gms | German Medical Science

Objective: Brain metastases outnumber primary neoplasms ten-fold, and are associated with a poor prognosis. Patients with metastatic, triple negative breast cancers are at high risk (25-46%) of developing brain metastases at some point in the course of their disease. Recently we demonstrated that high ADAM8 expression in breast tumors lead to increased numbers of circulating tumor cells and a higher frequency of brain metastasis in mouse tumor models. Thus, we evaluated the mechanistic role of ADAM8, a metalloprotease-disintegrin, in facilitating trans-endothelial migration and in the formation of brain metastases.

Methods: Brain metastases were analysed using immunohistochemistry for ADAM8. To model brain metastasis of breast cancer cells, stable shRNA ADAM8 knock-down clones of the breast cancer cell line MDA-MB-231 (shA8) and control (shCtrl) cells were generated and subjected to functional assays assessing migration, invasion, and transmigration through a Blood Brain Barrier model consisting of endothelial cells and astrocytes. In addition, MMP expression was analysed in MDA-MB-231 shCtrl and shA8 cells by qPCR. Furthermore, comparative phospho-kinase arrays were utilized. Cell model data were correlated to brain metastases from primary breast tumors.

Results: A significant increase in ADAM8 expression was identified in 34% of primary site tumors, and was found to be 2-fold higher in brain metastases of different origins, including breast cancer. In transendothelial migration assays, MDA-MB-231 ADAM8 knock-down cells showed a reduced endothelial adhesion as well as a reduced transmigratory capacity both in serum-induced transmigration and in transmigration triggered by the chemokine SDF-1, a mediator of metastasis. This was further supported by the blood-brain barrier in vitro model as well as in matrigel invasion assays. ADAM8 knockdown caused reduced ERK1/2 and CREB phosphorylation and affected expression levels of MMP9 specifically. We demonstrated that in the course of transendothelial migration, the P-selectin glycoprotein ligand, a membrane protein important in leukocyte extravasation, is shed by ADAM8.

Conclusion: Our results suggest that ADAM8 is an important mediator for brain metastasis of breast cancer by affecting transendothelial migration and may offer an attractive target for therapeutic interventions.