Toll-like receptor 9 expression is associated with breast cancer sensitivity to the growth inhibitory effects of bisphosphonates in vitro and in vivo

Bisphosphonates are standard treatments for bone metastases. When given in the adjuvant setting, they reduce breast cancer mortality and recurrence in bone but only among post-menopausal patients. Optimal drug use would require biomarker-based patient selection. Such biomarkers are not yet in clinical use. Based on the similarities in inflammatory responses to bisphosphonates and Toll-like receptor (TLR) agonists, we hypothesized that TLR9 expression may affect bisphosphonate responses in cells. We compared bisphosphonate effects in breast cancer cell lines with low or high TLR9 expression. We discovered that cells with decreased TLR9 expression are significantly more sensitive to the growth-inhibitory effects of bisphosphonates in vitro and in vivo. Furthermore, cancer growth-promoting effects seen with some bisphosphonates in some control shRNA cells were not detected in TLR9 shRNA cells. These differences were not associated with inhibition of Rap1A prenylation or p38 phosphorylation, which are known markers for bisphosphonate activity. However, TLR9 shRNA cells exhibited increased sensitivity to ApppI, a metabolite that accumulates in cells after bisphosphonate treatment. We conclude that decreased TLR9-expression sensitizes breast cancer cells to the growth inhibitory effects of bisphosphonates. Our results suggest that TLR9 should be studied as a potential biomarker for adjuvant bisphosphonate sensitivity among breast cancer patients.