissues, and systemically evaluated the correlation between FGF19 and clinicopathological factors and the overall survival for the first time according to our knowledge. Moreover, we performed a series of experiments in vitro to estimate the role of the FGF19-FGFR4 signaling pathway in ovarian cancer progression. Consequently, we found that FGF19 expression was significantly associated with lymphatic metastasis (P=0.033). FGF19 and FGFR4 expression could predict poorer prognosis (P=0.033 and 0.018, respectively), and FGF19-FGFR4 double high expression was a more accurate and sensitive prognostic factor in ovarian cancer (P<0.001). With experiments in vitro, we found that FGF19 existed in ovarian cancer cells and was secreted out. In addition, FGF19 stimulated the AKT-MAPK signaling pathway via activating FGFR4, by which FGF19 accelerated proliferation and invasion. Moreover, we proved that the FGF19 concentration in cell culture medium was high enough to promote cell proliferation and invasion, indicating that the FGF19-FGFR4 signaling pathway may be activated ectopically by an autocrine or paracrine pathway.

FGFR4 overexpression has been previously proven to be associated with progression and prognosis in many types of cancers, including gastric cancer, hepatocellular carcinoma, cholangiocarcinoma, colorectal and oropharynx cancer (1822). As the known specific ligand to FGFR4, FGF19 was also demonstrated to be involved in progression of cancers such as hepatocellular carcinoma, prostate cancer (12,18,22,23) and colon cancer. The amplification of the FGFR4 gene was discovered in gynecological cancers by Jaakkola et al in 1993 (24). A previous study pointed out that FGFR4 could predict prognosis and may be a potential therapeutic target in high-advanced ovarian cancer (10). However, the FGFR4 ligand was not detected and the underlying molecular mechanism was not well elucidated, which prompted us to explore the correlation between FGF19, FGFR4, progression and prognosis in ovarian cancer. Moreover, FGFR4 is widely expressed in many types of tumors and we demonstrated that not only ovarian cancer cell lines but also other cell lines exhibited a different response to FGF19-induced proliferation, which required the participation of FGFR4. Thus, FGF19-induced proliferation may be a general phenomenon regarding all cell lines with abundant FGFR4, indicating that the FGF19-FGFR4 signaling pathway may be a potential molecular target for ovarian cancer as well as other tumors with FGFR4 overexpression.

Among the human FGF factors, FGF19 is distinguished by its function as a hormone, regulating bile acid synthesis, with effects on glucose and lipid metabolism (25,26). Interestingly, ovarian cancer risk is believed to be positively correlated with glucose and lipid metabolism disorders such as obesity and diabetes (27,28). Recent evidence indicates that obesity, diabetes and ovarian cancer are more intricately related (29,30). Previous studies reported that diet and diabetes are associated with mortality and the prognosis of ovarian cancer (28,31). Based on our new findings, we boldly hypothesize that FGF19 ectopic expression and function may be one explanation why obesity and diabetes are related to high-risk or even a poorer prognosis of ovarian cancer. In our study, the FGF19 concentration of blood samples was detected but no significant difference between the patients and healthy individuals was observed. We believe that more cases should be enrolled and the cohort should be further divided into subgroups according to suspicious parameters such as diabetes and BMI to detect the role of FGF19 in the correlation between ovarian cancer and lipid metabolism.

The FGFR family has five members and can interact with several FGFs with different affinity, which may trigger different downstream signaling pathways and regulate different cellular biological processes (32,33). Moreover, the FGFR family is also proven to undergo crosstalk with other receptors such as the epidermal growth factor receptor family, which may increase the complexity of the FGFR signaling network exponentially (34). Even in ovarian cancer, the FGF19-FGFR4 signaling pathway is associated with obvious conundrums to solve, such as the difference in FGF19 secretion between biological and pathological patterns, and how FGF19 interacts with FGFR4 and which downstream pathways are dominantly triggered. These questions require further fundamental experiments, especially using animal models. Moreover, inhibitors mimicking the FGF19 structure are potential molecular drugs targeting FGFR4 in ovarian cancer based on our finding that FGF19 is also a prognostic marker. We hope our findings stimulate more research on the function of FGF19 in ovarian cancer, which may further help identify effective therapeutic drugs and increase the survival of patients with ovarian cancer.

