Platelet (hem)ITAM Signaling Mediated Cancer-Associated Venous Thrombosis

Background-Patients with cancer are at a high risk of venous thromboembolism (VTE) and underlying mechanisms are unclear. Recent studies have shown that podoplanin (PDPN) expressing highly in primary brain tumors is associated with increasingly risk of VTE by activate platelets. The C-type lectin-like receptor 2(CLEC-2) is a type II transmembrane protein expressed on the surface of platelets and PDPN is the only known endogenous ligand for it. CLEC-2 mediates platelet activation through a hem-immunoreceptor tyrosine-based activation motif ((hem)ITAM).

Objective: We tested the hypothesis that the platelet (hem)ITAM signaling is involved in the development of VTE in patients with malignant tumors, and targeted inhibitors may reduce thrombosis and improve prognosis in patients with cancer.

Materials and Methods-We selected two human malignant cell lines (NCI-H226 and C8161) expressing high levels of PDPN as models to study the role of the platelet (hem)ITAM signaling in cancer-associated venous thrombosis in vitro and in vivo.

Results and Conclusions-Our results showed that both types of cell lines expressing PDPN triggered platelet activation via CLEC-2 in vitro, which could be abrogated by an anti-PDPN antibody SZ-168 or syk inhibitor R406. Further, in vivo study showed that injection of NCI-H226 or C8161 in two mouse models of venous thrombosis activated platelets, increased platelet counts and enhanced thrombosis. These results suggested that platelet activation induced by PDPN was correlated with hypercoagulability, contributed to thrombosis in cancer patients. Importantly, PDPN enhanced thrombosis was reduced in mice treated with SZ168 or R406. Immunohistochemical staining using anti-PDPN antibody was also performed in pulmonary squamous cell carcinoma specimens of 166 patients. During up to 4-year-follow-up, 20 (12.05%) patients developed VTE. 105 tumor specimens stained positive for PDPN (27 high expression, 43 medium expression, 35 low expression). High PDPN expression was associated with an increased risk of VTE and poor prognosis, independently of age, sex and tumor type. In this study, we found that PDPN expression in tumors induced platelet aggregation and was associated with a high risk of VTE via platelet (hem)ITAM signaling. Our data also suggested that SZ168 inhibited PDPN-induced platelet aggregation in vitro and decreased the incidence of VTE in mice. In conclusion, our research demonstrated novel insights into the pathogenesis of cancer-associated venous thrombosis.