Dasatinib as a Bone-Modifying Agent: Anabolic and Anti-Resorptive Effects

Abstract

BackgroundBone loss, in malignant or non-malignant diseases, is caused by increased osteoclast resorption and/or reduced osteoblast bone formation, and is commonly associated with skeletal complications. Thus, there is a need to identify new agents capable of influencing bone remodeling. We aimed to further pre-clinically evaluate the effects of dasatinib (BMS-354825), a multitargeted tyrosine kinase inhibitor, on osteoblast and osteoclast differentiation and function.MethodsFor studies on osteoblasts, primary human bone marrow mensenchymal stem cells (hMSCs) together with the hMSC-TERT and the MG-63 cell lines were employed. Osteoclasts were generated from peripheral blood mononuclear cells (PBMC) of healthy volunteers. Skeletally-immature CD1 mice were used in the in vivo model.ResultsDasatinib inhibited the platelet derived growth factor receptor-β (PDGFR-β), c-Src and c-Kit phosphorylation in hMSC-TERT and MG-63 cell lines, which was associated with decreased cell proliferation and activation of canonical Wnt signaling. Treatment of MSCs from healthy donors, but also from multiple myeloma patients with low doses of dasatinib (2–5 nM), promoted its osteogenic differentiation and matrix mineralization. The bone anabolic effect of dasatinib was also observed in vivo by targeting endogenous osteoprogenitors, as assessed by elevated serum levels of bone formation markers, and increased trabecular microarchitecture and number of osteoblast-like cells. By in vitro exposure of hemopoietic progenitors to a similar range of dasatinib concentrations (1–2 nM), novel biological sequelae relative to inhibition of osteoclast formation and resorptive function were identified, including F-actin ring disruption, reduced levels of c-Fos and of nuclear factor of activated T cells 1 (NFATc1) in the nucleus, together with lowered cathepsin K, αVβ3 integrin and CCR1 expression.ConclusionsLow dasatinib concentrations show convergent bone anabolic and reduced bone resorption effects, which suggests its potential use for the treatment of bone diseases such as osteoporosis, osteolytic bone metastasis and myeloma bone disease.

Preclinical Activity of the Oral Proteasome Inhibitor MLN9708 in Myeloma Bone Disease

“…In vitro osteoclast formation, resorption pits, F-actin ring formation, and CD51/61 expression Peripheral blood mononuclear cells (PBMCs) from healthy donors were differentiated by culture in osteoclastogenic medium (containing 25 ng/mL M-CSF and 50 ng/mL RANKL) as described in Garcia-Gomez and colleagues (26). Alternatively, PBMCs from patients with myeloma were also used.…”

“…In vitro osteoblast differentiation, alkaline phosphatase activity, and mineralization assays Osteoblasts were generated from mesenchymal osteoprogenitors by culture in osteogenic medium (containing 5 mmol/L b-glycerophosphate, 50 mg/mL ascorbic acid, and 80 nmol/L dexamethasone) and assayed as in GarciaGomez and colleagues (26). Briefly, primary MSCs (passage 2-3) or the hMSC-TERT cell line were exposed to different MLN2238 or bortezomib concentrations and cultured in osteogenic medium for 11 or 21 days for alkaline phosphatase (ALP) activity and matrix mineralization assays, respectively.…”

“…Generation of protein lysates and Western blotting were performed following standard procedures (26). A luminol detection system with p-iodophenol enhancement for chemiluminescence was used for protein detection.…”

“…One femur of each animal was fixed in 10% formalin and scanned using a microcomputed tomography (microCT) system (MicroCATII; Siemens) as described previously (26). Analysis of trabecular microarchitecture in the distal femur was performed using BoneJ (29), a bone morphometry image analysis ImageJ plugin.…”

“…Bone histologic evaluation on paraffin sections was performed after hematoxylin-eosin or immunohistochemical staining for TCF4 as previously described (26).…”