scholarly journals Regulation of bone resorption and sealing zone formation in osteoclasts occurs through protein kinase b-mediated microtubule stabilization

2013 ◽  
Vol 28 (5) ◽  
pp. 1191-1202 ◽  
Author(s):  
Takumi Matsumoto ◽  
Yuichi Nagase ◽  
Jun Hirose ◽  
Naoto Tokuyama ◽  
Tetsuro Yasui ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Bone Resorption ◽  
Protein Kinase B ◽  
Zone Formation ◽  
Microtubule Stabilization ◽  
Sealing Zone

Specific Antagonists of NMDA Receptors Prevent Osteoclast Sealing Zone Formation Required for Bone Resorption

2000 ◽  
Vol 268 (1) ◽  
pp. 201-209 ◽  
Author(s):  
Cécile Itzstein ◽  
Léon Espinosa ◽  
Pierre D. Delmas ◽  
Chantal Chenu
Keyword(s):  
Bone Resorption ◽  
Nmda Receptors ◽  
Zone Formation ◽  
Sealing Zone

A Novel Bruton′s Tyrosine Kinase Inhibitor CC-292 In Combination With The Proteasome Inhibitor Carfilzomib Iimpacts Multiple Myeloma Bone Microenviroment With Resultant Anti-Myeloma Activity

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 682-682 ◽  
Author(s):  
Homare Eda ◽  
Loredana Santo ◽  
Diana D Cirstea ◽  
Andrew J Yee ◽  
Tyler A Scullen ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Tyrosine Kinase ◽  
Proteasome Inhibitor ◽  
Bone Volume ◽  
Employment Equity ◽  
Zone Formation ◽  
Sealing Zone ◽  
Btk Inhibitor ◽  
Equity Ownership ◽  
Rank Signaling

Abstract A member of the Tec family kinases, Bruton’s tyrosine kinase (Btk) modulates B-cell development and activation, and plays an important role in antibody production. Interestingly, Btk and Tec (the other Tec kinase family) regulate osteoclast (OC) differentiation via Receptor Activator of Nuclear Factor κ B (RANK) signaling. Moreover, OCs derived from X-linked agammaglobulinemia (XLA) patients who harbor Btk null mutations have impaired function. Here we show that a potent and specific Btk inhibitor, CC-292 inhibits OC function in multiple myeloma (MM) patients. CC-292 is a highly selective, covalent Btk inhibitor. OC derived from MM patient monocytes were assayed with or without CC-292. Interestingly, OC function was significantly inhibited in the presence of CC-292 (100 nM and 1000 nM) as demonstrated by pit formation assay. However, mRNA expression for TRAP and Cathepsin K, two OC differentiation markers were increased in the presence of CC-292 suggesting that CC-292 inhibits OC function without inhibiting OC differentiation. OC sealing zone contributes to OC bone resorption function. Given the role of c-Src and Proline-rich tyrosine Kinase 2 (Pyk2) signaling in sealing zone formation and OC function we next evaluated CC-292’s effect on Pyk2 and c-Src. Pyk2 plays a role in OC activation and localizes to the sealing zone in OC. RANK signaling activates c-Src, which phosphorylates Pyk2. Moreover c-Src controls OC bone resorption by regulating actin organization via cortactin. Interestingly, CC-292 (100 nM) inhibited c-Src total protein, c-Src phosphorylation and Pyk2 phosphorylation. Furthermore, CC-292 (100 nM) inhibited cortactin protein and mRNA expression, and upregulated c-Cbl protein (E3 ubiquitin ligase for c-Src) expression in OC derived from MM patient monocytes with resultant inhibition of OC sealing zone formation. However, at the same low doses (100 nM) CC-292 did not show any direct in vitro effect against MM cell viability. Because carfilzomib, a proteasome inhibitor that binds irreversibly to its target, has potent anti-MM activity and also inhibits OC resorptive activity, we studied CC-292 in combination with carfilzomib. Our data suggests that carfilzomib (1.25 nM) has no impact on OC sealing zone formation but inhibits OC differentiation. CC-292 (100 nM) in combination with carfilzomib (1.25 nM) inhibited not only sealing zone formation but also OC differentiation, resulting in stronger suppression of OC function than carfilzomib alone. The combination of CC-292 (30mg/kg p.o. for 5 days per week for 6 weeks) and carfilzomib (3 mg/kg i.v. x 2 days per week for 4 weeks and 2 mg/kg i.v. x 2 days per week for 2 weeks) significantly inhibited tumor burden and myeloma cell numbers in a diffuse NOD-SCID MM model. The calvarial cells derived from these mice treated with CC-292 alone, carfilzomib alone or the combination showed higher osteocalcin mRNA (osteoblast differentiation marker) expression. A specific bone resorption marker, carboxy-terminal telopeptide collagen crosslinks (CTX) in mouse serum was significantly inhibited in CC-292 and CC-292 in combination with carfilzomib treatment groups in comparison with control mice. Furthermore, 3D microCT reconstructions showed increase in cancellous bone volume in lumbar vertebrae in mice treated with CC-292 or carfilzomib, while the combination treatment resulted in an increase in cancellous bone volume in an additive manner. These data demonstrate that the novel BTK inhibitor CC-292 inhibits OC function through inhibition of OC sealing zone formation. Moreover, CC-292 in combination with carfilzomib augments effects against the bone microenvironment with resultant anti-MM activity. Disclosures: Arastu-Kapur: Onyx Pharmaceuticals, Inc.: Employment. Evans:Celgene Avilomics Research: Employment, Equity Ownership. Singh:Celgene Avilomics Research: Employment, Equity Ownership. Kirk:Onyx Pharmaceuticals, Inc.: Employment. Westlin:Celgene Avilomics Research: Employment, Equity Ownership. Raje:Celgene: Consultancy; Millenium: Consultancy; Onyx: Consultancy; Amgen: Consultancy; Acetylon: Research Funding; Eli Lilly: Research Funding.

