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Erlotinib inhibits osteolytic bone invasion of human non-small-cell lung cancer cell line NCI-H292.

Furugaki K, Moriya Y, Iwai T, Yorozu K, Yanagisawa M, Kondoh K, Fujimoto-Ohuchi K, Mori K - Clin. Exp. Metastasis (2011)

Bottom Line: In this model, NCI-H292 cells markedly activated osteoclasts in tibia, which resulted in osteolytic bone destruction.Furthermore, erlotinib also inhibited osteoblast/stromal cell proliferation in vitro and the development of osteoclasts induced by RANKL in vitro.In conclusion, erlotinib inhibits tumor-induced osteolytic invasion in bone metastasis by suppressing osteoclast activation through inhibiting tumor growth at the bone metastatic sites, osteolytic factor production in tumor cells, osteoblast/stromal cell proliferation and osteoclast differentiation from mouse bone marrow cells.

View Article: PubMed Central - PubMed

Affiliation: Product Research Department, Kamakura Research Laboratories, Chugai Pharmaceutical Co., Ltd., Kajiwara, Kamakura, Kanagawa, Japan.

ABSTRACT
Previous preclinical and clinical findings have suggested a potential role of epidermal growth factor receptor (EGFR) in osteoclast differentiation and the pathogenesis of bone metastasis in cancer. In this study, we investigated the effect of erlotinib, an orally active EGFR tyrosine kinase inhibitor (TKI), on the bone invasion of human non-small-cell lung cancer (NSCLC) cell line NCI-H292. First, we established a novel osteolytic bone invasion model of NCI-H292 cells which was made by inoculating cancer cells into the tibia of scid mice. In this model, NCI-H292 cells markedly activated osteoclasts in tibia, which resulted in osteolytic bone destruction. Erlotinib treatment suppressed osteoclast activation to the basal level through suppressing receptor activator of NF-κB ligand (RANKL) expression in osteoblast/stromal cell at the bone metastatic sites, which leads to inhibition of osteolytic bone destruction caused by NCI-H292 cells. Erlotinib inhibited the proliferation of NCI-H292 cells in in vitro. Erlotinib suppressed the production of osteolytic factors, such as parathyroid hormone-related protein (PTHrP), IL-8, IL-11 and vascular endothelial growth factor (VEGF) in NCI-H292 cells. Furthermore, erlotinib also inhibited osteoblast/stromal cell proliferation in vitro and the development of osteoclasts induced by RANKL in vitro. In conclusion, erlotinib inhibits tumor-induced osteolytic invasion in bone metastasis by suppressing osteoclast activation through inhibiting tumor growth at the bone metastatic sites, osteolytic factor production in tumor cells, osteoblast/stromal cell proliferation and osteoclast differentiation from mouse bone marrow cells.

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Effects of erlotinib on cell proliferation of mouse and human osteoblasts/stromal cells and osteoclast differentiation from mouse bone marrow cells. a EGFR expression detected by immunoblotting with anti-EGFR antibody. b and c Cell proliferation was determined by MTT assay after 4 days of treatment with erlotinib. Each point represents mean + SD of triplicates. d Mouse bone marrow cells were cultured in osteoclast differentiation medium with or without erlotinib for 12 days. Micrographs of TRAP staining. e The number of multinucleated (>3 nuclei) TRAP-positive osteoclasts was measured. Each point represents mean + SD of quadruplicates. *P < 0.05, by t test, compared with vehicle-treated group
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Fig5: Effects of erlotinib on cell proliferation of mouse and human osteoblasts/stromal cells and osteoclast differentiation from mouse bone marrow cells. a EGFR expression detected by immunoblotting with anti-EGFR antibody. b and c Cell proliferation was determined by MTT assay after 4 days of treatment with erlotinib. Each point represents mean + SD of triplicates. d Mouse bone marrow cells were cultured in osteoclast differentiation medium with or without erlotinib for 12 days. Micrographs of TRAP staining. e The number of multinucleated (>3 nuclei) TRAP-positive osteoclasts was measured. Each point represents mean + SD of quadruplicates. *P < 0.05, by t test, compared with vehicle-treated group

Mentions: Recently, the role of EGFR in osteoblast and osteoclast has been focused on [5, 24]. Therefore, we examined the effects of erlotinib on these cells. The mouse primary osteoblasts, mouse osteoblastic cell line MC3T3-E1, mouse stromal cell line ST2, human osteoblastic cell line MG-63 and human stromal cell line HS-5 expressed EGFR (Fig. 5a). Erlotinib inhibited the proliferation of these cell lines in vitro (Fig. 5b, c). In contrast, the reduced cell proliferative activity of osteoblasts/stromal cells in vivo by erlotinib treatment could not be estimated, because there was no detectable Ki-67 immunostaining in osteoblasts/stromal cells with or without erlotinib treatment (data not shown). In addition, erlotinib completely suppressed the formation of mononuclear and multinuclear TRAP-positive cells in a dose-dependent manner (Fig. 5d, e), although erlotinib did not exhibit any cytotoxity against bone marrow cells in this condition (data not shown).Fig. 5


Erlotinib inhibits osteolytic bone invasion of human non-small-cell lung cancer cell line NCI-H292.

