title: Modulation of endochondral ossification by MEK inhibitors PD0325901 and AZD6244 (Selumetinib).
abstract: MEK inhibitors (MEKi) PD0325901 and AZD6244 (Selumetinib) are drugs currently under clinical investigation for cancer treatment, however the Ras-MAPK pathway is also an important mediator of normal bone cell differentiation and function. In this study we examined the effects of these compounds on endochondral processes using both in vitro and in vivo models. Treatment with PD0325901 or AZD6244 significantly increased Runx2 and Alkaline phosphate gene expression in calvarial osteoblasts and decreased TRAP+ cells in induced osteoclast cultures. To test the effects of these drugs on bone healing, C57/Bl6 mice underwent a closed tibial fracture and were treated with PD0325901 or AZD6244 at 10mg/kg/day. Animals were culled at day 10 and at day 21 post-fracture for analysis of the fracture callus and the femoral growth plate in the contralateral leg. MEKi treatment markedly increased cartilage volume in the soft callus at day 10 post-fracture (+60% PD0325901, +20% AZD6244) and continued treatment led to a delay in cartilage remodeling. At the growth plate, we observed an increase in the height of the hypertrophic zone relative to the proliferative zone of +78% in PD0325901 treated mice. Osteoclast surface was significantly decreased both at the terminal end of the growth plate and within the fracture calluses of MEKi treated animals. The mechanistic effects of MEKi on genes encoding cartilage matrix proteins and catabolic enzymes were examined in articular chondrocyte cultures. PD0325901 or AZD6244 led to increased matrix protein expression (Col2a1 and Acan) and decreased expression of catabolic factors (Mmp13 and Adamts-5). Taken together, these data support the hypothesis that MEKi treatment can impact chondrocyte hypertrophy, matrix resorption, and fracture healing. These compounds can also affect bone architecture by expanding the hypertrophic zone of the growth plate and reducing osteoclast surface systemically.
authors.affiliation: Orthopaedic Research & Biotechnology Unit, The Children's Hospital at Westmead, Sydney, Australia; Discipline of Paediatrics and Child Health, Faculty of Medicine, University of Sydney, Sydney, Australia.
authors.forename: J
authors.initials: J
authors.lastname: El-Hoss
authors.affiliation: Orthopaedic Research & Biotechnology Unit, The Children's Hospital at Westmead, Sydney, Australia.
authors.forename: M
authors.initials: M
authors.lastname: Kolind
authors.affiliation: Raymond Purves Bone and Joint Research Laboratories, Kolling Institute of Medical Research, University of Sydney, Royal North Shore Hospital, Sydney, Australia.
authors.forename: M T
authors.initials: MT
authors.lastname: Jackson
authors.affiliation: Orthopaedic Research & Biotechnology Unit, The Children's Hospital at Westmead, Sydney, Australia; Discipline of Paediatrics and Child Health, Faculty of Medicine, University of Sydney, Sydney, Australia.
authors.forename: N
authors.initials: N
authors.lastname: Deo
authors.affiliation: Orthopaedic Research & Biotechnology Unit, The Children's Hospital at Westmead, Sydney, Australia.
authors.forename: K
authors.initials: K
authors.lastname: Mikulec
authors.affiliation: Bone Biology Group, Garvan Institute for Medical Research, Sydney, Australia.
authors.forename: M M
authors.initials: MM
authors.lastname: McDonald
authors.affiliation: Raymond Purves Bone and Joint Research Laboratories, Kolling Institute of Medical Research, University of Sydney, Royal North Shore Hospital, Sydney, Australia.
authors.forename: C B
authors.initials: CB
authors.lastname: Little
authors.affiliation: Orthopaedic Research & Biotechnology Unit, The Children's Hospital at Westmead, Sydney, Australia; Discipline of Paediatrics and Child Health, Faculty of Medicine, University of Sydney, Sydney, Australia.
authors.forename: D G
authors.initials: DG
authors.lastname: Little
authors.affiliation: Orthopaedic Research & Biotechnology Unit, The Children's Hospital at Westmead, Sydney, Australia; Discipline of Paediatrics and Child Health, Faculty of Medicine, University of Sydney, Sydney, Australia. Electronic address: aaron.schindeler@sydney.edu.au.
authors.forename: A
authors.initials: A
authors.lastname: Schindeler
mesh_terms: D000818:Animals; D001549:Benzamides / Q000494:pharmacology*; D001562:Benzimidazoles / Q000494:pharmacology*; D001854:Bone Marrow Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002146:Bony Callus / Q000187:drug effects / Q000473:pathology; D002356:Cartilage / Q000187:drug effects / Q000254:growth & development; D002454:Cell Differentiation / Q000187:drug effects; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D004159:Diphenylamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D017102:Fracture Healing / Q000187:drug effects; D020935:MAP Kinase Signaling System / Q000187:drug effects; D051379:Mice; D008810:Mice, Inbred C57BL; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D010010:Osteoclasts / Q000187:drug effects / Q000201:enzymology; D010012:Osteogenesis / Q000187:drug effects*; D053244:Osteoprotegerin / Q000097:blood; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D053245:RANK Ligand / Q000097:blood; D012756:Sheep; D012886:Skull / Q000166:cytology; D055114:X-Ray Microtomography
chemical_list: C517975:AZD 6244; D001549:Benzamides; D001562:Benzimidazoles; D053244:Osteoprotegerin; D047428:Protein Kinase Inhibitors; D053245:RANK Ligand; C506614:mirdametinib; D004159:Diphenylamine; D020929:Mitogen-Activated Protein Kinase Kinases
keywords: Cancer; Cartilage remodeling; Endochondral ossification; Fracture healing; Ras–MAPK