Antithrombin controls tumor migration, invasion and angiogenesis by inhibition of enteropeptidase.
Antithrombin is a key inhibitor of the coagulation cascade, but it may also function as an anti-inflammatory, anti-angiogenic, anti-viral and anti-apoptotic protein. Here, we report a novel function of antithrombin as a modulator of tumor cell migration and invasion. Antithrombin inhibited enteropeptidase on the membrane surface of HT-29, A549 and U-87 MG cells. The inhibitory process required the activation of antithrombin by heparin, and the reactive center loop and the heparin binding domain were essential. Surprisingly, antithrombin non-covalently inhibited enteropeptidase, revealing a novel mechanism of inhibition for this serpin. Moreover, as a consequence of this inhibition, antithrombin was cleaved, resulting in a molecule with anti-angiogenic properties that reduced vessel-like formation of endothelial cells. The addition of antithrombin and heparin to U-87 MG and A549 cells reduced motility in wound healing assays, inhibited the invasion in transwell assays and the degradation of a gelatin matrix mediated by invadopodia. These processes were controlled by enteropeptidase, as demonstrated by RNA interference experiments. Carcinoma cell xenografts in nude mice showed in vivo co-localization of enteropeptidase and antithrombin. Finally, treatment with heparin reduced experimental metastasis induced by HT29 cells in vivo. In conclusion, the inhibition of enteropeptidase by antithrombin may have a double anti-tumor effect through inhibiting a protease involved in metastasis and generating an anti-angiogenic molecule.
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Ginés
G
Luengo-Gil
Instituto de Investigaciones Biomédicas Alberto Sols, CSIC-UAM, Madrid, Spain.
María Inmaculada
MI
Calvo
Instituto de Investigaciones Biomédicas Alberto Sols, CSIC-UAM, Madrid, Spain.
Ester
E
Martín-Villar
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Sonia
S
Águila
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Nataliya
N
Bohdan
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Ana I
AI
Antón
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Salvador
S
Espín
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Francisco
F
Ayala de la Peña
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Vicente
V
Vicente
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Javier
J
Corral
Instituto de Investigaciones Biomédicas Alberto Sols, CSIC-UAM, Madrid, Spain.
Miguel
M
Quintanilla
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, Murcia, Spain.
Irene
I
Martínez-Martínez
D000072283:A549 Cells; D020533:Angiogenesis Inhibitors / Q000008:administration & dosage; D000818:Animals; D000991:Antithrombins / Q000008:administration & dosage*; D002465:Cell Movement / Q000187:drug effects; D042783:Endothelial Cells / Q000187:drug effects / Q000473:pathology; D004765:Enteropeptidase / Q000235:genetics*; D019073:HT29 Cells; D006801:Humans; D051379:Mice; D009361:Neoplasm Invasiveness / Q000235:genetics / Q000473:pathology; D009369:Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D009389:Neovascularization, Pathologic / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D011485:Protein Binding; D011487:Protein Conformation
D020533:Angiogenesis Inhibitors; D000991:Antithrombins; D004765:Enteropeptidase