Forelimbs, hindlimbs, and spine of the mice were collected upon euthanasia and fixed in 10% neutral buffered formalin for 48 h and decalcified in 10% EDTA for 2 weeks. After decalcification tissues were processed in a Shandon Excelsior automated tissue processor (Thermo Fisher Scientific, Grand Island, NY) and embedded in paraffin wax for sectioning. Longitudinal, mid-sagittal sections 3.5 μm in thickness from the tibia, femur, humerus and lumbar spines were cut using an automated Microm HM 355 S microtome (Thermo Fisher Scientific). Tissue sections were stained with hematoxylin and eosin (H&E) and prepared for histomorphometric analysis. All section are viewed on Leica DM2500 compound microscope (W. Nuhsbaum Inc., McHenry, IL) with Q-imaging micropublisher cooled CCD color digital camera. Images were captured and analyzed using BioQuant software v14.1.6 (Bioquant Image Analysis Corporation). Tumor burden per mouse, defined as area of bone occupied by the cancer cells, was calculated at the tibia, femur and humerus at 50x magnification on H&E stained sections, as previously described38 (link). Osteoclast number at the tumor-bone interface (OCL/mm bone surface) in the femur, tibia and humerus was measured on TRAP stained slides at 200× magnification. The investigators were blinded to treatment of subjects.