SFRP2 Drives Aerobic Glycolysis and Tumor Progression in Ovarian Cancer via Transcriptional Upregulation of PTK2B

SFRP2 Drives Aerobic Glycolysis and Tumor Progression in Ovarian Cancer via Transcriptional Upregulation of PTK2B | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article SFRP2 Drives Aerobic Glycolysis and Tumor Progression in Ovarian Cancer via Transcriptional Upregulation of PTK2B Jindong Sheng, Yiwen Xing, Jin Luan, Xiangyu Liu, Hualin Song, and 7 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8553543/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 4 You are reading this latest preprint version Abstract Background Ovarian cancer is a highly lethal gynecologic malignancy, with metabolic reprogramming being a key contributor to its progression and therapy resistance. Although Secreted Frizzled-Related Protein 2 (SFRP2) is implicated in cancer, its functional role and molecular mechanisms in ovarian cancer, particularly in regulating metabolic pathways, remain poorly defined. Methods Bioinformatics analysis of GEO (GSE66957) and TCGA-OV datasets was performed to assess SFRP2 expression and its correlation with prognosis. Immunohistochemistry (IHC) on a human ovarian cancer tissue microarray was used for clinical validation. SFRP2 was knocked down or overexpressed in ovarian cancer cell lines (HEY, SK-OV-3) using lentiviral shRNAs. Functional assays (CCK-8, colony formation, apoptosis, migration) and metabolic assays (glucose consumption, lactate/ATP production, ECAR/OCR) were conducted. The mechanistic link between SFRP2, transcription factor CEBPA, and downstream target PTK2B was investigated using co-immunoprecipitation (Co-IP), nuclear-cytoplasmic fractionation, chromatin immunoprecipitation (ChIP), and dual-luciferase reporter assays. Rescue experiments were performed both in vitro and in vivo (xenograft mouse models). Results SFRP2 was significantly overexpressed in ovarian cancer tissues and cell lines, and high SFRP2 expression correlated with advanced disease and poor patient survival. SFRP2 knockdown suppressed cell proliferation, colony formation, and migration, while promoting apoptosis. Gene set enrichment analysis linked SFRP2 to the CCR5 signaling pathway, prompting an investigation into glycolysis. SFRP2 depletion impaired aerobic glycolysis, reducing glucose uptake, lactate/ATP production, and ECAR, while increased OCR. Conversely, SFRP2 overexpression enhanced glycolytic flux and tumorigenic phenotypes, which were abrogated by the glycolytic inhibitor 2-DG. Mechanistically, SFRP2 interacted with the transcription factor CEBPA, promoted its nuclear translocation, and enhanced its binding to the PTK2B promoter, leading to PTK2B transcriptional activation. Crucially, PTK2B knockdown reversed the pro-glycolytic and pro-tumorigenic effects of SFRP2 overexpression both in vitro and in vivo. Conclusion Our findings identify a novel SFRP2/CEBPA/PTK2B signaling axis that drives aerobic glycolysis and malignant progression in ovarian cancer, highlighting SFRP2 and PTK2B as potential prognostic markers and therapeutic targets. Ovarian Cancer SFRP2 PTK2B Aerobic Glycolysis Transcriptional Regulation Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction Ovarian cancer is the most lethal gynecological malignancy, with ~ 80% of patients presenting with stage III–IV disease and widespread peritoneal dissemination. Although most initially respond to platinum-based chemotherapy, > 85% relapse because of intrinsic or acquired chemoresistance, resulting in a 5-year survival rate below 25% and underscoring the need for new therapeutic strategies[ 1 – 2 ]. A hallmark of cancer cells, including ovarian cancer, is metabolic reprogramming, particularly the Warburg effect or aerobic glycolysis. Under aerobic conditions, cancer cells preferentially metabolize glucose to lactate even in the presence of oxygen[ 3 ]. This metabolic shift provides cells with the necessary biomass and energy to support rapid proliferation, invasion, and resistance to apoptosis. Given the critical role of metabolic reprogramming in ovarian cancer pathogenesis and treatment resistance, clarifying its upstream regulators and downstream effectors is essential[ 4 ]. SFRP2 (Secreted Frizzled-Related Protein 2), a member of the secreted frizzled-related protein family, is a secreted modulator of Wnt signalling that can inhibit canonical Wnt/β-catenin or activate non-canonical Wnt/Ca²⁺ pathways in a context-dependent manner[ 5 – 7 ]. In cancer, SFRP2 often behaves as a “double-edged sword” but predominantly exerts tumor-promoting effects by stimulating angiogenesis, metabolic reprogramming and metastasis, and has emerged as a potential therapeutic target [ 8 ] [ 5 , 9 ]. In ovarian cancer, SFRP2 has been identified as an independent poor prognostic marker[ 10 ]. Its overexpression may promote disease progression by activating the Wnt/β-catenin pathway (e.g., through interaction with the miR-96-5p/MTSS1 axis[ 11 ]) or by cooperating with other oncogenic factors such as ARPC1B[ 12 ]. Additionally, SFRP2 is involved in processes such as fibrotic transformation[ 11 ] and chemotherapy resistance[ 13 ]. However, the precise function and mechanism of SFRP2 in ovarian cancer progression, particularly its impact on aerobic glycolysis and malignant phenotypes, are largely unknown. PTK2B (Protein Tyrosine Kinase 2 Beta), a non-receptor tyrosine kinase and member of the Focal Adhesion Kinase (FAK) family, regulates cell adhesion, migration and survival and has been implicated in synaptic plasticity and tumor progression[ 14 ] [ 15 ]. In ovarian cancer, emerging evidence implicates PTK2B in disease progression through multiple pathways. Notably, PTK2B/FAK inhibitors such as PF-431396 have been shown to reverse resistant cell phenotypes and act synergistically with chemotherapy, suggesting potential for overcoming platinum resistance[ 14 ]. Additionally, PTK2B may influence the peritoneal metastatic microenvironment—where adhesion and migration are critical—by modulating tumor cell-microenvironment interactions through LPXN phosphorylation[ 14 , 16 ]. Furthermore, while PTK2B overexpression has been linked to stem cell maintenance in leukemia, analogous mechanisms could potentially sustain stemness properties in ovarian cancer stem cells (OCSCs) [ 14 , 17 ]. However, its regulation and functional contribution to ovarian cancer metabolism, particularly in glycolysis, are unknown. In this study, we initially identified SFRP2 as a significantly overexpressed gene correlated with poor survival in ovarian cancer. We systematically demonstrate that SFRP2 acts as a potent oncoprotein that drives malignant progression by enhancing aerobic glycolysis. Furthermore, we elucidate a novel transcriptional mechanism whereby SFRP2 interacts with the transcription factor CCAAT/Enhancer-Binding Protein Alpha (CEBPA) to activate PTK2B expression. Our findings establish the SFRP2/CEBPA/PTK2B axis as a critical regulator of glycolytic metabolism and tumor progression in ovarian cancer, revealing new prognostic and therapeutic opportunities. Materials and Methods Core experimental procedures (qPCR, Western blotting, Co-IP, ChIP, luciferase assays, and xenograft models) are described in the main text, whereas supporting assays and extended protocols are provided in the Supplementary Information. Bioinformatic Analysis SFRP2 expression data were downloaded from the GEO database (GSE66957). Survival analysis based on SFRP2 expression was performed using RNA-seq and clinical data from the TCGA-OV project accessed via the GDC portal. Gene Set Enrichment Analysis (GSEA) was conducted using GSEA software (v4.3.2) with the Hallmark gene sets. Tissue Microarray and IHC Ovarian cancer and adjacent normal tissues from a commercial TMA (Shanghai Outdo Biotech, Cat. No. HOvaC143Su01-M-007) were used for IHC staining of SFRP2, with ethical approval by medical ethics committee of Tianjin medical university cancer institute and hospital (Ek2022065) and informed consent. TMA slides were deparaffinized in xylene, rehydrated through graded ethanol, and subjected to antigen retrieval in citrate buffer (pH 6.0). Endogenous peroxidase was blocked with 3% H₂O₂, followed by incubation with 5% normal goat serum. Anti-SFRP2 antibody (Wuhan Sanying, Cat. No. 66328-1-Ig; 1:200) was applied overnight at 4°C, then detected with HRP-conjugated secondary antibody (Proteintech, 1:500) and DAB chromogen. Hematoxylin was used for counterstaining. Staining intensity (0–3) and positive cell percentage (0–4) were scored, with the final IRS (0–12) categorized as low (0–3), moderate (4–6), or high (7–12) expression. Cell Culture and Transfection Human ovarian cancer cell lines (HEY, SK-OV-3, HO-8910) and the normal ovarian epithelial cell line (IOSE80) were obtained from American Type Culture Collection (ATCC, Manassas, VA, USA) and cultured in RPMI-1640 medium (Gibco, Thermo Fisher Scientific, USA; Cat. No. 11875119) supplemented with 10% fetal bovine serum (FBS; Gibco; Cat. No. 10099141) and 1% penicillin-streptomycin (Gibco; Cat. No. 15140122) at 37°C in a humidified incubator with 5% CO₂. For lentiviral transduction, cells were seeded at 50–60% confluence and infected with lentiviral vectors carrying shRNAs targeting SFRP2 or PTK2B (GeneChem Co., Ltd., Shanghai, China), or overexpression plasmids for SFRP2 (GeneChem), in the presence of 5 µg/mL polybrene (Sigma-Aldrich, USA; Cat. No. TR-1003-G). Stable cell lines were selected using puromycin (2 µg/mL; InvivoGen, USA; Cat. No. ant-pr-1) for 7–10 days. The glycolysis inhibitor 2-deoxy-D-glucose (2-DG; Sigma-Aldrich; Cat. No. D8375) was dissolved in sterile PBS and used at a final concentration of 10 mM in complete culture medium. Target sequences for shRNAs against SFRP2 and PTK2B are provided in Table S1 . All experiments were performed with cells at 70–80% confluence and within 15 passages to ensure consistency. Quantitative Real-Time PCR (qPCR) Assay Total RNA was extracted using TRIzol and reverse-transcribed into cDNA. qPCR was performed using SYBR Green on a CFX96 system, and relative expression was calculated by the 2 ^−ΔΔCt method using GAPDH as control. Primer sequences are provided in Supplementary Table S2 . Western Blotting (WB) Assay Protein lysates were prepared in RIPA buffer and analyzed by SDS-PAGE followed by transfer to PVDF membranes. After incubation with primary and HRP-secondary antibodies, signals were visualized using ECL. Antibody information is listed in Supplementary Table S3 . Co-Immunoprecipitation (Co-IP) To investigate the interaction between SFRP2 (Proteintech, 55309-1-AP) and CEBPA (OmnimAbs, OM284260), Co-IP was performed. Cells were lysed in RIPA buffer with protease inhibitors. Cell lysates (500 µg) were incubated with 5 µg anti-SFRP2 antibody overnight at 4°C, followed by 20 µL protein A/G beads (Thermo, 88802) for 2 h. Normal rabbit IgG was used as a negative control. Beads were washed, and bound proteins were eluted with SDS loading buffer. Inputs and immunoprecipitates were analyzed by Western blot using anti-SFRP2 and anti-CEBPA antibodies. This method confirmed the potential interaction between SFRP2 and CEBPA. Chromatin Immunoprecipitation (ChIP) Assay ChIP was performed to analyze CEBPA binding to the PTK2B promoter using a SimpleChIP® Kit (Cell Signaling, Cat. No. 9003). Cells were cross-linked with 1% formaldehyde, and chromatin was fragmented by sonication. Chromatin (5 µg) was immunoprecipitated