scholarly journals Apatite-mediated Actin Dynamics in Resorbing Osteoclasts

2004 ◽  
Vol 15 (12) ◽  
pp. 5231-5241 ◽  
Author(s):  
Frédéric Saltel ◽  
Olivier Destaing ◽  
Frédéric Bard ◽  
Diane Eichert ◽  
Pierre Jurdic
Keyword(s):  
Bone Resorption ◽  
Actin Dynamics ◽  
Cell Periphery ◽  
Main Function ◽  
Zone Formation ◽  
Actin Structure ◽  
Sealing Zone ◽  
Mineralized Matrix ◽  
And Migration

The actin cytoskeleton is essential for osteoclasts main function, bone resorption. Two different organizations of actin have been described in osteoclasts, the podosomes belt corresponding to numerous F-actin columns arranged at the cell periphery, and the sealing zone defined as a unique large band of actin. To compare the role of these two different actin organizations, we imaged osteoclasts on various substrata: glass, dentin, and apatite. Using primary osteoclasts expressing GFP-actin, we found that podosome belts and sealing zones, both very dynamic actin structures, were present in mature osteoclasts; podosome belts were observed only in spread osteoclasts adhering onto glass, whereas sealing zone were seen in apico-basal polarized osteoclasts adherent on mineralized matrix. Dynamic observations of several resorption cycles of osteoclasts seeded on apatite revealed that 1) podosomes do not fuse together to form the sealing zone; 2) osteoclasts alternate successive stationary polarized resorption phases with a sealing zone and migration, nonresorption phases without any specific actin structure; and 3) apatite itself promotes sealing zone formation though c-src and Rho signaling. Finally, our work suggests that apatite-mediated sealing zone formation is dependent on both c-src and Rho whereas apico-basal polarization requires only Rho.

AVL-292, a Bruton's Tyrosine Kinase Inhibitor Impacts Bone Resorption by Abrogating Osteoclast Sealing Zone Formation in Multiple Myeloma

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1822-1822
Author(s):  
Homare Eda ◽  
Loredana Santo ◽  
Diana Cirstea ◽  
Andrew J. Yee ◽  
Anuj Mahindra ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Tyrosine Kinase ◽  
Mrna Expression ◽  
Mechanism Of Action ◽  
Research Funding ◽  
Bruton's Tyrosine Kinase ◽  
Zone Formation ◽  
Sealing Zone ◽  
Btk Inhibitor ◽  
Rank Signaling

