Bisphosphonate drugs (BPs) rapidly target the skeleton and are the gold
standard treatment to inhibit bone destruction in patients with osteoporosis
and metastatic bone disease. However, BPs also have anti-cancer effects outside
bone; in mouse models they reduce tumour growth and metastasis and there is
some evidence that adjuvant BP therapy increases survival in post-menopausal
women with breast cancer. The exact mechanisms underlying these anti-cancer
effects are unknown since BPs are considered to only affect bone-resident
osteoclasts in vivo. To address this,
we determined the cell types capable of
internalising fluorescently-labelled BP in mice bearing 4T1 mammary
tumours. Within minutes of tail vein injection, intravital 2-photon imaging
revealed the diffusion of BP into tumour tissue from the leaky, disorganised
tumour vasculature. BP then appeared to bind to small, granular
microcalcifications within the tumour tissue. Intravital imaging revealed that
tumour-associated macrophages (TAMs) rapidly internalised BP by pinocytosis and
by engulfing these BP-coated microcalcifications. Flow cytometric analysis of
the tumours 24hr later confirmed that uptake occurred predominantly in TAMs and
not tumour epithelial cells.
We also identified a patient with breast cancer in which the BP 99mTc-MDP
(used for SPECT/CT bone scintigraphy) localised to the primary mammary
carcinoma. Histological analysis of the resected tumour post-surgery revealed
the presence of granular microcalcifications similar in appearance to those in
the mouse 4T1 tumours, and some of which were closely associated with CD68+
TAMs.
These studies provide clear evidence that BPs can be rapidly
internalised by macrophages outside the skeleton. The leaky vasculature of
tumours facilitates the local diffusion of BP, where it binds to
microcalcifications within the tumour that are engulfed by TAMs. Given the
important role of TAMs in promoting tumour progression and metastasis, our
studies suggest that the anti-tumour activity of BPs in cancer patients occurs indirectly
via effects on these cells.