overnight at 4°C with 5 µg anti-CEBPA antibody (OmnimAbs, Cat. No. OM284260) or normal rabbit IgG (negative control). Immune complexes were captured with Protein G magnetic beads, washed, and eluted. Cross-links were reversed, and DNA was purified. PTK2B promoter enrichment was quantified by qPCR using primers targeting the predicted CEBPA-binding site (sequences in ​Table S2 ). Binding levels were calculated as % input. IgG controls confirmed specificity. Dual-Luciferase Reporter Assay The interaction between CEBPA and the PTK2B promoter was assessed using the dual-luciferase system (Promega, Cat. No. E1910). The PTK2B promoter fragment was cloned into pGL3-basic (Firefly luciferase), with pRL-TK (Renilla luciferase) as an internal control. HEK293T cells were co-transfected with pGL3-PTK2B-Luc, pRL-TK, and either pcDNA3.1-CEBPA (overexpression) or siCEBPA (knockdown). Firefly luciferase activity (normalized to Renilla) was measured 48 h post-transfection using a luminometer. Relative luciferase activity (%) reflected CEBPA-dependent PTK2B promoter regulation. Triplicate wells and ≥ 3 biological replicates ensured reproducibility. In Vivo Xenograft Model Female nude mice (4–6 weeks old) were purchased from Jiangsu GemPharmatech Co., Ltd. (Nanjing, China). All animal procedures were approved by the Institutional Animal Care and Use Committee (IACUC) of Stamp of The Animal Ethical and Welfare Committee of Tianjin Medical University Cancer Institute and Hospital (Approval No. [NSFC-AE-2022061]) and conducted in accordance with the Guide for the Care and Use of Laboratory Animals. For tumor xenografts, 5×10⁶ cells (resuspended in 100 µL PBS) were subcutaneously injected into the right flanks of mice (n = 6 per group). Experimental groups included: ​NC​, ​SFRP2, ​shPTK2B​, and ​SFRP2 + shPTK2B​. Tumor size was measured every 3 days using calipers, and tumor volume (mm³) was calculated as: Volume = 0.5 × length × width². After 33 days, mice were euthanized by CO₂ asphyxiation followed by cervical dislocation, and tumors were excised, weighed, and photographed. Tumor tissues were either fixed in 4% paraformaldehyde for histological analysis or snap-frozen in liquid nitrogen for further molecular studies. Statistical Analysis Data are presented as mean ± SD. Differences between groups were analyzed using Student's t-test (two groups) or one-way ANOVA (multiple groups). Correlation analysis was performed using Pearson's chi-square test. Survival curves were plotted using the Kaplan-Meier method and compared by the log-rank test. P < 0.05 was considered statistically significant. Results SFRP2 is Overexpressed in Ovarian Cancer and Correlates with Poor Prognosis To define the clinical relevance of SFRP2 in ovarian cancer, we first assessed its expression and prognostic value. Bioinformatic analysis of the GEO dataset (GSE66957) revealed that SFRP2 mRNA levels were significantly elevated in ovarian cancer tissues compared to normal ovarian tissues (Fig. 1 A). Consistent with this, analysis of the TCGA-OV cohort demonstrated that high SFRP2 expression was associated with significantly shorter overall survival (OS) (Fig. 1 B). We validated these findings at the protein level using immunohistochemistry (IHC) on a human ovarian cancer tissue microarray, which confirmed marked upregulation of SFRP2 in cancer tissues (Fig. 1 C). Furthermore, correlation analysis between SFRP2 expression and clinicopathological parameters revealed that high SFRP2 expression was positively correlated with higher T stage, lymph node metastasis (N stage), distant metastasis (M stage), larger tumor size, and advanced pathological stage (FIGO stage) (Tables 1 and 2 ). Critically, Kaplan-Meier survival analysis of our patient cohort confirmed that high SFRP2 protein expression predicted a poorer prognosis (Fig. 1 D). These data firmly establish SFRP2 as a clinically relevant oncoprotein in ovarian cancer. Table 1 Relationship between SFRP2 expression and tumor characteristics in patients with ovarian cancer Features No. of patients SFRP2 expression p value low high All patients 137 66 71 Age 0.438 ≤ 51years 70 36 34 > 51years 67 30 37 Stage P 11.9cm 68 26 42 T Infiltrate P < 0.001 T1 12 10 2 T2 34 25 9 T3 91 31 60 Lymphatic metastasis(N) 0.003 N0 105 58 47 N1 32 8 24 Metastasis 0.002 M0 112 61 51 M1 25 5 20 Table 2 Spearman correlation analysis between SFRP2 expression and tumor characteristics in patients with ovarian cancer SFRP2 T Infiltrate Spearman correlation 0.400 Significance (two-tailed) P < 0.001 N 137 Lymphatic metastasis(N) Spearman correlation 0.256 Significance (two-tailed) 0.003 N 137 Metastasis Spearman correlation 0.266 Significance (two-tailed) 0.002 N 137 Stage Spearman correlation 0.421 Significance (two-tailed) P < 0.001 N 137 Tumor size Spearman correlation 0.197 Significance (two-tailed) 0.021 N 137 Knockdown of SFRP2 Suppresses Malignant Phenotypes In Vitro To further validate the oncogenic role of SFRP2 in ovarian cancer, we performed a series of in vitro functional assays. qPCR analysis confirmed that SFRP2 mRNA was highly expressed in ovarian cancer cell lines such as HEY, SK-OV-3 compared to the normal ovarian epithelial cell line IOSE80 (Fig. 2 A). Based on these expression profiles, we selected HEY and SK-OV-3 cells for subsequent functional studies. To establish stable SFRP2-knockdown models, we designed three specific shRNAs targeting SFRP2. Transfection of these shRNAs into HEY and SK-OV-3 cells, followed by qPCR and western blot analyses, identified shSFRP2-#1 and shSFRP2-#3 as the most efficient sequences for SFRP2 knockdown (Figure S1 A-C). Functional assays demonstrated that SFRP2 knockdown significantly inhibited cell proliferation, as evidenced by CCK-8 assays (Fig. 2 B). Flow cytometric analysis showed a significant increase in the apoptosis rate in SFRP2-knockdown cells compared with the negative control (shCtrl) (Fig. 2 C). Furthermore, the clonogenic survival of SFRP2-knockdown cells was drastically impaired, as indicated by fewer and smaller colonies formed in the colony formation assay (Fig. 2 D). Cell migration, assessed by both wound healing and Transwell assays, was also markedly suppressed in SFRP2-knockdown cells compared to controls (Fig. 2 E, 2 F). These results demonstrate that SFRP2 is essential for maintaining the proliferation, survival, and migratory capacity of ovarian cancer cells. SFRP2 Drives Tumor Progression by Enhancing Aerobic Glycolysis To elucidate the mechanism by which SFRP2 drives ovarian cancer progression, we performed GSEA on TCGA-OV transcriptomic data. Intriguingly, the C-C motif chemokine receptor 5 (CCR5) signaling pathway was the most significantly enriched gene set in the SFRP2 high-expression group (Fig. 3 A). Given that CCR5 has been previously linked to tumor glycolysis[ 18 ], we hypothesized that SFRP2 might contribute to ovarian cancer progression by modulating aerobic glycolysis. To test this hypothesis, we first examined the effect of SFRP2 on key glycolytic indicators. Western blot analysis showed that knockdown of SFRP2 downregulated the expression of critical glycolytic enzymes, including PFKFB4 and LDHA (Fig. 3 B). Functionally, SFRP2 knockdown led to a significant reduction in glucose consumption, lactate production, and intracellular ATP levels (Fig. 3 C), all of which are hallmarks of glycolytic activity. Seahorse XF extracellular flux analysis further confirmed that SFRP2 depletion significantly decreased the ECAR (a measure of glycolytic flux) and increased the OCR (a measure of mitochondrial respiration) (Fig. 3 D), indicating a metabolic shift away from glycolysis. To ascertain whether glycolysis is essential for SFRP2-mediated oncogenic effects, we overexpressed SFRP2 and treated cells with the glycolytic inhibitor 2-Deoxy-D-glucose (2-DG). SFRP2 overexpression potently enhanced the expression of PKM2, HK2, GLUT1, ALDOC, ALDOA, ADH6 and LDHA (Fig. 3 E), and increased glucose uptake, lactate and ATP production, (Fig. 3 F). These metabolic changes were accompanied by a promotion of cell proliferation and colony formation, key indicators of tumorigenic potential (Fig. 3 G, H). Importantly, 2-DG treatment effectively abolished these pro-glycolytic and pro-tumorigenic effects induced by SFRP2 overexpression (Fig. 3 E-H). These data conclusively demonstrate that SFRP2 facilitates ovarian cancer progression by augmenting aerobic glycolysis. SFRP2 Promotes CEBPA-Mediated Transcriptional Upregulation of PTK2B To identify the downstream effector mediating the oncogenic function of SFRP2, we focused on the CCR5 pathway-related genes. qPCR analysis revealed that knockdown of SFRP2 significantly reduced mRNA levels of PTK2B, but not other candidates, identifying PTK2B as a key potential downstream target (Fig. 4 A). We next investigated how SFRP2 regulates PTK2B. As SFRP2 is a secreted protein, we hypothesized an indirect transcriptional mechanism. Using the STRING database to find SFRP2-interacting proteins and the hTFtarget database to predict transcription factors for the PTK2B promoter, we identified CEBPA as a common candidate (data not shown). This hypothesis was supported by Co-IP experiments, which confirmed a direct physical interaction between endogenous SFRP2 and CEBPA proteins in ovarian cancer cells (Fig. 4 B). Notably, SFRP2 knockdown markedly reduced the nuclear accumulation of CEBPA, as shown by nuclear-cytoplasmic fractionation followed by WB (Fig. 4 C), suggesting SFRP2 facilitates CEBPA's nuclear translocation. We then examined whether the SFRP2/CEBPA complex regulates PTK2B transcription. ChIP assays demonstrated that SFRP2 overexpression significantly enhanced the enrichment of CEBPA on the PTK2B promoter region (Fig. 4 D). Consistent with this, dual-luciferase reporter assays showed that co-expression of SFRP2 and CEBPA synergistically activated the activity of a luciferase reporter driven by the PTK2B promoter (Fig. 4 E). Taken together, these results delineate a novel mechanism whereby SFRP2 binds to CEBPA, promotes its nuclear translocation, and enhances its transcriptional activation of PTK2B. The SFRP2/PTK2B Axis Promotes Malignant Progression through Aerobic Glycolysis Finally, we conducted rescue experiments to confirm that PTK2B is the critical functional mediator of SFRP2. We established SFRP2-overexpressing HEY and SK-OV-3 cells, in which PTK2B was simultaneously knocked down (Figure S1 D-E). SFRP2 overexpression significantly promoted cell proliferation and colony formation, phenotypes that were effectively reversed upon PTK2B knockdown (Fig. 5 A, B). These results indicate that PTK2B is indispensable for SFRP2-driven proliferative and clonogenic capacities. We next extended these findings to an in vivo xenograft mouse model to assess the functional relevance of the SFRP2-PTK2B axis in tumor growth. Tumors derived from SFRP2-overexpressing cells exhibited significantly accelerated growth kinetics, resulting in markedly larger and heavier tumor masses compared to those derived from control cells. Strikingly, co-knockdown of PTK2B in SFRP2-overexpressing cells largely abolished these pro-tumorigenic effects, leading to markedly impaired tumor growth (Fig. 5 C-E). Western blot analysis of tumor lysates showed that SFRP2 overexpression upregulated the glycolytic enzyme PKM2, and this effect was again abolished by PTK2B knockdown (Fig. 5 F). Furthermore, IHC analysis of tumor sections revealed that SFRP2 overexpression increased the expression of