Furugaki K, Moriya Y, Iwai T, Yorozu K, Yanagisawa M, Kondoh K, Fujimoto-Ohuchi K, Mori K - Clin. Exp. Metastasis (2011)

Effects of erlotinib on cell proliferation of mouse and human osteoblasts/stromal cells and osteoclast differentiation from mouse bone marrow cells. a EGFR expression detected by immunoblotting with anti-EGFR antibody. b and c Cell proliferation was determined by MTT assay after 4 days of treatment with erlotinib. Each point represents mean + SD of triplicates. d Mouse bone marrow cells were cultured in osteoclast differentiation medium with or without erlotinib for 12 days. Micrographs of TRAP staining. e The number of multinucleated (>3 nuclei) TRAP-positive osteoclasts was measured. Each point represents mean + SD of quadruplicates. *P < 0.05, by t test, compared with vehicle-treated group
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3198194&req=5

Fig5: Effects of erlotinib on cell proliferation of mouse and human osteoblasts/stromal cells and osteoclast differentiation from mouse bone marrow cells. a EGFR expression detected by immunoblotting with anti-EGFR antibody. b and c Cell proliferation was determined by MTT assay after 4 days of treatment with erlotinib. Each point represents mean + SD of triplicates. d Mouse bone marrow cells were cultured in osteoclast differentiation medium with or without erlotinib for 12 days. Micrographs of TRAP staining. e The number of multinucleated (>3 nuclei) TRAP-positive osteoclasts was measured. Each point represents mean + SD of quadruplicates. *P < 0.05, by t test, compared with vehicle-treated group
Mentions: Recently, the role of EGFR in osteoblast and osteoclast has been focused on [5, 24]. Therefore, we examined the effects of erlotinib on these cells. The mouse primary osteoblasts, mouse osteoblastic cell line MC3T3-E1, mouse stromal cell line ST2, human osteoblastic cell line MG-63 and human stromal cell line HS-5 expressed EGFR (Fig. 5a). Erlotinib inhibited the proliferation of these cell lines in vitro (Fig. 5b, c). In contrast, the reduced cell proliferative activity of osteoblasts/stromal cells in vivo by erlotinib treatment could not be estimated, because there was no detectable Ki-67 immunostaining in osteoblasts/stromal cells with or without erlotinib treatment (data not shown). In addition, erlotinib completely suppressed the formation of mononuclear and multinuclear TRAP-positive cells in a dose-dependent manner (Fig. 5d, e), although erlotinib did not exhibit any cytotoxity against bone marrow cells in this condition (data not shown).Fig. 5

Bottom Line: In this model, NCI-H292 cells markedly activated osteoclasts in tibia, which resulted in osteolytic bone destruction.Furthermore, erlotinib also inhibited osteoblast/stromal cell proliferation in vitro and the development of osteoclasts induced by RANKL in vitro.In conclusion, erlotinib inhibits tumor-induced osteolytic invasion in bone metastasis by suppressing osteoclast activation through inhibiting tumor growth at the bone metastatic sites, osteolytic factor production in tumor cells, osteoblast/stromal cell proliferation and osteoclast differentiation from mouse bone marrow cells.

View Article: PubMed Central - PubMed

Affiliation: Product Research Department, Kamakura Research Laboratories, Chugai Pharmaceutical Co., Ltd., Kajiwara, Kamakura, Kanagawa, Japan.

ABSTRACT
Previous preclinical and clinical findings have suggested a potential role of epidermal growth factor receptor (EGFR) in osteoclast differentiation and the pathogenesis of bone metastasis in cancer. In this study, we investigated the effect of erlotinib, an orally active EGFR tyrosine kinase inhibitor (TKI), on the bone invasion of human non-small-cell lung cancer (NSCLC) cell line NCI-H292. First, we established a novel osteolytic bone invasion model of NCI-H292 cells which was made by inoculating cancer cells into the tibia of scid mice. In this model, NCI-H292 cells markedly activated osteoclasts in tibia, which resulted in osteolytic bone destruction. Erlotinib treatment suppressed osteoclast activation to the basal level through suppressing receptor activator of NF-κB ligand (RANKL) expression in osteoblast/stromal cell at the bone metastatic sites, which leads to inhibition of osteolytic bone destruction caused by NCI-H292 cells. Erlotinib inhibited the proliferation of NCI-H292 cells in in vitro. Erlotinib suppressed the production of osteolytic factors, such as parathyroid hormone-related protein (PTHrP), IL-8, IL-11 and vascular endothelial growth factor (VEGF) in NCI-H292 cells. Furthermore, erlotinib also inhibited osteoblast/stromal cell proliferation in vitro and the development of osteoclasts induced by RANKL in vitro. In conclusion, erlotinib inhibits tumor-induced osteolytic invasion in bone metastasis by suppressing osteoclast activation through inhibiting tumor growth at the bone metastatic sites, osteolytic factor production in tumor cells, osteoblast/stromal cell proliferation and osteoclast differentiation from mouse bone marrow cells.

Show MeSH
Related in: MedlinePlus