Tumour necrosis factor receptor associated factors (TRAFs) play a key role in signal transduction in mammalian cells. Several members of the TRAF family have been identified but only TRAF2 and TRAF6 are implicated in the regulation of osteoclastogenesis. Here we studied the role of TRAF2 and TRAF6 in breast cancer induced bone cell activity and osteolysis. We observed that TRAF2, but not TRAF6, is highly expressed in the human MDA-MB-231 (MDA-231) bone-seeking breast cancer cells when compared to parental cells (TRAF2; 43% increase, P<0.01). Targeted knockdown of TRAF2, but not TRAF6, in MDA-231 cells by small interfering RNAs markedly reduced cell migration (38% reduction, P<0.05), significantly reduced the ability of MDA-231 cells (41% reduction, P<0.05) and their conditioned medium (69% reduction, P<0.05) to induce osteoclast formation in bone marrow cultures. Next, we successfully generated stable parental and bone seeking MDA-231 cell lines over-expressing TRAF2 using a retroviral approach. Over-expression of TRAF2 in the parental MDA cell lines (eightfold increase) significantly enhanced cell migration (30% increase, P<0.001) and invasion (50% increase, P<0.05). Over-expression of TRAF2 in bone-seeking MDA-231 cells significantly enhanced the stimulatory effects of these cells (187% increase, P< 0.001) and their conditioned medium (24% increase, P<0.005) on osteoclast formation in RANKL stimulated bone marrow cultures. Moreover, studies in human MDA-231mouse calvaria organ co-cultures showed that conditioned medium obtained from MDA-231 cells over-expressing TRAF2 caused profound increase in osteolysis (20% increase, P< 0.05) when compared to conditioned medium from control cells. In conclusion, our studies showed that TRAF2, but not TRAF6, activity in breast cancer cells regulates breast cancer cell motility in vitro and osteolysis ex vivo. However, the role of TRAF2 in bone metastasis and osteolytic bone loss associated with breast cancer in vivo will require further investigation.
17 May 2014 - 20 May 2014