PTK2B and the proliferation marker Ki-67, which was attenuated by PTK2B knockdown (Fig. 5 E). These in vivo data robustly validate that SFRP2 exerts its pro-tumorigenic and pro-glycolytic effects primarily through PTK2B, underscoring the critical role of the SFRP2-PTK2B axis in deriving ovarian cancer progression and metabolic reprogramming. Discussion This study identifies a new oncogenic pathway—SFRP2/CEBPA/PTK2B—that drives malignant progression by activating aerobic glycolysis. We provide evidence that SFRP2, beyond its canonical roles, exerts an intracellular function by directly binding CEBPA, facilitating its nuclear translocation and co-activating PTK2B expression. The subsequent upregulation of PTK2B is indispensable for glycolytic flux and aggressive tumor phenotypes driven by SFRP2, as validated in vitro and in vivo . SFRP2 is a key regulator of the Wnt signaling pathway and plays complex roles in various cancers. In colorectal cancer, SFRP2 activates enolase 2 (ENO2) through the TCF4/β-catenin axis, significantly promoting glycolysis and metastatic phenotypes. Inhibition of ENO2 reverses SFRP2-induced metastasis, and co-expression of SFRP2/ENO2 is associated with poorer survival and higher recurrence rates[ 19 ]. In non-small cell lung cancer, m6A-methylated SFRP2 is recognized and interacts with YTHDF2; knockdown of SFRP2 suppresses glycolysis and cancer stem cell properties[ 20 ]. Current research on SFRP2 in ovarian cancer still has several gaps, such as its interaction with known glycolytic regulators (e.g., HIF-1α, FOXK2 [21–22] ) and its subtype-specific roles in different ovarian cancer subtypes[ 23 ]. Existing evidence suggests that targeting SFRP2 and its downstream glycolytic pathways may offer new strategies for improving ovarian cancer prognosis[ 19 , 24 ]. Our findings robustly position SFRP2 as an oncogene in ovarian cancer, consistently demonstrating that its high expression correlates with advanced disease and poor survival. We extend the understanding of SFRP2's oncogenic functions by establishing its critical role in promoting proliferation, clonogenicity, migration, and anti-apoptosis in ovarian cancer cells. More importantly, we uncover a previously unrecognized function of SFRP2 in regulating cancer cell metabolism. While a recent study suggested SFRP2's involvement in metabolism in other contexts[ 19 ], its direct role in driving aerobic glycolysis, the Warburg effect, in ovarian cancer is a novel finding. Our data demonstrate that SFRP2 significantly enhances glycolytic flux, and this metabolic reprogramming is essential for its tumor-promoting effects, as inhibition of glycolysis abrogated SFRP2-driven oncogenic phenotypes. Our GSEA hinting at the CCR5 pathway was crucial, as it provided a rationale to connect SFRP2 to glycolysis, leveraging existing literature on CCR5's metabolic roles [ 25 ]. This led us to investigate downstream effectors. Among CCR5 pathway-related candidates, PTK2B emerged as a key target transcriptionally regulated by SFRP2. PTK2B is a non-receptor tyrosine kinase known for its roles in cell adhesion, migration, and survival in various cancers[ 26 – 28 ]. Emerging evidence has begun to link PTK2B to metabolic regulation. For instance, the interaction between PTK2B and SIRPα in macrophages regulates both autophagy and necroptosis, processes closely linked to cellular metabolic reprogramming such as glycolysis and mitochondrial function, further supporting the role of PTK2B in immune-metabolic crosstalk[ 29 ]. However, its role in ovarian cancer metabolism, particularly in glycolysis, remained unexplored. Our study clearly demonstrates that PTK2B is a critical mediator of SFRP2-induced aerobic glycolysis and tumor progression in ovarian cancer. Knockdown of PTK2B reversed the pro-glycolytic and pro-tumorigenic effects of SFRP2 both in vitro and in vivo . This aligns with and extends the emerging concept of PTK2B as a metabolic regulator in cancer, providing a direct link between SFRP2 signaling and PTK2B-dependent glycolytic reprogramming. The transcriptional mechanism delineated here is particularly intriguing. SFRP2, typically considered a secreted Wnt antagonist, is shown to have an intracellular role by directly interacting with CEBPA and influencing its subcellular localization and transcriptional activity. This non-canonical, intracellular function expands the paradigm of SFRP2's actions in cancer and suggests potential novel therapeutic strategies aimed at disrupting this protein-protein interaction. CEBPA is a transcription factor involved in differentiation and metabolism[ 30 ]. Our ChIP and luciferase reporter assays confirm that the SFRP2/CEBPA complex directly binds to and activates the PTK2B promoter, establishing a linear pathway from SFRP2 to PTK2B upregulation. In summary, our work delineates a previously unrecognized SFRP2/CEBPA/PTK2B axis that is critical for aerobic glycolysis and malignant progression in ovarian cancer. These findings not only deepen our understanding of the molecular mechanisms driving metabolic reprogramming in this deadly disease but also highlight SFRP2 and PTK2B as promising prognostic biomarkers and attractive therapeutic targets. Targeting this axis, perhaps with monoclonal antibodies against SFRP2 or small molecule inhibitors of PTK2B, could represent a novel metabolic therapy for ovarian cancer patients. Declarations Ethics approval and consent to participate We hereby confirm that all experimental procedures involving mice in this study were reviewed and approved by the Institutional Animal Care and Use Committee (IACUC) of Tianjin Medical University Cancer Institute and Hospital (Approval No. NSFC-AE-2022061). All procedures were strictly conducted in accordance with the approved protocol and internationally recognized guidelines for the care and use of laboratory animals. Consent for publication Not applicable. This study involved experimental animals only and did not include any human participants or individual person’s data. Competing interests The authors declare no conflict of interest. Funding This work was supported by the National Natural Science Foundation of China (No. 82202863, Jindong Sheng; No. 82272946, Ke Wang). Author contribution J. S., K. W. and W. L. designed this program. J. S., Y. X., J. L., X. L., H. S., L. S., M. Y. operated the cell and animal experiments. J. S., Y. X., X. F., H. W. and Y. C. conducted the data collection and analysis. J. S. and Y. X. produced the manuscript which was checked by J. S., K. W. and W. L. All the authors have confirmed the submission of this manuscript. Acknowledgements The authors gratefully acknowledge the support from the National Natural Science Foundation of China. Availability of data and materials The data that support the findings of this study are available from the corresponding author upon reasonable request. References Caruso G, Weroha SJ, Cliby W. Ovarian Cancer: A Review. JAMA. 2025;334:1278–91. https://doi.org/10.1001/jama.2025.9495 . Webb PM, Jordan SJ. Global epidemiology of epithelial ovarian cancer. Nat Rev Clin Oncol. 2024;21:389–400. https://doi.org/10.1038/s41571-024-00881-3 . Liao M, Yao D, Wu L, Luo C, Wang Z, Zhang J, et al. Targeting the Warburg effect: A revisited perspective from molecular mechanisms to traditional and innovative therapeutic strategies in cancer. Acta Pharm Sin B. 2024;14:953–1008. https://doi.org/10.1016/j.apsb.2023.12.003 . Zhang Y, Wang Y, Zhao G, Orsulic S, Matei D. Metabolic dependencies and targets in ovarian cancer. 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The significance of proline-rich tyrosine kinase2 (Pyk2) on hepatocellular carcinoma progression and recurrence. Br J Cancer. 2007;97:50–7. https://doi.org/10.1038/sj.bjc.6603827 . Wang D, Lin Y, Xu F, Zhang H, Zhu X, Liu Z, et al. SIRPα maintains macrophage homeostasis by interacting with PTK2B kinase in Mycobacterium tuberculosis infection and through autophagy and necroptosis. EBioMedicine. 2022;85:104278. https://doi.org/10.1016/j.ebiom.2022.104278 . Johnson PF. Molecular stop signs: regulation of cell-cycle arrest by C/EBP transcription factors. J Cell Sci. 2005;118:2545–55. https://doi.org/10.1242/jcs.02459 . Supplementary Files FigS1.tif Supplementary Figure 1. Validation of SFRP2 knockdown and rescue models in ovarian cancer cells. (A) Relative SFRP2 mRNA levels in HEY cells transfected with control (CON), shCtrl, or three distinct shRNA constructs targeting SFRP2 (shSFRP2-1, -2, -3). qRT-PCR shows significant reduction in SFRP2 expression by shSFRP2-1 and shSFRP2-3 compared to control. **p < 0.01, ***p < 0.001. (B) Relative SFRP2 mRNA levels in HEY and SK-OV-3 cells transfected with shCtrl, shSFRP2-1, or shSFRP2-3. Both shSFRP2-1 and shSFRP2-3 effectively reduce SFRP2 expression in both cell lines. *p < 0.05, ***p < 0.001. (C) Western blot analysis of SFRP2 protein levels in HEY and SK-OV-3 cells after transfection with shCtrl, shSFRP2-1, or shSFRP2-3. GAPDH serves as loading control. Both shSFRP2-1 and shSFRP2-3 significantly reduce SFRP2 protein expression, confirming efficient knockdown at the protein level. (D) Relative SFRP2 and PTK2B mRNA levels in HEY and SK-OV-3 cells following SFRP2 overexpression (SFRP2 vs. NC) or PTK2B knockdown (shPTK2B vs. shCtrl). SFRP2 overexpression increases SFRP2 mRNA levels, while shPTK2B efficiently reduces PTK2B expression. GAPDH serves as loading control. **p < 0.01, ***p < 0.001. (E) Western blot analysis of SFRP2 and PTK2B protein levels in HEY and SK-OV-3 cells following indicated plasmids transfection. GAPDH serves as loading control. MaterialsandMethods.docx TableS1.docx TableS2.docx TableS3.docx Cite Share Download PDF Status: Under Review Version 1 posted Reviewers agreed at journal 13 Jan, 2026 Reviewers invited by journal 13 Jan, 2026 Editor assigned by journal 10 Jan, 2026 First submitted to journal 09 Jan, 2026 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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1","display":"","copyAsset":false,"role":"figure","size":7198076,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSFRP2 is highly expressed in ovarian cancer and correlates with poor prognosis.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(A) \u003c/strong\u003eBoxplot showing SFRP2 mRNA expression levels in ovarian cancer (Tumor) versus adjacent non-tumor tissues (Normal) based on RNA-seq data. Expression values are log₂-transformed and normalized (log₂(normalized counts + 1)). Each dot represents an individual sample.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(B) \u003c/strong\u003eKaplan–Meier survival curve illustrating overall survival (OS) in patients stratified by SFRP2 expression level (high vs. low). High SFRP2 expression (n = 60) is associated with significantly shorter OS compared to low expression (n = 239). Hazard ratio (HR) = 1.57 (95% CI: 1.12–2.21), Log-rank p = 0.00819. Numbers at risk are indicated below the curve.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(C)\u003c/strong\u003eRepresentative immunohistochemical (IHC) staining of SFRP2 in ovarian cancer tissues at different stages (II, III, IV). Staining intensity increases with advancing tumor stage. Images are shown at 200× and 400× magnification. Brown staining indicates positive SFRP2 expression. Scale bar=50 μm.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(D)\u003c/strong\u003eKaplan–Meier survival analysis of overall survival in ovarian cancer patients grouped by SFRP2 protein expression (low vs. high) based on IHC results. Patients with high SFRP2 expression exhibit significantly worse survival outcomes (Log-rank p \u0026lt; 0.001).