Effect of MEKi on NF1 pseudarthrosis primary cells

Background: Neurofibromatosis type 1 (NF1) is a genetic condition predisposing children to fracture pseudarthroses. MEK inhibitors are FDA-approved or under study for the treatment of malignant pathologies associated with NF1, though their potential to treat pseudarthrosis is largely unknown. Methods: Primary cells cultured from control bone (i.e., iliac crest) or fracture pseudarthroses from children with NF1 were treated with vehicle or the MEK inhibitors trametinib or selumetinib. Gene expression was evaluated by transcriptome sequencing (RNAseq), and activation of the downstream signaling pathway was evaluated by western blotting. Results were replicated in an independent cohort of patient pseudarthrosis-derived primary cells. Results: We confirmed increased MAPK signaling in pseudarthrosis samples compared to patient-matched control samples. Pseudarthrosis samples were reproducibly associated with reduced expression of gene signatures implicated in osteoblast differentiation, skeletal development, and formation of the extracellular matrix. Expression of these gene signatures was significantly rescued following treatment with selumetinib, and to a lesser extent trametinib, concomitant with reduced MAPK signaling activation. Conclusions: Our study utilized patient-derived primary cells to demonstrate the molecular signatures associated with fracture pseudarthrosis and how this molecular dysregulation was reversed with MEK inhibitor treatment. Clinical relevance: MEK inhibitors may be repurposed to promote healing of fracture pseudarthroses in children with NF1. Overall design: Primary cells were cultured from two cohorts of patients. The first cohort includes cells cultured from patient-matched control or fracture pseudarthrosis bone, while the second cohort includes only cells cultured from pseudarthrosis bone. All samples were treated with vehicle (dmso) or the MEK inhibitors trametinib or selumetinib prior to expression proviling by RNA-sequencing.