J El-Hoss1, T Cheng1, E C Carpenter1, K Sullivan1, N Deo1, K Mikulec1, D G Little1, A Schindeler1. 1. Orthopaedic Research & Biotechnology Unit, The Children's Hospital at Westmead, Locked Bag 4001, Westmead, NSW 2145, Australia. E-mail address for J. El-Hoss: jel-hoss@ccia.unsw.edu.au. E-mail address for T. Cheng: tegan.cheng@sydney.edu.au. E-mail address for E.C. Carpenter: carpie@doctors.org.uk. E-mail address for K. Sullivan: kates3@live.com. E-mail address for N. Deo: nikita.deo@sydney.edu.au. E-mail address for K. Mikulec: kathy.mikulec@health.nsw.gov.au. E-mail address for D.G. Little: david.little@health.nsw.gov.au. E-mail address for A. Schindeler: aaron.schindeler@sydney.edu.au.
Abstract
BACKGROUND: Congenital tibial dysplasia is a severe pediatric condition that classically results in a persistent pseudarthrosis. A majority of these cases are associated with neurofibromatosis type I (NF1), a genetic disorder in which inactivation of the NF1 gene leads to overactivity of the Ras-MEK-MAPK (mitogen-activated protein kinase) signaling pathway. We therefore hypothesized that pharmaceutical inhibition of MEK-MAPK may be a beneficial therapeutic strategy. METHODS: In vitro methods were used to demonstrate a role for the MEK inhibitor PD0325901 in promoting osteogenic differentiation in Nf1-/- calvarial osteoblasts. Local applications of rhBMP-2 and/or PD0325901 were then tested in a mouse model of NF1 tibial pseudarthrosis featuring localized double inactivation of the Nf1 gene in a fracture. Mice received no treatment, PD0325901 (10 mg/kg/day from two days before fracture to ten days after fracture), rhBMP-2 (10 μg), or a combination of rhBMP-2 and PD0325901. RESULTS: Animals treated with the delivery vehicle alone, PD0325901, rhBMP-2, or the PD0325901 + rhBMP-2 combination showed union rates of 0%, 8%, 69% (p < 0.01), or 80% (p < 0.01), respectively, at twenty-one days after fracture. Mice treated with the rhBMP-2 + PD0325901 combination displayed a callus volume sixfold greater than the vehicle controls and twofold greater than the group receiving rhBMP-2 alone. Although MEK inhibition combined with rhBMP-2 led to increases in bone formation and union, the proportion of fibrous tissue in the callus was not significantly reduced. CONCLUSIONS: The data suggest that MEK inhibition can promote bone formation in combination with rhBMP-2 in the context of an NF1 pseudarthrosis. However, PD0325901 did not promote substantive bone anabolism in the absence of an exogenous anabolic stimulus and did not suppress fibrosis. CLINICAL RELEVANCE: This study examines a signaling pathway-based approach to treating poor bone healing in a model of NF1 pseudarthrosis.
BACKGROUND:Congenital tibial dysplasia is a severe pediatric condition that classically results in a persistent pseudarthrosis. A majority of these cases are associated with neurofibromatosis type I (NF1), a genetic disorder in which inactivation of the NF1 gene leads to overactivity of the Ras-MEK-MAPK (mitogen-activated protein kinase) signaling pathway. We therefore hypothesized that pharmaceutical inhibition of MEK-MAPK may be a beneficial therapeutic strategy. METHODS: In vitro methods were used to demonstrate a role for the MEK inhibitor PD0325901 in promoting osteogenic differentiation in Nf1-/- calvarial osteoblasts. Local applications of rhBMP-2 and/or PD0325901 were then tested in a mouse model of NF1tibial pseudarthrosis featuring localized double inactivation of the Nf1 gene in a fracture. Mice received no treatment, PD0325901 (10 mg/kg/day from two days before fracture to ten days after fracture), rhBMP-2 (10 μg), or a combination of rhBMP-2 and PD0325901. RESULTS: Animals treated with the delivery vehicle alone, PD0325901, rhBMP-2, or the PD0325901 + rhBMP-2 combination showed union rates of 0%, 8%, 69% (p < 0.01), or 80% (p < 0.01), respectively, at twenty-one days after fracture. Mice treated with the rhBMP-2 + PD0325901 combination displayed a callus volume sixfold greater than the vehicle controls and twofold greater than the group receiving rhBMP-2 alone. Although MEK inhibition combined with rhBMP-2 led to increases in bone formation and union, the proportion of fibrous tissue in the callus was not significantly reduced. CONCLUSIONS: The data suggest that MEK inhibition can promote bone formation in combination with rhBMP-2 in the context of an NF1pseudarthrosis. However, PD0325901 did not promote substantive bone anabolism in the absence of an exogenous anabolic stimulus and did not suppress fibrosis. CLINICAL RELEVANCE: This study examines a signaling pathway-based approach to treating poor bone healing in a model of NF1pseudarthrosis.