\u003c/p\u003e","description":"","filename":"OnlineFig11.png","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/88a3256d660b3de7d9bd5de8.png"},{"id":100421863,"identity":"3bd63039-b2bf-4335-a0d2-c1a436ecd49c","added_by":"auto","created_at":"2026-01-16 13:58:31","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":830399,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eKnockdown of SFRP2 suppresses malignant phenotypes of ovarian cancer cells.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(A)\u003c/strong\u003eQuantitative real-time PCR analysis of SFRP2 mRNA expression in human ovarian epithelial cell line IOSE80 and ovarian cancer cell lines HEY, SK-OV-3 and HO-8910 cells. Data are presented as mean ± SD (n = 3). *p \u0026lt; 0.05, **p \u0026lt; 0.01 vs. IOSE80.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(B)\u003c/strong\u003eCell proliferation assays performed using CCK-8 kit in HEY and SK-OV-3 cells transfected with shCtrl, shSFRP2-1, or shSFRP2-3. Optical density (OD) at 450 nm was measured daily over 5 days. SFRP2 knockdown significantly reduced cell proliferation compared to control. **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(C)\u003c/strong\u003eApoptosis analysis by flow cytometry using Annexin V-FITC/PI staining in HEY and SK-OV-3 cells after SFRP2 knockdown. Representative dot plots show early (Q4) and late (Q3) apoptotic populations. Bar graphs summarize percentage of apoptotic cells. SFRP2 silencing increased apoptosis in both cell lines. *p \u0026lt; 0.05, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(D)\u003c/strong\u003eColony formation assay assessing long-term proliferative capacity. Representative images and quantification of colonies formed by HEY and SK-OV-3 cells following SFRP2 knockdown. Knockdown of SFRP2 significantly reduced colony number. ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(E)\u003c/strong\u003eWound healing assay evaluating cell migration in HEY and SK-OV-3 cells. Representative images at 0 and 24 hours post-wounding. Migration rate was calculated as the ratio of wound closure at 24 h to initial wound width. SFRP2 depletion significantly impaired migratory ability. *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(F)\u003c/strong\u003eTranswell migration assay assessing invasive potential. Representative images of migrated cells stained with crystal violet. Quantification shows fold change (FC) in migrated cell numbers relative to control. SFRP2 knockdown markedly reduced cell migration. ***p \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"OnlineFig21.png","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/19aab86268c7929e0049d6fa.png"},{"id":100408630,"identity":"ff9cffea-7ff8-47e4-9e67-da2b6a3f2ed2","added_by":"auto","created_at":"2026-01-16 13:06:22","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":91993,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSFRP2 promotes ovarian cancer progression by enhancing aerobic glycolysis.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(A) Gene set enrichment analysis (GSEA) of TCGA-OV transcriptomic data showing significant enrichment of the \"HALLMARK_CCR5_SIGNALING\" gene set in tumors with high SFRP2 expression (normalized enrichment score, NES = 2.18; false discovery rate, FDR q-value \u0026lt; 0.001). The leading-edge analysis heatmap displays expression of core-enriched genes in the CCR5 pathway.\u003c/p\u003e\n\u003cp\u003e(B) Western blot analysis of key glycolytic proteins PFKFB4 and LDHA in SK-OV-3 and HEY cells following SFRP2 knockdown (shSFRP2) or control (shCtrl). Actin serves as loading control. SFRP2 silencing reduces protein expression of PFKFB4 and LDHA.\u003c/p\u003e\n\u003cp\u003e(C) Metabolic assays measuring glucose consumption, lactate production, and ATP levels in HEY and SK-OV-3 cells after SFRP2 knockdown. Glucose uptake and lactate release are significantly reduced, while ATP levels are markedly decreased in shSFRP2 cells compared to controls. *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(D) Real-time extracellular flux analysis assessing oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) in HEY and SK-OV-3 cells. OCR reflects mitochondrial respiration, while ECAR indicates glycolytic activity. SFRP2 knockdown increases OCR (indicating enhanced oxidative phosphorylation) and decreases ECAR (indicating suppressed glycolysis), consistent with metabolic shift away from aerobic glycolysis.\u003c/p\u003e\n\u003cp\u003e(E) Western blot analysis of glycolytic enzymes in HEY and SK-OV-3 cells treated with SFRP2 overexpression (SFRP2), SFRP2 + 2-DG (a glycolysis inhibitor), or negative control (NC). Expression of PKM2, HK2, GLUT1, ALDOC, ALDOA, ADH6, and LDHA is increased by SFRP2 overexpression but attenuated by 2-DG co-treatment. β-Actin serves as loading control.\u003c/p\u003e\n\u003cp\u003e(F) Metabolic phenotyping of HEY and SK-OV-3 cells under SFRP2 overexpression or SFRP2 + 2-DG treatment. Glucose consumption, ATP production, and lactate generation are significantly elevated by SFRP2 overexpression and reversed by 2-DG. *p \u0026lt; 0.05, **p \u0026lt; 0.01.\u003c/p\u003e\n\u003cp\u003e(G) Cell proliferation assays using CCK-8 kit in HEY and SK-OV-3 cells transfected with NC, SFRP2, or SFRP2 + 2-DG. Overexpression of SFRP2 enhances proliferation, which is significantly blocked by 2-DG treatment. *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(H) Colony formation assay showing long-term proliferative capacity in HEY and SK-OV-3 cells under the same conditions. SFRP2 overexpression increases colony number, which is abolished by co-treatment with 2-DG. ***p \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"OnlineFig31.png","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/fc2e74f590ffd75d9ddd5ffc.png"},{"id":100408553,"identity":"cf7bc0fb-ebd1-4f25-a852-adad10ab0638","added_by":"auto","created_at":"2026-01-16 13:06:20","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":3326444,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSFRP2 promotes CEBPA-mediated transcriptional upregulation of PTK2B.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(A) Quantitative real-time PCR analysis of mRNA levels for SFRP2, CXCL12, CCL2, PLCG1, and PTK2B in HEY and SK-OV-3 cells following SFRP2 knockdown (shSFRP2) or control (shCtrl). Knockdown of SFRP2 significantly reduces PTK2B expression. *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(B) Co-immunoprecipitation (Co-IP) assay demonstrating a direct physical interaction between endogenous SFRP2 and CEBPA proteins in SK-OV-3 and HEY cells. Cell lysates were immunoprecipitated with anti-SFRP2 antibody or IgG control, followed by Western blotting for CEBPA and SFRP2.\u003c/p\u003e\n\u003cp\u003e(C) Nuclear-cytoplasmic fractionation followed by Western blotting showing subcellular localization of CEBPA in HEY and SK-OV-3 cells after SFRP2 knockdown. α-Tubulin and histone H3 serve as cytoplasmic and nuclear loading controls, respectively.\u003c/p\u003e\n\u003cp\u003e(D) Chromatin immunoprecipitation (ChIP) assay assessing the enrichment of CEBPA on the PTK2B promoter region in HEY and SK-OV-3 cells overexpressing SFRP2 (SFRP2 OE) or control (NC). qPCR amplification of the PTK2B promoter region shows increased CEBPA binding in SFRP2-overexpressing cells compared to control. **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(E) Dual-luciferase reporter assays evaluating the transcriptional activity of the PTK2B promoter. HEY and SK-OV-3 cells were co-transfected with wild-type (wt) or mutant (mut) PTK2B promoter luciferase constructs, along with SFRP2, CEBPA, or empty vector controls. SFRP2 overexpression enhances PTK2B promoter activity only when CEBPA is co-expressed, and this activation is abolished in the mutant promoter. ***p \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"OnlineFig4.png","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/b5bb55b0af232144df42e95f.png"},{"id":100408227,"identity":"5af9cf1b-cc94-4161-8ef3-21ade340e31a","added_by":"auto","created_at":"2026-01-16 13:05:48","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":972830,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe SFRP2/PTK2B signaling axis promotes malignant progression through aerobic glycolysis.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(A) CCK-8 showing growth kinetics of HEY and SK-OV-3 cells transfected with control (NC), SFRP2 overexpression (SFRP2), PTK2B knockdown (shPTK2B), or dual manipulation (SFRP2 + shPTK2B). SFRP2 overexpression significantly enhances cell proliferation, an effect reversed by co-knockdown of PTK2B. *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(B) Colony formation assay demonstrating long-term proliferative capacity. Representative images and quantification show that SFRP2 overexpression increases colony number, which is abrogated by PTK2B knockdown. *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(C) Representative image of subcutaneous xenograft tumors derived from HEY cells expressing NC, shPTK2B, SFRP2, or SFRP2 + shPTK2B after 33 days of inoculation in nude mice.\u003c/p\u003e\n\u003cp\u003e(D) Tumor volume measured over time post-inoculation. SFRP2-overexpressing tumors grow significantly faster than controls, and this acceleration is blocked by PTK2B knockdown. **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(E) Tumor weight at endpoint. SFRP2 overexpression results in significantly heavier tumors compared to control, and this effect is rescued by PTK2B knockdown. **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(F) Western blot analysis of tumor lysates showing protein levels of SFRP2 and PKM2. SFRP2 overexpression upregulates PKM2 expression, an effect abolished by PTK2B knockdown. GAPDH serves as loading control.\u003c/p\u003e\n\u003cp\u003e(G) IHC staining of tumor sections for Ki-67 (proliferation marker), PTK2B, and SFRP2. SFRP2 overexpression increases Ki-67 and PTK2B expression, while co-knockdown of PTK2B attenuates these signals.Scale bar=50 μm.\u003c/p\u003e","description":"","filename":"OnlineFig51.png","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/c0057f5dd8b8977550d354a3.png"},{"id":100546431,"identity":"9fd65900-dc7d-46f0-9c5a-6f4b62e25d3a","added_by":"auto","created_at":"2026-01-19 08:08:35","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":4293737,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/b592aed9-ea85-45f6-9127-91b1d34a5690.pdf"},{"id":100408532,"identity":"b8f652c1-3273-448f-8300-bd1f170af1b9","added_by":"auto","created_at":"2026-01-16 13:06:19","extension":"tif","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":5345228,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Figure 1.\u003c/strong\u003e \u003cstrong\u003eValidation of SFRP2 knockdown and rescue models in ovarian cancer cells.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(A) Relative SFRP2 mRNA levels in HEY cells transfected with control (CON), shCtrl, or three distinct shRNA constructs targeting SFRP2 (shSFRP2-1, -2, -3). qRT-PCR shows significant reduction in SFRP2 expression by shSFRP2-1 and shSFRP2-3 compared to control. **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(B) Relative SFRP2 mRNA levels in HEY and SK-OV-3 cells transfected with shCtrl, shSFRP2-1, or shSFRP2-3. Both shSFRP2-1 and shSFRP2-3 effectively reduce SFRP2 expression in both cell lines. *p \u0026lt; 0.05, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(C) Western blot analysis of SFRP2 protein levels in HEY and SK-OV-3 cells after transfection with shCtrl, shSFRP2-1, or shSFRP2-3. GAPDH serves as loading control. Both shSFRP2-1 and shSFRP2-3 significantly reduce SFRP2 protein expression, confirming efficient knockdown at the protein level.