Gefitinib reduces the production of osteoclast differentiation factors in human bone marrow stromal cells: Implications for treatment of bone metastasis

Abstract

3119 Background: Clinical studies of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) as monotherapy in breast cancer patients have shown some evidence of activity. Significant relief of bone pain in patients with bone metastases was observed in a clinical trial of the EGFR-TKI gefitinib (IRESSA) in patients with metastatic breast cancer. Methods: The expression and functional role of EGFR, as well as the effects of gefitinib (4 μM) on the ability to produce osteoclastogenic factors, were investigated by reverse-transcriptase PCR, Real-Time PCR, Western blot analysis, immunoprecipitation, and/or ELISAs in 2 human bone marrow-derived continuous mesenchymal-stem-cell-like cell lines, HDS-1 and HDS-2. Results: Immunohistochemical analysis of bone marrow biopsies showed that human primary bone marrow stromal cells (BMSC) express immunoreactive EGFR. Expression of EGFR mRNA and protein was also demonstrated in both HDS cell lines. The levels of activated EGFR were significantly increased in HDS cells following treatment with EGF; this activation was blocked by gefitinib. Gefitinib reduced basal levels of activated EGFR and Akt in HDS cells. Treatment of HDS cells with gefitinib significantly reduced levels of secreted macrophage colony-stimulating factor (p<0.001, Student t-test) and cell-associated receptor activator of NF-kB ligand (RANKL) [p<0.05, Student t-test]. In contrast, no significant effects of gefitinib on the secretion of osteoprotegerin, a RANKL decoy receptor that inhibits osteoclast differentiation, were observed in HDS cells. Finally, the ability of conditioned medium from gefitinib-treated HDS cells to sustain the differentiation of pre-osteoclasts was reduced by ∼45% compared with untreated HDS cells (p<0.001, Student t-test). Conclusions: These data demonstrate that EGFR regulates the ability of BMSC to induce osteoclast differentiation and strongly support clinical trials of gefitinib in patients with breast cancer and bone metastases. IRESSA is a trademark of the AstraZeneca group of companies Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration AstraZeneca