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A4.18 The effects of SRC-family kinase inhibitors on osteoclast development in vitro
  1. D Csete1,
  2. D Győri1,
  3. T Vántus2,
  4. G Kéri2,
  5. C Szántai-Kis2,
  6. A Mócsai1
  1. 1Department of Physiology, Semmelweis University School of Medicine and MTA-SE "Lendület" Inflammation Physiology Research Group, Budapest, Hungary
  2. 2Vichem Chemie Research Ltd., Budapest, Hungary

Abstract

Background and objectives Osteoclasts are the unique bone-resorbing cells of hematopoietic origin; their development is regulated primarily by M-CSF, RANKL and integrin-mediated interactions. Our workgroup has previously tested thousands of Src-family kinase inhibitors produced by the Vichem Ltd on neutrophil granulocytes, which cells show many similarities with osteoclasts regarding signalling pathways. One of the most effective inhibitors was the PD166326, an anti-leukaemia drug candidate. Here we tested the effect of PD166326 on osteoclasts.

Materials and methods Bone marrow cells were isolated from the long bones of C57Bl/6 mice on the 0. day and differentiated into osteoclasts in vitro in the presence of recombinant M-CSF and RANKL. Cultures were treated with PD166326 during different stages of osteoclastogenesis. Vehicle control samples received dimethyl sulfoxide (DMSO). Osteoclast differentiation was examined after 4 or 6 days by osteoclast-specific tartrate-resistant acid phosphatase (TRAP)-staining. Survival assay was performed at the 4th day using AnnexinV-PE and 7-AAD apoptosis and necrosis kit. Actin ring formation was observed with Lifeact EGFP transgenic mice.

Results The number and the size of developing osteoclasts strongly decreased in case of the early administration of the inhibitor. The treatment of the cultures by 10 nM PD166326 practically blocked osteoclast development. When we administered the inhibitor later, from the 4th day – when the last main point of osteoclast development happens – only minor reduction was observed. The tendency to apoptosis did not increase in the PD166326 treated cultures. The administration of PD166326 did not influence the degradation of actin rings.

Conclusions The anti-cancer drug PD166326 inhibited osteoclastogenesis in a very small concentration. We suppose that it has an effect on the early stage of osteoclast development. These data may be considered in the therapy of different cancers especially in metastatic bone diseases.

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