\u003c/p\u003e\n\u003cp\u003e(D) Relative SFRP2 and PTK2B mRNA levels in HEY and SK-OV-3 cells following SFRP2 overexpression (SFRP2 vs. NC) or PTK2B knockdown (shPTK2B vs. shCtrl). SFRP2 overexpression increases SFRP2 mRNA levels, while shPTK2B efficiently reduces PTK2B expression. GAPDH serves as loading control. **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e\n\u003cp\u003e(E) Western blot analysis of SFRP2 and PTK2B protein levels in HEY and SK-OV-3 cells following indicated plasmids transfection. GAPDH serves as loading control.\u003c/p\u003e","description":"","filename":"FigS1.tif","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/ae9a765d3a19b8bdeb3e44af.tif"},{"id":100408203,"identity":"26f17eda-7f7f-433b-aa97-3c52273046ed","added_by":"auto","created_at":"2026-01-16 13:05:44","extension":"docx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":16472,"visible":true,"origin":"","legend":"","description":"","filename":"MaterialsandMethods.docx","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/745c7a1ef9d4bca21cc86130.docx"},{"id":100408583,"identity":"f7f5d281-c64c-40f2-a651-9840ed225858","added_by":"auto","created_at":"2026-01-16 13:06:21","extension":"docx","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":15932,"visible":true,"origin":"","legend":"","description":"","filename":"TableS1.docx","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/4cabe703d20adefc7e449737.docx"},{"id":100408192,"identity":"9d562e43-c66d-4dc7-a50d-760ed75a354b","added_by":"auto","created_at":"2026-01-16 13:05:42","extension":"docx","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":16207,"visible":true,"origin":"","legend":"","description":"","filename":"TableS2.docx","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/6a33c8cc3af62d598c374140.docx"},{"id":100408368,"identity":"ab4b1fa0-a098-46e2-ac7c-59bc15af430b","added_by":"auto","created_at":"2026-01-16 13:06:03","extension":"docx","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":17279,"visible":true,"origin":"","legend":"","description":"","filename":"TableS3.docx","url":"https://assets-eu.researchsquare.com/files/rs-8553543/v1/a7d2bf81ca8228ebda114656.docx"}],"financialInterests":"","formattedTitle":"SFRP2 Drives Aerobic Glycolysis and Tumor Progression in Ovarian Cancer via Transcriptional Upregulation of PTK2B","fulltext":[{"header":"Introduction","content":"\u003cp\u003eOvarian cancer is the most lethal gynecological malignancy, with ~\u0026thinsp;80% of patients presenting with stage III\u0026ndash;IV disease and widespread peritoneal dissemination. Although most initially respond to platinum-based chemotherapy, \u0026gt;\u0026thinsp;85% relapse because of intrinsic or acquired chemoresistance, resulting in a 5-year survival rate below 25% and underscoring the need for new therapeutic strategies[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. A hallmark of cancer cells, including ovarian cancer, is metabolic reprogramming, particularly the Warburg effect or aerobic glycolysis. Under aerobic conditions, cancer cells preferentially metabolize glucose to lactate even in the presence of oxygen[\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. This metabolic shift provides cells with the necessary biomass and energy to support rapid proliferation, invasion, and resistance to apoptosis. Given the critical role of metabolic reprogramming in ovarian cancer pathogenesis and treatment resistance, clarifying its upstream regulators and downstream effectors is essential[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eSFRP2 (Secreted Frizzled-Related Protein 2), a member of the secreted frizzled-related protein family, is a secreted modulator of Wnt signalling that can inhibit canonical Wnt/β-catenin or activate non-canonical Wnt/Ca\u0026sup2;⁺ pathways in a context-dependent manner[\u003cspan additionalcitationids=\"CR6\" citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. In cancer, SFRP2 often behaves as a \u0026ldquo;double-edged sword\u0026rdquo; but predominantly exerts tumor-promoting effects by stimulating angiogenesis, metabolic reprogramming and metastasis, and has emerged as a potential therapeutic target [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e] [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. In ovarian cancer, SFRP2 has been identified as an independent poor prognostic marker[\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Its overexpression may promote disease progression by activating the Wnt/β-catenin pathway (e.g., through interaction with the miR-96-5p/MTSS1 axis[\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]) or by cooperating with other oncogenic factors such as ARPC1B[\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Additionally, SFRP2 is involved in processes such as fibrotic transformation[\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e] and chemotherapy resistance[\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. However, the precise function and mechanism of SFRP2 in ovarian cancer progression, particularly its impact on aerobic glycolysis and malignant phenotypes, are largely unknown.\u003c/p\u003e \u003cp\u003ePTK2B (Protein Tyrosine Kinase 2 Beta), a non-receptor tyrosine kinase and member of the Focal Adhesion Kinase (FAK) family, regulates cell adhesion, migration and survival and has been implicated in synaptic plasticity and tumor progression[\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e] [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. In ovarian cancer, emerging evidence implicates PTK2B in disease progression through multiple pathways. Notably, PTK2B/FAK inhibitors such as PF-431396 have been shown to reverse resistant cell phenotypes and act synergistically with chemotherapy, suggesting potential for overcoming platinum resistance[\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Additionally, PTK2B may influence the peritoneal metastatic microenvironment\u0026mdash;where adhesion and migration are critical\u0026mdash;by modulating tumor cell-microenvironment interactions through LPXN phosphorylation[\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Furthermore, while PTK2B overexpression has been linked to stem cell maintenance in leukemia, analogous mechanisms could potentially sustain stemness properties in ovarian cancer stem cells (OCSCs) [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. However, its regulation and functional contribution to ovarian cancer metabolism, particularly in glycolysis, are unknown.\u003c/p\u003e \u003cp\u003eIn this study, we initially identified SFRP2 as a significantly overexpressed gene correlated with poor survival in ovarian cancer. We systematically demonstrate that SFRP2 acts as a potent oncoprotein that drives malignant progression by enhancing aerobic glycolysis. Furthermore, we elucidate a novel transcriptional mechanism whereby SFRP2 interacts with the transcription factor CCAAT/Enhancer-Binding Protein Alpha (CEBPA) to activate PTK2B expression. Our findings establish the SFRP2/CEBPA/PTK2B axis as a critical regulator of glycolytic metabolism and tumor progression in ovarian cancer, revealing new prognostic and therapeutic opportunities.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003eCore experimental procedures (qPCR, Western blotting, Co-IP, ChIP, luciferase assays, and xenograft models) are described in the main text, whereas supporting assays and extended protocols are provided in the Supplementary Information.\u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eBioinformatic Analysis\u003c/h2\u003e \u003cp\u003eSFRP2 expression data were downloaded from the GEO database (GSE66957). Survival analysis based on SFRP2 expression was performed using RNA-seq and clinical data from the TCGA-OV project accessed via the GDC portal. Gene Set Enrichment Analysis (GSEA) was conducted using GSEA software (v4.3.2) with the Hallmark gene sets.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eTissue Microarray and IHC\u003c/h3\u003e\n\u003cp\u003eOvarian cancer and adjacent normal tissues from a commercial TMA (Shanghai Outdo Biotech, Cat. No. HOvaC143Su01-M-007) were used for IHC staining of SFRP2, with ethical approval by medical ethics committee of Tianjin medical university cancer institute and hospital (Ek2022065) and informed consent. TMA slides were deparaffinized in xylene, rehydrated through graded ethanol, and subjected to antigen retrieval in citrate buffer (pH 6.0). Endogenous peroxidase was blocked with 3% H₂O₂, followed by incubation with 5% normal goat serum. Anti-SFRP2 antibody (Wuhan Sanying, Cat. No. 66328-1-Ig; 1:200) was applied overnight at 4\u0026deg;C, then detected with HRP-conjugated secondary antibody (Proteintech, 1:500) and DAB chromogen. Hematoxylin was used for counterstaining. Staining intensity (0\u0026ndash;3) and positive cell percentage (0\u0026ndash;4) were scored, with the final IRS (0\u0026ndash;12) categorized as low (0\u0026ndash;3), moderate (4\u0026ndash;6), or high (7\u0026ndash;12) expression.\u003c/p\u003e\n\u003ch3\u003eCell Culture and Transfection\u003c/h3\u003e\n\u003cp\u003eHuman ovarian cancer cell lines (HEY, SK-OV-3, HO-8910) and the normal ovarian epithelial cell line (IOSE80) were obtained from American Type Culture Collection (ATCC, Manassas, VA, USA) and cultured in RPMI-1640 medium (Gibco, Thermo Fisher Scientific, USA; Cat. No. 11875119) supplemented with 10% fetal bovine serum (FBS; Gibco; Cat. No. 10099141) and 1% penicillin-streptomycin (Gibco; Cat. No. 15140122) at 37\u0026deg;C in a humidified incubator with 5% CO₂. For lentiviral transduction, cells were seeded at 50\u0026ndash;60% confluence and infected with lentiviral vectors carrying shRNAs targeting SFRP2 or PTK2B (GeneChem Co., Ltd., Shanghai, China), or overexpression plasmids for SFRP2 (GeneChem), in the presence of 5 \u0026micro;g/mL polybrene (Sigma-Aldrich, USA; Cat. No. TR-1003-G). Stable cell lines were selected using puromycin (2 \u0026micro;g/mL; InvivoGen, USA; Cat. No. ant-pr-1) for 7\u0026ndash;10 days. The glycolysis inhibitor 2-deoxy-D-glucose (2-DG; Sigma-Aldrich; Cat. No. D8375) was dissolved in sterile PBS and used at a final concentration of 10 mM in complete culture medium. Target sequences for shRNAs against SFRP2 and PTK2B are provided in Table \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e. All experiments were performed with cells at 70\u0026ndash;80% confluence and within 15 passages to ensure consistency.\u003c/p\u003e\n\u003ch3\u003eQuantitative Real-Time PCR (qPCR) Assay\u003c/h3\u003e\n\u003cp\u003eTotal RNA was extracted using TRIzol and reverse-transcribed into cDNA. qPCR was performed using SYBR Green on a CFX96 system, and relative expression was calculated by the 2\u003csup\u003e^\u0026minus;ΔΔCt\u003c/sup\u003e method using GAPDH as control. Primer sequences are provided in Supplementary Table \u003cspan refid=\"MOESM2\" class=\"InternalRef\"\u003eS2\u003c/span\u003e.