Inhibition of angiogenesis by leflunomide via targeting the soluble ephrin-A1/EphA2 system in bladder cancer

Abstract

Angiogenesis plays an important role in bladder cancer (BCa). The immunosuppressive drug leflunomide has attracted worldwide attention. However, the effects of leflunomide on angiogenesis in cancer remain unclear. Here, we report the increased expression of soluble ephrin-A1 (sEphrin-A1) in supernatants of BCa cell lines (RT4, T24, and TCCSUP) co-cultured with human umbilical vein endothelial cells (HUVECs) compared with that in immortalized uroepithelial cells (SV-HUC-1) co-cultured with HUVECs. sEphrin-A1 is released from BCa cells as a monomeric protein that is a functional form of the ligand. The co-culture supernatants containing sEphrin-A1 caused the internalization and down-regulation of EphA2 on endothelial cells and dramatic functional activation of HUVECs. This sEphrin-A1/EphA2 system is mainly functional in regulating angiogenesis in BCa tissue. We showed that leflunomide (LEF) inhibited angiogenesis in a N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-induced bladder carcinogenesis model and a tumor xenograft model, as well as in BCa cell and HUVEC co-culture systems, via significant inhibition of the sEphrin-A1/EphA2 system. Ephrin-A1 overexpression could partially reverse LEF-induced suppression of angiogenesis and subsequent tumor growth inhibition. Thus, LEF has a significant anti-angiogenesis effect on BCa cells and BCa tissue via its inhibition of the functional angiogenic sEphrin-A1/EphA2 system and may have potential for treating BCa beyond immunosuppressive therapy.

<p>Leflunomide Inhibits Proliferation and Induces Apoptosis via Suppressing Autophagy and PI3K/Akt Signaling Pathway in Human Bladder Cancer Cells</p>

“…9 A recent study showed the anti-angiogenesis effect of leflunomide on bladder cancer. 21 In the present study, we demonstrated that leflunomide reduced bladder cancer cell viability via inducing apoptosis and cell cycle arrest. Additionally, autophagy and Akt/mTOR signaling pathway were involved in the cytotoxicity of leflunomide on bladder cancer cells.…”

“…Recently, leflunomide was reported to inhibit tumor angiogenesis by suppressing ephrin-A1/EphA2 in bladder cancer. 21 However, other pathways affected by leflunomide such as apoptosis and autophagy mechanism in bladder cancer have not been well elucidated. In this study, we reported that leflunomide suppressed bladder cancer cells growth via inducing tumor cell apoptosis and cell cycle arrest in S phase.…”

C1GALT1 is associated with poor survival and promotes soluble Ephrin A1-mediated cell migration through activation of EPHA2 in gastric cancer

“…Ephrin A1 is the ligand for EPHA receptors and is overexpressed in gastric cancer. In addition, soluble Ephrin A1 has been reported to be secreted in conditioned media of several cancer cell lines and be detected in serum of cancer patients [38][39][40]. We found that soluble Ephrin A1 was present in conditioned media of AGS and MKN cells Nine patients presented with metastatic disease.…”

“…Soluble Ephrin A1 can promote metastasis in several cancer types [38,39,42]. We found that C1GALT1 modified O-glycans on EPHA2 and regulated EPHA2 phosphorylation.…”

Roles of EphA1/A2 and ephrin‐A1 in cancer

“…Therefore, roles of EphB4 and ephrin‐B2 in tumor growth and agiogenesis are still controversial. In pathological conditions, EphA1/A2 and ephrin‐A1 have been implicated in tumor angiogenesis that is important for tumor growth to supply O 2 and nutrients . However, the detailed molecular mechanisms remain to be elucidated.…”