Abstract Abstract 1822 The activation of Bruton's tyrosine kinase (Btk), a member of the Tec family of tyrosine kinases regulates B-cell development and activation, and plays a key role in antibody production. Interestingly, Btk and the other Tec kinase family, Tec, regulate OC differentiation via Receptor Activator of Nuclear Factor κ B (RANK) signaling. Moreover, patients with X-linked agammaglobulinemia (XLA) who harbor Btk null mutations have impaired OC function. Here we show that, a potent and specific inhibitor of Btk, AVL-292, inhibits OC function in MM patients. AVL-292 is a highly selective, covalent Btk inhibitor that potently silences Btk enzymatic activity (IC50 < 0.5nM) with high selectivity towards Btk with lack of significant inhibition against other kinases involved in BCR signaling (Syk, Lyn). We examined the mechanism of action of AVL-292 in the context of OCs function. OC derived from MM patient monocytes were assayed with or without AVL-292. Interestingly, OC function was inhibited in the presence AVL-292 as demonstrated by pit formation assay. However, mRNA expression of Cathepsin K and TRAP, markers of OC differentiation, were increased in the presence of AVL-292. These data suggest AVL-292 inhibits OC function without affecting the OC differentiation. It has been shown that BTK and Tec regulation of OC differentiation is related to calcium (Ca2+) signaling by increasing Ca2+ flux through direct phosphorylation of phospholipase C2 (PLCγ2). To delineate the mechanism of action of the BTK inhibitor against OC function, the RANK signaling proteins were detected by western blotting and Ca2+ concentration was measured by flourescence. AVL-292 inhibited phosphorylation of the BTK substrate, PLCγ2 in OCs. This was associated with an inhibition of intracellular Ca2+ release by AVL-292 which otherwise increased with RANKL stimulation in OCs. Given the effect of AVL-292 on RANK signaling we next evaluated its effect on Proline-rich tyrosine Kinase 2 (Pyk2) and c-Src. Pyk2 plays a role in OC activation and localizes to the sealing zone in OC. RANK signaling activates c-Src, which phosphorylates Pyk2. Moreover c-Src controls OC bone resorption by regulating actin organization via cortactin. Interestingly, AVL-292 inhibited c-Src and Pyk2 phosphorylation. Furthermore, AVL-292 inhibited cortactin protein and mRNA expression, and upregulated c-Cbl protein (E3 ubiquitin ligase for c-Src) expression in OCs derived from MM patient monocytes. These data demonstrate that the novel BTK inhibitor AVL-292 inhibits OC function through inhibition of OC sealing zone formation. These results suggest a potential novel mechanism of Btk inhibition with AVL-292 on osteoclast function and therefore bone resorption. Disclosures: Evans: Celgene: Employment. Singh:Celgene: Employment. Westlin:Celgene: Employment. Raje:Onyx: Consultancy; Celgene: Consultancy; Millenium: Consultancy; Acetylon: Research Funding; Amgen: Research Funding; Eli-Lilly: Research Funding.

scholarly journals Defective microtubule-dependent podosome organization in osteoclasts leads to increased bone density in Pyk2−/− mice

2007 ◽  
Vol 178 (6) ◽  
pp. 1053-1064 ◽  
Author(s):  
Hava Gil-Henn ◽  
Olivier Destaing ◽  
Natalie A. Sims ◽  
Kazuhiro Aoki ◽  
Neil Alles ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Protein Tyrosine Kinase ◽  
Ectopic Expression ◽  
Cell Periphery ◽  
Wild Type ◽  
Zone Formation ◽  
Sealing Zone ◽  
Osteoclast Function ◽  
Actin Rings

The protein tyrosine kinase Pyk2 is highly expressed in osteoclasts, where it is primarily localized in podosomes. Deletion of Pyk2 in mice leads to mild osteopetrosis due to impairment in osteoclast function. Pyk2-null osteoclasts were unable to transform podosome clusters into a podosome belt at the cell periphery; instead of a sealing zone only small actin rings were formed, resulting in impaired bone resorption. Furthermore, in Pyk2-null osteoclasts, Rho activity was enhanced while microtubule acetylation and stability were significantly reduced. Rescue experiments by ectopic expression of wild-type or a variety of Pyk2 mutants in osteoclasts from Pyk2−/− mice have shown that the FAT domain of Pyk2 is essential for podosome belt and sealing zone formation as well as for bone resorption. These experiments underscore an important role of Pyk2 in microtubule-dependent podosome organization, bone resorption, and other osteoclast functions.

A role for the actin cytoskeleton in the hormonal and growth-factor-mediated activation of protein kinase B

2001 ◽  
Vol 353 (3) ◽  
pp. 735
Author(s):  
K. PEYROLLIER ◽  
E. HAJDUCH ◽  
A. GRAY ◽  
G. J. LITHERLAND ◽  
A. R. PRESCOTT ◽  
...  
Keyword(s):  
Growth Factor ◽  
Protein Kinase ◽  
Actin Cytoskeleton ◽  
Protein Kinase B
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