\u003c/p\u003e\n\u003ch3\u003eWestern Blotting (WB) Assay\u003c/h3\u003e\n\u003cp\u003eProtein lysates were prepared in RIPA buffer and analyzed by SDS-PAGE followed by transfer to PVDF membranes. After incubation with primary and HRP-secondary antibodies, signals were visualized using ECL. Antibody information is listed in Supplementary Table \u003cspan refid=\"MOESM3\" class=\"InternalRef\"\u003eS3\u003c/span\u003e.\u003c/p\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eCo-Immunoprecipitation (Co-IP)\u003c/h2\u003e \u003cp\u003eTo investigate the interaction between SFRP2 (Proteintech, 55309-1-AP) and CEBPA (OmnimAbs, OM284260), Co-IP was performed. Cells were lysed in RIPA buffer with protease inhibitors. Cell lysates (500 \u0026micro;g) were incubated with 5 \u0026micro;g anti-SFRP2 antibody overnight at 4\u0026deg;C, followed by 20 \u0026micro;L protein A/G beads (Thermo, 88802) for 2 h. Normal rabbit IgG was used as a negative control. Beads were washed, and bound proteins were eluted with SDS loading buffer. Inputs and immunoprecipitates were analyzed by Western blot using anti-SFRP2 and anti-CEBPA antibodies. This method confirmed the potential interaction between SFRP2 and CEBPA.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eChromatin Immunoprecipitation (ChIP) Assay\u003c/h3\u003e\n\u003cp\u003eChIP was performed to analyze CEBPA binding to the PTK2B promoter using a SimpleChIP\u0026reg; Kit (Cell Signaling, Cat. No. 9003). Cells were cross-linked with 1% formaldehyde, and chromatin was fragmented by sonication. Chromatin (5 \u0026micro;g) was immunoprecipitated overnight at 4\u0026deg;C with 5 \u0026micro;g anti-CEBPA antibody (OmnimAbs, Cat. No. OM284260) or normal rabbit IgG (negative control). Immune complexes were captured with Protein G magnetic beads, washed, and eluted. Cross-links were reversed, and DNA was purified. PTK2B promoter enrichment was quantified by qPCR using primers targeting the predicted CEBPA-binding site (sequences in ​Table \u003cspan refid=\"MOESM2\" class=\"InternalRef\"\u003eS2\u003c/span\u003e). Binding levels were calculated as % input. IgG controls confirmed specificity.\u003c/p\u003e\n\u003ch3\u003eDual-Luciferase Reporter Assay\u003c/h3\u003e\n\u003cp\u003eThe interaction between CEBPA and the PTK2B promoter was assessed using the dual-luciferase system (Promega, Cat. No. E1910). The PTK2B promoter fragment was cloned into pGL3-basic (Firefly luciferase), with pRL-TK (Renilla luciferase) as an internal control. HEK293T cells were co-transfected with pGL3-PTK2B-Luc, pRL-TK, and either pcDNA3.1-CEBPA (overexpression) or siCEBPA (knockdown). Firefly luciferase activity (normalized to Renilla) was measured 48 h post-transfection using a luminometer. Relative luciferase activity (%) reflected CEBPA-dependent PTK2B promoter regulation. Triplicate wells and \u0026ge;\u0026thinsp;3 biological replicates ensured reproducibility.\u003c/p\u003e \u003cp\u003e \u003cb\u003eIn Vivo\u003c/b\u003e \u003cb\u003eXenograft Model\u003c/b\u003e\u003c/p\u003e \u003cp\u003eFemale nude mice (4\u0026ndash;6 weeks old) were purchased from Jiangsu GemPharmatech Co., Ltd. (Nanjing, China). All animal procedures were approved by the Institutional Animal Care and Use Committee (IACUC) of Stamp of The Animal Ethical and Welfare Committee of Tianjin Medical University Cancer Institute and Hospital (Approval No. [NSFC-AE-2022061]) and conducted in accordance with the Guide for the Care and Use of Laboratory Animals. For tumor xenografts, 5\u0026times;10⁶ cells (resuspended in 100 \u0026micro;L PBS) were subcutaneously injected into the right flanks of mice (n\u0026thinsp;=\u0026thinsp;6 per group). Experimental groups included: ​NC​, ​SFRP2, ​shPTK2B​, and ​SFRP2\u0026thinsp;+\u0026thinsp;shPTK2B​. Tumor size was measured every 3 days using calipers, and tumor volume (mm\u0026sup3;) was calculated as: Volume\u0026thinsp;=\u0026thinsp;0.5 \u0026times; length \u0026times; width\u0026sup2;. After 33 days, mice were euthanized by CO₂ asphyxiation followed by cervical dislocation, and tumors were excised, weighed, and photographed. Tumor tissues were either fixed in 4% paraformaldehyde for histological analysis or snap-frozen in liquid nitrogen for further molecular studies.\u003c/p\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eStatistical Analysis\u003c/h2\u003e \u003cp\u003eData are presented as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD. Differences between groups were analyzed using Student's t-test (two groups) or one-way ANOVA (multiple groups). Correlation analysis was performed using Pearson's chi-square test. Survival curves were plotted using the Kaplan-Meier method and compared by the log-rank test. P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 was considered statistically significant.\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003eSFRP2 is Overexpressed in Ovarian Cancer and Correlates with Poor Prognosis\u003c/h2\u003e \u003cp\u003eTo define the clinical relevance of SFRP2 in ovarian cancer, we first assessed its expression and prognostic value. Bioinformatic analysis of the GEO dataset (GSE66957) revealed that SFRP2 mRNA levels were significantly elevated in ovarian cancer tissues compared to normal ovarian tissues (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA). Consistent with this, analysis of the TCGA-OV cohort demonstrated that high SFRP2 expression was associated with significantly shorter overall survival (OS) (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eWe validated these findings at the protein level using immunohistochemistry (IHC) on a human ovarian cancer tissue microarray, which confirmed marked upregulation of SFRP2 in cancer tissues (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC). Furthermore, correlation analysis between SFRP2 expression and clinicopathological parameters revealed that high SFRP2 expression was positively correlated with higher T stage, lymph node metastasis (N stage), distant metastasis (M stage), larger tumor size, and advanced pathological stage (FIGO stage) (Tables\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e and \u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Critically, Kaplan-Meier survival analysis of our patient cohort confirmed that high SFRP2 protein expression predicted a poorer prognosis (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD). These data firmly establish SFRP2 as a clinically relevant oncoprotein in ovarian cancer.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eRelationship between SFRP2 expression and tumor characteristics in patients with ovarian cancer\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eFeatures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eNo. of patients\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c4\" namest=\"c3\"\u003e \u003cp\u003eSFRP2 expression\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003ep value\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003elow\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ehigh\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAll patients\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e137\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e66\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e71\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAge\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.438\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026le;\u0026thinsp;51years\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e70\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e36\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e34\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;51years\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e67\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e37\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStage\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eI\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e12\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eII\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e34\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIII\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e66\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e26\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e40\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIV\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTumor size\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.021\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026le;\u0026thinsp;11.9cm\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e69\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e40\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e29\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026gt;\u0026thinsp;11.9cm\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e68\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e26\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e42\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT Infiltrate\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e12\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e34\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e91\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e31\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e60\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eLymphatic metastasis(N)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.003\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eN0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e105\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e58\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e47\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eN1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e32\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e24\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMetastasis\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.002\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e112\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e61\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e51\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eM1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSpearman correlation analysis between SFRP2 expression and tumor characteristics in patients with ovarian cancer\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSFRP2\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT Infiltrate\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSpearman correlation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.400\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSignificance (two-tailed)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eN\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e137\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eLymphatic metastasis(N)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSpearman correlation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.256\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSignificance (two-tailed)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.003\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eN\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e137\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMetastasis\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSpearman correlation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.266\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSignificance (two-tailed)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.002\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eN\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e137\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStage\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSpearman correlation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.421\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSignificance (two-tailed)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eN\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e137\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTumor size\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSpearman correlation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.197\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSignificance (two-tailed)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.021\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eN\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e137\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003eKnockdown of SFRP2 Suppresses Malignant Phenotypes In Vitro\u003c/h2\u003e \u003cp\u003eTo further validate the oncogenic role of SFRP2 in ovarian cancer, we performed a series of \u003cem\u003ein vitro\u003c/em\u003e functional assays. qPCR analysis confirmed that SFRP2 mRNA was highly expressed in ovarian cancer cell lines such as HEY, SK-OV-3 compared to the normal ovarian epithelial cell line IOSE80 (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA). Based on these expression profiles, we selected HEY and SK-OV-3 cells for subsequent functional studies. To establish stable SFRP2-knockdown models, we designed three specific shRNAs targeting SFRP2. Transfection of these shRNAs into HEY and SK-OV-3 cells, followed by qPCR and western blot analyses, identified shSFRP2-#1 and shSFRP2-#3 as the most efficient sequences for SFRP2 knockdown (Figure \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003eA-C).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eFunctional assays demonstrated that SFRP2 knockdown significantly inhibited cell proliferation, as evidenced by CCK-8 assays (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB). Flow cytometric analysis showed a significant increase in the apoptosis rate in SFRP2-knockdown cells compared with the negative control (shCtrl) (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC). Furthermore, the clonogenic survival of SFRP2-knockdown cells was drastically impaired, as indicated by fewer and smaller colonies formed in the colony formation assay (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD). Cell migration, assessed by both wound healing and Transwell assays, was also markedly suppressed in SFRP2-knockdown cells compared to controls (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eE, \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eF). These results demonstrate that SFRP2 is essential for maintaining the proliferation, survival, and migratory capacity of ovarian cancer cells.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003eSFRP2 Drives Tumor Progression by Enhancing Aerobic Glycolysis\u003c/h2\u003e \u003cp\u003eTo elucidate the mechanism by which SFRP2 drives ovarian cancer progression, we performed GSEA on TCGA-OV transcriptomic data. Intriguingly, the C-C motif chemokine receptor 5 (CCR5) signaling pathway was the most significantly enriched gene set in the SFRP2 high-expression group (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA). Given that CCR5 has been previously linked to tumor glycolysis[\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e], we hypothesized that SFRP2 might contribute to ovarian cancer progression by modulating aerobic glycolysis. To test this hypothesis, we first examined the effect of SFRP2 on key glycolytic indicators. Western blot analysis showed that knockdown of SFRP2 downregulated the expression of critical glycolytic enzymes, including PFKFB4 and LDHA (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB). Functionally, SFRP2 knockdown led to a significant reduction in glucose consumption, lactate production, and intracellular ATP levels (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eC), all of which are hallmarks of glycolytic activity. Seahorse XF extracellular flux analysis further confirmed that SFRP2 depletion significantly decreased the ECAR (a measure of glycolytic flux) and increased the OCR (a measure of mitochondrial respiration) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eD), indicating a metabolic shift away from glycolysis.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTo ascertain whether glycolysis is essential for SFRP2-mediated oncogenic effects, we overexpressed SFRP2 and treated cells with the glycolytic inhibitor 2-Deoxy-D-glucose (2-DG). SFRP2 overexpression potently enhanced the expression of PKM2, HK2, GLUT1, ALDOC, ALDOA, ADH6 and LDHA (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eE), and increased glucose uptake, lactate and ATP production, (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eF). These metabolic changes were accompanied by a promotion of cell proliferation and colony formation, key indicators of tumorigenic potential (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eG, H). Importantly, 2-DG treatment effectively abolished these pro-glycolytic and pro-tumorigenic effects induced by SFRP2 overexpression (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eE-H). These data conclusively demonstrate that SFRP2 facilitates ovarian cancer progression by augmenting aerobic glycolysis.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003eSFRP2 Promotes CEBPA-Mediated Transcriptional Upregulation of PTK2B\u003c/h2\u003e \u003cp\u003eTo identify the downstream effector mediating the oncogenic function of SFRP2, we focused on the CCR5 pathway-related genes. qPCR analysis revealed that knockdown of SFRP2 significantly reduced mRNA levels of PTK2B, but not other candidates, identifying PTK2B as a key potential downstream target (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA). We next investigated how SFRP2 regulates PTK2B. As SFRP2 is a secreted protein, we hypothesized an indirect transcriptional mechanism. Using the STRING database to find SFRP2-interacting proteins and the hTFtarget database to predict transcription factors for the PTK2B promoter, we identified CEBPA as a common candidate (data not shown). This hypothesis was supported by Co-IP experiments, which confirmed a direct physical interaction between endogenous SFRP2 and CEBPA proteins in ovarian cancer cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eB). Notably, SFRP2 knockdown markedly reduced the nuclear accumulation of CEBPA, as shown by nuclear-cytoplasmic fractionation followed by WB (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eC), suggesting SFRP2 facilitates CEBPA's nuclear translocation.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eWe then examined whether the SFRP2/CEBPA complex regulates PTK2B transcription. ChIP assays demonstrated that SFRP2 overexpression significantly enhanced the enrichment of CEBPA on the PTK2B promoter region (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eD). Consistent with this, dual-luciferase reporter assays showed that co-expression of SFRP2 and CEBPA synergistically activated the activity of a luciferase reporter driven by the PTK2B promoter (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eE). Taken together, these results delineate a novel mechanism whereby SFRP2 binds to CEBPA, promotes its nuclear translocation, and enhances its transcriptional activation of PTK2B.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003eThe SFRP2/PTK2B Axis Promotes Malignant Progression through Aerobic Glycolysis\u003c/h2\u003e \u003cp\u003eFinally, we conducted rescue experiments to confirm that PTK2B is the critical functional mediator of SFRP2. We established SFRP2-overexpressing HEY and SK-OV-3 cells, in which PTK2B was simultaneously knocked down (Figure \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003eD-E). SFRP2 overexpression significantly promoted cell proliferation and colony formation, phenotypes that were effectively reversed upon PTK2B knockdown (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eA, B). These results indicate that PTK2B is indispensable for SFRP2-driven proliferative and clonogenic capacities.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eWe next extended these findings to an \u003cem\u003ein vivo\u003c/em\u003e xenograft mouse model to assess the functional relevance of the SFRP2-PTK2B axis in tumor growth. Tumors derived from SFRP2-overexpressing cells exhibited significantly accelerated growth kinetics, resulting in markedly larger and heavier tumor masses compared to those derived from control cells. Strikingly, co-knockdown of PTK2B in SFRP2-overexpressing cells largely abolished these pro-tumorigenic effects, leading to markedly impaired tumor growth (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eC-E). Western blot analysis of tumor lysates showed that SFRP2 overexpression upregulated the glycolytic enzyme PKM2, and this effect was again abolished by PTK2B knockdown (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eF). Furthermore, IHC analysis of tumor sections revealed that SFRP2 overexpression increased the expression of PTK2B and the proliferation marker Ki-67, which was attenuated by PTK2B knockdown (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eE). These \u003cem\u003ein vivo\u003c/em\u003e data robustly validate that SFRP2 exerts its pro-tumorigenic and pro-glycolytic effects primarily through PTK2B, underscoring the critical role of the SFRP2-PTK2B axis in deriving ovarian cancer progression and metabolic reprogramming.\u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eThis study identifies a new oncogenic pathway\u0026mdash;SFRP2/CEBPA/PTK2B\u0026mdash;that drives malignant progression by activating aerobic glycolysis. We provide evidence that SFRP2, beyond its canonical roles, exerts an intracellular function by directly binding CEBPA, facilitating its nuclear translocation and co-activating PTK2B expression. The subsequent upregulation of PTK2B is indispensable for glycolytic flux and aggressive tumor phenotypes driven by SFRP2, as validated \u003cem\u003ein vitro\u003c/em\u003e and \u003cem\u003ein vivo\u003c/em\u003e.