RESUMO
A series of (hetero)arylpyrimidines agonists of the Wnt-beta-catenin cellular messaging system have been prepared. These compounds show activity in U2OS cells transfected with Wnt-3a, TCF-luciferase, Dkk-1 and tk-Renilla. Selected compounds show minimal GSK-3beta inhibition indicating that the Wnt-beta-catenin agonism activity most likely comes from interaction at Wnt-3a/Dkk-1. Two examples 1 and 25 show in vivo osteogenic activity in a mouse calvaria model. One example 1 is shown to activate non-phosphorylated beta-catenin formation in bone.
Assuntos
Imidazóis/química , Pirimidinas/química , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animais , Desenvolvimento Ósseo/efeitos dos fármacos , Linhagem Celular Tumoral , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Humanos , Imidazóis/síntese química , Imidazóis/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Pirimidinas/síntese química , Pirimidinas/farmacologia , Proteínas Recombinantes de Fusão/agonistas , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais , Crânio/metabolismo , Proteínas Wnt/agonistas , Proteínas Wnt/genética , Proteína Wnt3 , Proteína Wnt3A , beta Catenina/agonistasRESUMO
The biological activities of parathyroid hormone (PTH) on bone are quite complex as demonstrated by its catabolic and anabolic activities on the skeleton. Although there have been many reports describing genes that are regulated by PTH in osteoblast cells, the goal of this study was to utilize a well-established in vivo PTH anabolic treatment regimen to identify genes that mediate bone anabolic effects of PTH. We identified a gene we named PTH anabolic induced gene in bone (PAIGB) that has been reported as brain and acute leukemia cytoplasmic (BAALC). Therefore, using the latter nomenclature, we have discovered that BAALC is a PTH-regulated gene whose mRNA expression was selectively induced in rat tibiae nearly 100-fold (maximal) by a PTH 1-34 anabolic treatment regimen in a time-dependent manner. Although BAALC is broadly expressed, PTH did not regulate BAALC expression in other PTH receptor expressing tissues and we find that the regulation of BAALC protein by PTH in vivo is confined to mature osteoblasts. Further in vitro studies using rat UMR-106 osteoblastic cells show that PTH 1-34 rapidly induces BAALC mRNA expression maximally by 4 h while the protein was induced by 8 h. In addition to being regulated by PTH 1-34, BAALC expression can also be induced by other bone forming factors including PGE(2) and 1,25 dihydroxy vitamin D(3). We determined that BAALC is regulated by PTH predominantly through the cAMP/PKA pathway. Finally, we demonstrate in MC3T3-E1 osteoblastic cells that BAALC overexpression regulates markers of osteoblast differentiation, including downregulating alkaline phosphatase and osteocalcin expression while inducing osteopontin expression. We also demonstrate that these transcriptional responses mediated by BAALC are similar to the responses elicited by PTH 1-34. These data, showing BAALC overexpression can mimic the effect of PTH on markers of osteoblast differentiation, along with the observations that BAALC is induced selectively with a bone anabolic treatment regimen of PTH (not a catabolic treatment regimen), suggest that BAALC may be an important mediator of the PTH anabolic action on bone cell function.