Bone disease is commonly seen in patients suffering from multiple myeloma (MM), which in turn can severely affect the morbidity and quality of life. The proteasome inhibitor, bortezomib, a clinical potent antimyeloma agent has gained interest due to its osteo-anabolic effect. Numerous clinical and pre-clinical studies have suggested a positive impact on bone density and biomarkers associated with osteoblast and osteoclast activity. This work exploited the in vivo system to study the first effects of bortezomib on nonmyelamatous bones when applied locally and provide a hypothesis for the underlying mechanism of mode of action of bortezomib in this setting.
80 female Sprague-Dawley rats were randomized into five different treatment groups (n=16/group): 1) Empty (2) Calcium phosphate cement: CPC (3) CPC+ 100?gb (4) CPC+ 250?gb (5) CPC+ 2500?gb. A 2.5 mm drill hole was then created in the metaphysis of the left femur. The defect was then either filled with the previously mentioned substitutes or left empty to serve as a control. After 4 weeks femora were harvested followed by histological, histomorphometrical and immunohistochemical (BMP2; bone-morphogenic protein 2, OPG; osteoprotegerin, RANKL; Receptor activator of nuclear factor kappa-B ligand, ASMA; alpha smooth muscle actin, ED1; CD68 and DKK1 antibody) analysis. Cellular populations were tested using enzymehistochemistry Mass spectrometry and TOF-SIMS was used to assess the distribution of released bortezomib molecule. ECM proteins were investigated through immunohistochemistry. Statistical analysis was performed using Mann Whitney U test with Bonferroni correction.p<0.05 was considered significant.
Histomorphometric analysis showed a statistically significant increased bone formation in CPC250b when compared to empty (p=0.001), CPC (p=0.000), CPC100b (p=0.049) and CPC500b (p=0.0.049), respectively. The increased bone formation in case of CPC250b was associated to the increase in prominent bone formation markers i.e. BMP2 and OPG/RANKL levels when compared to all the other groups (p=0.000).There was a simultaneous down-regulation of the bone resorption markers in CPC250b (Oc/ BS; p=0.002) when compared to other groups. Mass spectrometry analysis could attribute this to the released bortezomib detected mostly around the drill hole defect. This was further confirmed by TOF-SIMS analysis which also detected boron peaks with increasing concentrations (250 and 500?g) in the same ROI. Finally low expression levels of Dkk1 were also detected in the CPC250b.
Optimal doses of bortezomib promote bone formation in non-myelomatous bones by stimulating osteoblastogenesis and inhibiting osteoclastogenesis.Our results shows bortezomib positive effect on bone remodeling which when combined with its antimyelomatic activity can be a potential candidate for MM treatment.