\u003c/p\u003e \u003cp\u003eSFRP2 is a key regulator of the Wnt signaling pathway and plays complex roles in various cancers. In colorectal cancer, SFRP2 activates enolase 2 (ENO2) through the TCF4/β-catenin axis, significantly promoting glycolysis and metastatic phenotypes. Inhibition of ENO2 reverses SFRP2-induced metastasis, and co-expression of SFRP2/ENO2 is associated with poorer survival and higher recurrence rates[\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. In non-small cell lung cancer, m6A-methylated SFRP2 is recognized and interacts with YTHDF2; knockdown of SFRP2 suppresses glycolysis and cancer stem cell properties[\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. Current research on SFRP2 in ovarian cancer still has several gaps, such as its interaction with known glycolytic regulators (e.g., HIF-1α, FOXK2\u003csup\u003e[21\u0026ndash;22]\u003c/sup\u003e) and its subtype-specific roles in different ovarian cancer subtypes[\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Existing evidence suggests that targeting SFRP2 and its downstream glycolytic pathways may offer new strategies for improving ovarian cancer prognosis[\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. Our findings robustly position SFRP2 as an oncogene in ovarian cancer, consistently demonstrating that its high expression correlates with advanced disease and poor survival. We extend the understanding of SFRP2's oncogenic functions by establishing its critical role in promoting proliferation, clonogenicity, migration, and anti-apoptosis in ovarian cancer cells. More importantly, we uncover a previously unrecognized function of SFRP2 in regulating cancer cell metabolism. While a recent study suggested SFRP2's involvement in metabolism in other contexts[\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e], its direct role in driving aerobic glycolysis, the Warburg effect, in ovarian cancer is a novel finding. Our data demonstrate that SFRP2 significantly enhances glycolytic flux, and this metabolic reprogramming is essential for its tumor-promoting effects, as inhibition of glycolysis abrogated SFRP2-driven oncogenic phenotypes.\u003c/p\u003e \u003cp\u003eOur GSEA hinting at the CCR5 pathway was crucial, as it provided a rationale to connect SFRP2 to glycolysis, leveraging existing literature on CCR5's metabolic roles [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. This led us to investigate downstream effectors. Among CCR5 pathway-related candidates, PTK2B emerged as a key target transcriptionally regulated by SFRP2. PTK2B is a non-receptor tyrosine kinase known for its roles in cell adhesion, migration, and survival in various cancers[\u003cspan additionalcitationids=\"CR27\" citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. Emerging evidence has begun to link PTK2B to metabolic regulation. For instance, the interaction between PTK2B and SIRPα in macrophages regulates both autophagy and necroptosis, processes closely linked to cellular metabolic reprogramming such as glycolysis and mitochondrial function, further supporting the role of PTK2B in immune-metabolic crosstalk[\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e]. However, its role in ovarian cancer metabolism, particularly in glycolysis, remained unexplored. Our study clearly demonstrates that PTK2B is a critical mediator of SFRP2-induced aerobic glycolysis and tumor progression in ovarian cancer. Knockdown of PTK2B reversed the pro-glycolytic and pro-tumorigenic effects of SFRP2 both \u003cem\u003ein vitro\u003c/em\u003e and \u003cem\u003ein vivo\u003c/em\u003e. This aligns with and extends the emerging concept of PTK2B as a metabolic regulator in cancer, providing a direct link between SFRP2 signaling and PTK2B-dependent glycolytic reprogramming.\u003c/p\u003e \u003cp\u003eThe transcriptional mechanism delineated here is particularly intriguing. SFRP2, typically considered a secreted Wnt antagonist, is shown to have an intracellular role by directly interacting with CEBPA and influencing its subcellular localization and transcriptional activity. This non-canonical, intracellular function expands the paradigm of SFRP2's actions in cancer and suggests potential novel therapeutic strategies aimed at disrupting this protein-protein interaction. CEBPA is a transcription factor involved in differentiation and metabolism[\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]. Our ChIP and luciferase reporter assays confirm that the SFRP2/CEBPA complex directly binds to and activates the PTK2B promoter, establishing a linear pathway from SFRP2 to PTK2B upregulation.\u003c/p\u003e \u003cp\u003eIn summary, our work delineates a previously unrecognized SFRP2/CEBPA/PTK2B axis that is critical for aerobic glycolysis and malignant progression in ovarian cancer. These findings not only deepen our understanding of the molecular mechanisms driving metabolic reprogramming in this deadly disease but also highlight SFRP2 and PTK2B as promising prognostic biomarkers and attractive therapeutic targets. Targeting this axis, perhaps with monoclonal antibodies against SFRP2 or small molecule inhibitors of PTK2B, could represent a novel metabolic therapy for ovarian cancer patients.\u003c/p\u003e"},{"header":"Declarations","content":" \u003cp\u003e \u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e \u003cp\u003e We hereby confirm that all experimental procedures involving mice in this study were reviewed and approved by the Institutional Animal Care and Use Committee (IACUC) of Tianjin Medical University Cancer Institute and Hospital (Approval No. NSFC-AE-2022061). All procedures were strictly conducted in accordance with the approved protocol and internationally recognized guidelines for the care and use of laboratory animals.\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eConsent for publication\u003c/strong\u003e \u003cp\u003eNot applicable. This study involved experimental animals only and did not include any human participants or individual person\u0026rsquo;s data.\u003c/p\u003e \u003ch2\u003eCompeting interests\u003c/h2\u003e \u003cp\u003eThe authors declare no conflict of interest.\u003c/p\u003e \u003ch2\u003eFunding\u003c/h2\u003e \u003cp\u003eThis work was supported by the National Natural Science Foundation of China (No. 82202863, Jindong Sheng; No. 82272946, Ke Wang).\u003c/p\u003e\u003ch2\u003eAuthor contribution\u003c/h2\u003e \u003cp\u003eJ. S., K. W. and W. L. designed this program. J. S., Y. X., J. L., X. L., H. S., L. S., M. Y. operated the cell and animal experiments. J. S., Y. X., X. F., H. W. and Y. C. conducted the data collection and analysis. J. S. and Y. X. produced the manuscript which was checked by J. S., K. W. and W. L. All the authors have confirmed the submission of this manuscript.\u003c/p\u003e\u003ch2\u003eAcknowledgements\u003c/h2\u003e \u003cp\u003eThe authors gratefully acknowledge the support from the National Natural Science Foundation of China.\u003c/p\u003e\u003ch2\u003eAvailability of data and materials\u003c/h2\u003e \u003cp\u003eThe data that support the findings of this study are available from the corresponding author upon reasonable request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eCaruso G, Weroha SJ, Cliby W. Ovarian Cancer: A Review. JAMA. 2025;334:1278\u0026ndash;91. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1001/jama.2025.9495\u003c/span\u003e\u003cspan address=\"10.1001/jama.2025.9495\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWebb PM, Jordan SJ. 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Molecular stop signs: regulation of cell-cycle arrest by C/EBP transcription factors. J Cell Sci. 2005;118:2545\u0026ndash;55. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1242/jcs.02459\u003c/span\u003e\u003cspan address=\"10.1242/jcs.02459\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"journal-of-translational-medicine","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jtrm","sideBox":"Learn more about [Journal of Translational Medicine](http://translational-medicine.biomedcentral.com)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/jtrm/default.aspx","title":"Journal of Translational Medicine","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Ovarian Cancer, SFRP2, PTK2B, Aerobic Glycolysis, Transcriptional Regulation","lastPublishedDoi":"10.21203/rs.3.rs-8553543/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8553543/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003eOvarian cancer is a highly lethal gynecologic malignancy, with metabolic reprogramming being a key contributor to its progression and therapy resistance. Although Secreted Frizzled-Related Protein 2 (SFRP2) is implicated in cancer, its functional role and molecular mechanisms in ovarian cancer, particularly in regulating metabolic pathways, remain poorly defined.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eBioinformatics analysis of GEO (GSE66957) and TCGA-OV datasets was performed to assess SFRP2 expression and its correlation with prognosis. Immunohistochemistry (IHC) on a human ovarian cancer tissue microarray was used for clinical validation. SFRP2 was knocked down or overexpressed in ovarian cancer cell lines (HEY, SK-OV-3) using lentiviral shRNAs. Functional assays (CCK-8, colony formation, apoptosis, migration) and metabolic assays (glucose consumption, lactate/ATP production, ECAR/OCR) were conducted. The mechanistic link between SFRP2, transcription factor CEBPA, and downstream target PTK2B was investigated using co-immunoprecipitation (Co-IP), nuclear-cytoplasmic fractionation, chromatin immunoprecipitation (ChIP), and dual-luciferase reporter assays. Rescue experiments were performed both in vitro and in vivo (xenograft mouse models).\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eSFRP2 was significantly overexpressed in ovarian cancer tissues and cell lines, and high SFRP2 expression correlated with advanced disease and poor patient survival. SFRP2 knockdown suppressed cell proliferation, colony formation, and migration, while promoting apoptosis. Gene set enrichment analysis linked SFRP2 to the CCR5 signaling pathway, prompting an investigation into glycolysis. SFRP2 depletion impaired aerobic glycolysis, reducing glucose uptake, lactate/ATP production, and ECAR, while increased OCR. Conversely, SFRP2 overexpression enhanced glycolytic flux and tumorigenic phenotypes, which were abrogated by the glycolytic inhibitor 2-DG. Mechanistically, SFRP2 interacted with the transcription factor CEBPA, promoted its nuclear translocation, and enhanced its binding to the PTK2B promoter, leading to PTK2B transcriptional activation. Crucially, PTK2B knockdown reversed the pro-glycolytic and pro-tumorigenic effects of SFRP2 overexpression both in vitro and in vivo.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eOur findings identify a novel SFRP2/CEBPA/PTK2B signaling axis that drives aerobic glycolysis and malignant progression in ovarian cancer, highlighting SFRP2 and PTK2B as potential prognostic markers and therapeutic targets.\u003c/p\u003e","manuscriptTitle":"SFRP2 Drives Aerobic Glycolysis and Tumor Progression in Ovarian Cancer via Transcriptional Upregulation of PTK2B","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-01-16 11:02:46","doi":"10.21203/rs.3.rs-8553543/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewerAgreed","content":"","date":"2026-01-13T14:40:46+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-01-13T07:32:51+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-01-10T12:41:23+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Translational Medicine","date":"2026-01-09T09:15:31+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-translational-medicine","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jtrm","sideBox":"Learn more about [Journal of Translational Medicine](http://translational-medicine.biomedcentral.com)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/jtrm/default.aspx","title":"Journal of Translational Medicine","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ab4eb1cd-0aba-4ede-8a90-6b3b575e4346","owner":[],"postedDate":"January 16th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-01-16T11:02:46+00:00","versionOfRecord":[],"versionCreatedAt":"2026-01-16 11:02:46","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8553543","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8553543","identity":"rs-8553543","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}