In conclusion, we demonstrated that both FGF19 and FGFR4 overexpression predict an unfavorable prognosis in ovarian cancer, while FGF19-FGFR4 double high expression is a better and more sensitive prognostic biomarker. Moreover, either recombinant FGF19 or FGF19 secreted by ovarian cancer cells promotes proliferation and invasion by FGFR4 activation and the subsequent AKT-MAPK signaling pathway. This indicates that FGF19-FGFR4 signaling may be auto-activated in a paracrine or autocrine manner and that FGF19 could be a potential drug target for ovarian cancer.

References

1 

Jemal A, Thomas A, Murray T and Thun M: Cancer statistics, 2002. CA Cancer J Clin. 52:23–47. 2002. View Article : Google Scholar : PubMed/NCBI

2 

Colombo PE, Fabbro M, Theillet C, Bibeau F, Rouanet P and Ray-Coquard I: Sensitivity and resistance to treatment in the primary management of epithelial ovarian cancer. Crit Rev Oncol Hematol. 89:207–216. 2014. View Article : Google Scholar

3 

Siegel R, Naishadham D and Jemal A: Cancer statistics, 2012. CA Cancer J Clin. 62:10–29. 2012. View Article : Google Scholar : PubMed/NCBI

4 

Seltzer V; NIH Consensus Development Panel on Ovarian Cancer: NIH consensus conference. Ovarian cancer. Screening, treatment, and follow-up. JAMA. 273:491–497. 1995. View Article : Google Scholar

5 

Kang KW, Lee MJ, Song JA, Jeong JY, Kim YK, Lee C, Kim TH, Kwak Kb, Kim OJ and An HJ: Overexpression of goosecoid homeobox is associated with chemoresistance and poor prognosis in ovarian carcinoma. Oncol Rep. 32:189–198. 2014.PubMed/NCBI

6 

McGuire WP, Hoskins WJ, Brady MF, Kucera PR, Partridge EE, Look KY, Clarke-Pearson DL and Davidson M: Cyclophosphamide and cisplatin compared with paclitaxel and cisplatin in patients with stage III and stage IV ovarian cancer. N Engl J Med. 334:1–6. 1996. View Article : Google Scholar : PubMed/NCBI

7 

Turner N and Grose R: Fibroblast growth factor signalling: From development to cancer. Nat Rev Cancer. 10:116–129. 2010. View Article : Google Scholar : PubMed/NCBI

8 

Eswarakumar VP, Lax I and Schlessinger J: Cellular signaling by fibroblast growth factor receptors. Cytokine Growth Factor Rev. 16:139–149. 2005. View Article : Google Scholar : PubMed/NCBI

9 

Wiedemann M and Trueb B: Characterization of a novel protein (FGFRL1) from human cartilage related to FGF receptors. Genomics. 69:275–279. 2000. View Article : Google Scholar : PubMed/NCBI

10 

Zaid TM, Yeung TL, Thompson MS, Leung CS, Harding T, Co NN, Schmandt RS, Kwan SY, Rodriguez-Aguay C, Lopez-Berestein G, et al: Identification of FGFR4 as a potential therapeutic target for advanced-stage, high-grade serous ovarian cancer. Clin Cancer Res. 19:809–820. 2013. View Article : Google Scholar : PubMed/NCBI

11 

Wang D, Zhu W, Li J, An C and Wang Z: Serum concentrations of fibroblast growth factors 19 and 21 in women with gestational diabetes mellitus: Association with insulin resistance, adiponectin, and polycystic ovary syndrome history. PLoS One. 8:e811902013. View Article : Google Scholar : PubMed/NCBI

12 

Feng S, Dakhova O, Creighton CJ and Ittmann M: Endocrine fibroblast growth factor FGF19 promotes prostate cancer progression. Cancer Res. 73:2551–2562. 2013. View Article : Google Scholar : PubMed/NCBI

13 

Song KH, Li T, Owsley E, Strom S and Chiang JY: Bile acids activate fibroblast growth factor 19 signaling in human hepatocytes to inhibit cholesterol 7alpha-hydroxylase gene expression. Hepatology. 49:297–305. 2009. View Article : Google Scholar :

