Study on the Role of Coagulation Factor X in Anti-androgen Resistance in Prostate Cancer

Research Background

Prostate cancer (PCa) is the second most common cancer in men worldwide and is one of the leading causes of cancer-related deaths. The occurrence and progression of prostate cancer are highly dependent on the androgen receptor (AR) signaling pathway. Consequently, various AR signaling inhibitors (ARSI) have been developed in recent years for treating prostate cancer. However, despite the initial efficacy of these treatments, most patients eventually develop into castration-resistant prostate cancer (CRPC), a stage where the tumors are resistant to traditional immunotherapies, described as “immune cold” tumors. The formation of CRPC is often accompanied by the accumulation of immune-suppressive myeloid-derived suppressor cells (MDSCs) and chronic inflammation, showing a high tendency for coagulation. However, whether coagulation factors directly contribute to tumor proliferation is not clear. This study aims to reveal the mechanism by which coagulation factor X (Factor X, FX) in the prostate cancer microenvironment affects the proliferation and resistance of CRPC cells.

Research Overview

This study, conducted by researchers Bianca Calì, Martina Troiani, Andrea Alimonti, and others from institutions such as the Institute of Oncology Research in Switzerland and Università della Svizzera Italiana, was published in the journal Cancer Cell in October 2024. Using single-cell RNA sequencing (scRNA-seq) to analyze the tumor microenvironment in mouse CRPC models, the study discovered that immunosuppressive neutrophils (PMN-MDSCs) are a major source of FX. The factor activates the protease-activated receptor 2 (PAR2) and ERK1/2 pathways in prostate cancer cells, promoting androgen-independent tumor proliferation and resistance. This research offers new potential targets for CRPC treatment strategies.

Research Methods and Procedures

1. Research Process and Sample Handling

In this study, two mouse CRPC models were used: a PTEN gene deletion mouse model (Pten^pc-/-) and the TRAMP-C1 cell line derived from the TRAMP (transgenic adenocarcinoma mouse) model. Hormone-sensitive (HS) and resistant (CR) states of tumors were created in both models. To identify tumor-infiltrating neutrophils, the team employed scRNA-seq technology, with Uniform Manifold Approximation and Projection (UMAP) for dimensionality reduction analysis, comparing the gene expression characteristics to those of immunosuppressive neutrophils (PMN-MDSCs).

2. Identification and Expression Analysis of Coagulation Factors

Through differential gene expression analysis, the researchers found that FX was significantly upregulated in immunosuppressive neutrophils (PMNs) in both mouse models and further increased in PMNs during the resistance phase. Activation of FX depends on tissue factor (TF) and FVII expressed in the tumor microenvironment, forming an activated Factor Xa (FXa) that promotes tumor cell proliferation. The study also showed that FX-expressing neutrophils maintained high expression levels after treatment with the CXCR2 inhibitor and anti-IL-23 antibody, indicating a low sensitivity of these PMNs to CXCR2 inhibitors.

3. Functional Experiments of FX In Vitro and In Vivo

In vitro experiments revealed that adding recombinant FXa to prostate cancer cells under androgen deprivation culture conditions significantly increased tumor cell proliferation rates. Further Western blot analysis showed that FXa activated the ERK1/2 signaling pathway in prostate cancer cells, and phosphorylation of ERK1/2 was blocked by specific PAR2 antagonists AZ3451 and ENMD-1068, confirming that FXa promotes cell proliferation through PAR2. Additionally, in vivo experiments involving FX gene knockout in mouse models significantly inhibited tumor growth, highlighting the crucial role of FX in CRPC.

4. Drug Experiments: Application of FX Inhibitors

The anticancer effects of the FXa inhibitor Rivaroxaban were also evaluated. In mouse CRPC models treated with androgen deprivation and Enzalutamide, Rivaroxaban significantly reduced tumor proliferation rates, inhibited the ERK signaling pathway, and enhanced the therapeutic efficacy of Enzalutamide. Further pathological analysis showed reduced Ki-67 positive cells and decreased proliferation activity in tumor tissues from the combination therapy group, demonstrating the potential clinical application of FXa inhibition in CRPC.

Research Results

1. Expression and Function of FX in CRPC

The study found that immunosuppressive neutrophils (PMNs) in the prostate cancer microenvironment are a major source of FX. FX activation does not occur via the classic coagulation pathway but rather through binding to PAR2 to mediate signaling, promoting prostate cancer cell proliferation and resistance formation. Under androgen deprivation conditions, FXa activation significantly promotes tumor cell proliferation and enhances ERK1/2 phosphorylation.

2. Key Role of PAR2

PAR2, as a G protein-coupled receptor, is involved in FXa-mediated cancer cell proliferation signaling. It was observed that PAR2 expression in prostate cancer cells is significantly higher than in normal tissues and increases with cancer progression, indicating PAR2’s critical role in CRPC. Targeted inhibition of PAR2 can effectively reduce ERK1/2 activity, thereby inhibiting tumor cell proliferation.

3. Expression of FX and CD84 in Immunosuppressive Neutrophils and Disease Prognosis

In CRPC patients, CD84+ PMNs exhibited high levels of FX expression. Further analysis showed a positive correlation between plasma FX levels and peripheral blood neutrophil counts, with high levels of FX serving as an independent prognostic factor for shorter survival in CRPC patients. Moreover, tumor tissues with high expression of CD84 and F10 (the gene encoding FX) marked increased numbers of immunosuppressive neutrophils, which is associated with poor patient prognosis.

Research Conclusion and Significance

This study reveals for the first time that immunosuppressive neutrophils (PMNs) in the CRPC tumor microenvironment are a key source of coagulation factor FX. FX promotes tumor cell proliferation and resistance in vivo by activating the PAR2 signaling pathway. The study indicates that inhibiting FXa activity can effectively block tumor proliferation and enhance the therapeutic effect of anti-androgen drugs like Enzalutamide. Additionally, it suggests that CD84 serves as a marker for FX high-expressing neutrophils and is significantly associated with poor prognosis in CRPC patients, representing new therapeutic targets.

Research Highlights and Innovation

1. Innovative Discovery: First revelation of FX’s non-coagulation function in CRPC, specifically through PAR2 activation to promote cancer cell proliferation.
2. Potential Therapeutic Target: The study shows that directly inhibiting FXa helps block CRPC progression and enhances Enzalutamide efficacy, providing new drug targets for CRPC treatment.
3. Clinical Prognostic Marker: The study confirms that CD84+ FX high-expressing immunosuppressive neutrophils are associated with poorer prognosis in CRPC patients, offering new biomarkers for clinical management.

Research Outlook

This study provides new insights into the complex interactions between coagulation factors and immune cells in CRPC and offers theoretical support for developing FXa-targeted therapies. Future research should further investigate the clinical application potential of FXa inhibitors in CRPC patients, along with conducting more clinical trials to validate the study’s findings. Exploring the role of the PAR2 signaling pathway in other cancer types will also hold significant academic value.

Summary

This study reveals the critical role of coagulation factor FX in prostate cancer resistance, expanding our understanding of the interactions between the coagulation system and immune system in cancer progression. The research offers potential new treatment routes for CRPC patients who have failed anti-androgen therapy, and lays the foundation for future immunosuppressive neutrophil-targeted therapeutic strategies.