Youwei Gong,1,2,* Binjie Chen,2,3,* Qixing Tan,2,3 Wei Wei2,3 

1Department of Blood Transfusion, Guangxi Medical University Cancer Hospital, Nanning, Guangxi, People’s Republic of China; 2Key Laboratory of Breast Cancer Diagnosis and Treatment Research of Guangxi Department of Education, Nanning, Guangxi, People’s Republic of China; 3Department of Breast Surgery, Guangxi Medical University Cancer Hospital, Nanning, Guangxi, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Wei Wei, Department of Breast Surgery, the Guangxi Medical University Cancer Hospital, 71 hedi Road, Nanning, 530021, People’s Republic of China, Email wei-wei1018@hotmail.com

Objective: Neutrophil extracellular traps (NETs) are associated with poor prognosis and an increased risk of venous thromboembolism (VTE) in metastatic breast cancer (MBC). This study aims to determine whether NETs promote hypercoagulability and if NETs and plasma hypercoagulability markers are biomarkers of survival in MBC.
Methods: Circulating levels of neutrophil extracellular trap (NET) markers and hypercoagulability markers (TAT, fibrinogen, and D-dimer) were assessed in 112 MBC patients before treatment, compared to 55 healthy controls. Stratified by NET levels and plasma TAT, fibrinogen, and D-dimer, the correlation with overall survival was analyzed. The NET procoagulant activity was evaluated using fibrin and purified coagulation complex production assays, and by measuring coagulation time (CT).
Results: MBC patients exhibited significantly elevated plasma NET levels compared to healthy controls (all P< 0.05), circulating MPO-DNA and NE-DNA levels were positively correlated with plasma TAT, fibrinogen, D-dimer, CT, FVIIIa, and platelet (PLT) counts. Additionally, we observed a significant increase in NETs formation in control neutrophils exposed to MBC plasma compared to those exposed to control plasma. NETs from MBC neutrophils significantly increased the potency of control plasma to generate thrombin and fibrin, effects that were notably attenuated by DNase I. Plasma TAT and D-dimer levels were significantly higher in MBC patients who died within three years post-recruitment compared to those who survived beyond three year. Plasma TAT and D-dimer were inversely correlated with survival. High plasma levels of MPO-DNA were associated with significantly worse overall survival (HR: 2.445, 95% CI: 1.255– 4.762, P=0.007). MBC patients with both high D-dimer and high MPO-DNA had significantly reduced survival (HR: 2.450, 95% CI: 1.332– 4.488, P=0.002).
Conclusion: Our results highlight the increased release of NETs in MBC patients and reveal that NET formation enhances hypercoagulability and cancer progression. Targeting NETs may be a potential therapeutic strategy to inhibit MBC progression and mitigate thrombotic complications in MBC.

Keywords: neutrophil extracellular traps, procoagulant activity, hypercoagulability, metastatic breast cancer, prognosis