14 

Stejskal D, Karpísek M, Hanulová Z and Stejskal P: Fibroblast growth factor-19: Development, analytical characterization and clinical evaluation of a new ELISA test. Scand J Clin Lab Invest. 68:501–507. 2008. View Article : Google Scholar : PubMed/NCBI

15 

Xie MH, Holcomb I, Deuel B, Dowd P, Huang A, Vagts A, Foster J, Liang J, Brush J, Gu Q, et al: FGF-19, a novel fibroblast growth factor with unique specificity for FGFR4. Cytokine. 11:729–735. 1999. View Article : Google Scholar : PubMed/NCBI

16 

Zhang X, Ibrahimi OA, Olsen SK, Umemori H, Mohammadi M and Ornitz DM: Receptor specificity of the fibroblast growth factor family. The complete mammalian FGF family. J Biol Chem. 281:15694–15700. 2006. View Article : Google Scholar : PubMed/NCBI

17 

Kurosu H, Choi M, Ogawa Y, Dickson AS, Goetz R, Eliseenkova AV, Mohammadi M, Rosenblatt KP, Kliewer SA and Kuro-o M: Tissue-specific expression of betaKlotho and fibroblast growth factor (FGF) receptor isoforms determines metabolic activity of FGF19 and FGF21. J Biol Chem. 282:26687–26695. 2007. View Article : Google Scholar : PubMed/NCBI

18 

Miura S, Mitsuhashi N, Shimizu H, Kimura F, Yoshidome H, Otsuka M, Kato A, Shida T, Okamura D and Miyazaki M: Fibroblast growth factor 19 expression correlates with tumor progression and poorer prognosis of hepatocellular carcinoma. BMC Cancer. 12:562012. View Article : Google Scholar : PubMed/NCBI

19 

Ye YW, Zhou Y, Yuan L, Wang CM, Du CY, Zhou XY, Zheng BQ, Cao X, Sun MH, Fu H, et al: Fibroblast growth factor receptor 4 regulates proliferation and antiapoptosis during gastric cancer progression. Cancer. 117:5304–5313. 2011. View Article : Google Scholar : PubMed/NCBI

20 

Xu YF, Yang XQ, Lu XF, Guo S, Liu Y, Iqbal M, Ning SL, Yang H, Suo N and Chen YX: Fibroblast growth factor receptor 4 promotes progression and correlates to poor prognosis in cholangiocarcinoma. Biochem Biophys Res Commun. 446:54–60. 2014. View Article : Google Scholar : PubMed/NCBI

21 

Dutra RL, de Carvalho MB, Dos Santos M, Mercante AM, Gazito D, de Cicco R, Group G, Tajara EH, Louro ID and da Silva AM: FGFR4 profile as a prognostic marker in squamous cell carcinoma of the mouth and oropharynx. PLoS One. 7:e507472012. View Article : Google Scholar : PubMed/NCBI

22 

Li CS, Zhang SX, Liu HJ, Shi YL, Li LP, Guo XB and Zhang ZH: Fibroblast growth factor receptor 4 as a potential prognostic and therapeutic marker in colorectal cancer. Biomarkers. 19:81–85. 2014. View Article : Google Scholar : PubMed/NCBI

23 

Sawey ET, Chanrion M, Cai C, Wu G, Zhang J, Zender L, Zhao A, Busuttil RW, Yee H, Stein L, et al: Identification of a therapeutic strategy targeting amplified FGF19 in liver cancer by oncogenomic screening. Cancer Cell. 19:347–358. 2011. View Article : Google Scholar : PubMed/NCBI

24 

Jaakkola S, Salmikangas P, Nylund S, Partanen J, Armstrong E, Pyrhönen S, Lehtovirta P and Nevanlinna H: Amplification of fgfr4 gene in human breast and gynecological cancers. Int J Cancer. 54:378–382. 1993. View Article : Google Scholar : PubMed/NCBI

25 

Jones SA: Physiology of FGF15/19. Adv Exp Med Biol. 728:171–182. 2012. View Article : Google Scholar : PubMed/NCBI

26 

Potthoff MJ, Kliewer SA and Mangelsdorf DJ: Endocrine fibroblast growth factors 15/19 and 21: From feast to famine. Genes Dev. 26:312–324. 2012. View Article : Google Scholar : PubMed/NCBI

