WFDC3: A Novel Prognostic Biomarker in Pancreatic Cancer

Introduction:

Pancreatic cancer remains one of the most lethal malignancies, with a five-year survival rate of just 10%. The difficulty in early diagnosis, coupled with its aggressive nature and resistance to conventional treatments, makes it a formidable challenge for clinicians. While immunotherapies have shown promise in treating other cancers, they have limited efficacy in pancreatic ductal adenocarcinoma (PAAD) due to its immunosuppressive tumor microenvironment. As a result, identifying reliable biomarkers for prognosis and potential therapeutic targets remains a critical priority for advancing treatment in this devastating disease.

WFDC3 and Its Role in Cancer

The Whey-Acidic-Protein Four-Disulfide Core (WFDC) family comprises 18 small, secreted proteins, each characterized by a conserved "WFDC" domain stabilized by four disulfide bonds. These proteins are involved in various biological processes, including protease regulation, host defense, and immune responses. Abnormal expression of WFDC proteins has been linked to cancer progression, particularly in ovarian and lung cancers. However, the role of WFDC3 in PAAD had not been fully explored until now.

Study Overview

In a groundbreaking study, Liu et al. investigated the role of WFDC3 in PAAD through a comprehensive multi-omic analysis. The study integrated several cutting-edge techniques to explore WFDC3 expression, its effects on tumor biology, and its potential as a prognostic biomarker. This included:

• Pan-cancer bioinformatics: Using data from The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx) databases to analyze WFDC3 expression and survival associations.
• Single-cell RNA sequencing: Examining the localization of WFDC3 in different cell types within PAAD tissues.
• Functional assays: Testing the effects of WFDC3 on PAAD cell migration, invasion, and immune interactions in vitro.
• Prognostic modeling: Constructing a WFDC3-based risk score using LASSO regression to assess its predictive power for patient survival.

Key Findings

1. WFDC3 Overexpression Is Linked to Poor Prognosis in PAAD
   • Tumor vs. normal: Analysis of TCGA-GTEx data revealed that WFDC3 mRNA expression is significantly higher in PAAD tissues compared to normal pancreas tissue.
     
   • Survival impact: High WFDC3 expression correlated with worse overall survival (OS), disease-specific survival (DSS), and progression-free survival (PFS). Notably, patients with high WFDC3 had an OS hazard ratio (HR) of 1.89 (P = 0.003), underscoring its potential as a prognostic biomarker.
     
2. Malignant Epithelial Specificity Revealed by Single-Cell Analysis
   • Cell type localization: Single-cell RNA-seq analysis (GSE154778) showed that WFDC3 is predominantly expressed in epithelial cell clusters, with minimal expression in stromal or immune cells.
     
   • Tumor-specific expression: InferCNV analysis indicated that malignant ductal cells, exhibiting high chromosomal instability, express significantly higher levels of WFDC3 compared to normal epithelium (P < 1e-12).
     
3. WFDC3 Drives Tumor Migration, Invasion, and Epithelial-Mesenchymal Transition (EMT)
   • Functional assays: Overexpression of WFDC3 in PANC-1 cells led to increased cell migration and invasion. Conversely, knockdown of WFDC3 in MIA PaCa-2 cells resulted in decreased migration and invasion (Transwell assays; Figures 7A–B).
     
   • EMT regulation: Upregulation of WFDC3 was associated with increased mesenchymal markers (e.g., N-cadherin, Vimentin, Snail) and decreased epithelial markers (e.g., E-cadherin, ZO-1), while WFDC3 knockdown reversed these changes (Figures 7C–F).
     
4. WFDC3 Suppresses T-Cell Cytotoxicity
   • Immune profiling: High WFDC3 expression in tumors correlated with reduced numbers of cytotoxic T cells (CD8⁺) and increased levels of immunosuppressive cells such as M0 macrophages and regulatory T cells (Tregs) (CIBERSORT analysis; Figures 6A–C).
     
   • Functional assays: Recombinant WFDC3 or WFDC3-rich conditioned media reduced Granzyme B expression in Jurkat T cells (flow cytometry; P < 0.01). In co-culture experiments, T cells interacted with WFDC3-overexpressing tumor cells, leading to decreased immune responses, including reduced expression of IFNG, IL2, Perforin, Granzyme B, and Granulysin (qRT-PCR; P < 0.05).
     
   • Tumor-killing potential: Knockdown of WFDC3 enhanced T cell-mediated tumor killing in CCK-8 and LDH assays.
     
5. A Four-Gene Prognostic Signature Based on WFDC3
   • Risk modeling: Using LASSO regression, the study identified a four-gene signature (WFDC3, WFIKKN1, PI3, and SLPI) that was prognostic in PAAD.
     
   • Risk score: The formula for the WFDC3-based risk score is:

     Risk Score = 0.068 × WFDC3—0.540 × WFIKKN1 + 0.041 × PI3 + 0.074 × SLPI
     

   • Performance: High-risk patients in the model showed significantly worse survival outcomes, with a 5-year AUC of 0.84, and the model was validated in two independent GEO cohorts (GSE62452 & GSE78229).
     

Practical Implications and Next Steps

• Prognostic tool: The WFDC3-based signature could serve as a valuable assay for stratifying PAAD patients by risk, potentially guiding personalized treatment plans and improving patient monitoring.
• Therapeutic targeting: As a secreted protein, WFDC3 presents a promising therapeutic target. Neutralizing antibodies or small molecules designed to inhibit WFDC3 may not only reduce metastasis but also relieve immunosuppressive effects in the tumor microenvironment.
• Mechanistic studies: Future research should focus on identifying WFDC3's binding partners and downstream signaling pathways (e.g., STAT, NF-κB), as well as testing the therapeutic potential of WFDC3-targeted approaches in vivo.
• Clinical translation: Measuring circulating WFDC3 levels could offer a non-invasive means of early detection and could be used to assess patients' responses to immunotherapies.

Conclusion

This study identifies WFDC3 as a critical driver of PAAD progression, demonstrating its role in promoting epithelial-mesenchymal transition and suppressing T cell-mediated immunity. Additionally, it highlights WFDC3's potential as a novel prognostic biomarker and a target for therapeutic interventions. With further validation and clinical testing, WFDC3 may become a valuable tool in the fight against this aggressive and hard-to-treat cancer.

More information:

Bohan Liu et al, WFDC3 identified as a prognostic and immune biomarker in pancreatic cancer. Biomol Biomed [Internet]. 2025 May 8 [cited 2025 Nov. 14];25(11):2447–2465.

Available from: https://doi.org/10.17305/bb.2025.12444

Journal information: Biomolecules and Biomedicine

Provided by: Association of Basic Medical Sciences of FBIH

Provided by Association of Basic Medical Sciences of FBIH