27 

Doll KM, Kalinowski AK, Snavely AC, Irwin DE, Bensen JT, Bae-Jump VL, Kim KH, Van Le L, Clarke-Pearson DL and Gehrig PA: Obesity is associated with worse quality of life in women with gynecologic malignancies: An opportunity to improve patient-centered outcomes. Cancer. 121:395–402. 2014. View Article : Google Scholar : PubMed/NCBI

28 

Shah MM, Erickson BK, Matin T, McGwin G Jr, Martin JY, Daily LB, Pasko D, Haygood CW, Fauci JM and Leath CA III: Diabetes mellitus and ovarian cancer: More complex than just increasing risk. Gynecol Oncol. 135:273–277. 2014. View Article : Google Scholar : PubMed/NCBI

29 

Thomson CA, E Crane T, Wertheim BC, Neuhouser ML, Li W, Snetselaar LG, Basen-Engquist KM, Zhou Y and Irwin ML: Diet quality and survival after ovarian cancer: Results from the Women's Health Initiative. J Natl Cancer Inst. 106:1062014. View Article : Google Scholar

30 

Olsen CM, Nagle CM, Whiteman DC, Ness R, Pearce CL, Pike MC, Rossing MA, Terry KL, Wu AH, Risch HA, et al Australian Cancer Study (Ovarian Cancer); Australian Ovarian Cancer Study Group: Ovarian Cancer Association Consortium: Obesity and risk of ovarian cancer subtypes: Evidence from the Ovarian Cancer Association Consortium. Endocr Relat Cancer. 20:251–262. 2013. View Article : Google Scholar : PubMed/NCBI

31 

Bae HS, Kim HJ, Hong JH, Lee JK, Lee NW and Song JY: Obesity and epithelial ovarian cancer survival: A systematic review and meta-analysis. J Ovarian Res. 7:412014. View Article : Google Scholar : PubMed/NCBI

32 

Katoh M and Nakagama H: FGF receptors: Cancer biology and therapeutics. Med Res Rev. 34:280–300. 2014. View Article : Google Scholar

33 

Kotsopoulos J, Moody JR, Fan I, Rosen B, Risch HA, McLaughlin JR, Sun P and Narod SA: Height, weight, BMI and ovarian cancer survival. Gynecol Oncol. 127:83–87. 2012. View Article : Google Scholar : PubMed/NCBI

34 

Ding W, Shi W, Bellusci S, Groffen J, Heisterkamp N, Minoo P and Warburton D: Sprouty2 downregulation plays a pivotal role in mediating crosstalk between TGF-beta1 signaling and EGF as well as FGF receptor tyrosine kinase-ERK pathways in mesenchymal cells. J Cell Physiol. 212:796–806. 2007. View Article : Google Scholar : PubMed/NCBI

Related Articles

Journal Cover

November-2015
Volume 34 Issue 5

Print ISSN: 1021-335X
Online ISSN:1791-2431

Sign up for eToc alerts

Recommend to Library

Copy and paste a formatted citation
x
Spandidos Publications style
Hu L and Cong L: Fibroblast growth factor 19 is correlated with an unfavorable prognosis and promotes progression by activating fibroblast growth factor receptor 4 in advanced-stage serous ovarian cancer. Oncol Rep 34: 2683-2691, 2015.
APA
Hu, L., & Cong, L. (2015). Fibroblast growth factor 19 is correlated with an unfavorable prognosis and promotes progression by activating fibroblast growth factor receptor 4 in advanced-stage serous ovarian cancer. Oncology Reports, 34, 2683-2691. https://doi.org/10.3892/or.2015.4212
MLA
Hu, L., Cong, L."Fibroblast growth factor 19 is correlated with an unfavorable prognosis and promotes progression by activating fibroblast growth factor receptor 4 in advanced-stage serous ovarian cancer". Oncology Reports 34.5 (2015): 2683-2691.
Chicago
Hu, L., Cong, L."Fibroblast growth factor 19 is correlated with an unfavorable prognosis and promotes progression by activating fibroblast growth factor receptor 4 in advanced-stage serous ovarian cancer". Oncology Reports 34, no. 5 (2015): 2683-2691. https://doi.org/10.3892/or.2015.4212