JNK inhibitors increase osteogenesis in Nf1-deficient cells

Neurofibromatosis type 1 (NF1) is an autosomal dominant disorder that is associated with a variety of manifestations, including orthopedic complications such as scoliosis and tibial pseudarthrosis. Orthopedic management of these skeletal complications is rendered more challenging due to a lack of standardized adjunctive pharmacotherapies. NF1 leads to disruption of the canonical Ras/Raf-1/MEK/ERK axis, and this has been associated with defects in bone anabolism. The roles of other non-canonical Ras effector pathways, such as the c-Jun N-terminal Kinase (JNK) pathway, are less well understood. In this study we examine the effects of an anthrapyrazolone inhibitor of JNK (SP600125) on inducible osteoprogenitors as well as Nf1-deficient and Nf1-null primary osteoblasts. C2C12 cells, which are highly responsive to rhBMP-2, were examined with exogenous rhBMP-2 and a range of SP600125 doses. Based on the expression of early and late bone markers and matrix mineralization, 10 μM SP600125 was found to be pro-osteogenic whether delivered concurrent with or following 2 days of rhBMP-2 treatment. Aberrant JNK activity was identified in Nf1-deficient osteoprogenitors (increased rhBMP-2 induced phospho-c-Jun) and in Nf1-null mature osteoblasts (increased total c-Jun). Next, SP600125 was used to treat these cells and was found to facilitate osteogenesis in Nf1-deficient osteoprogenitors, and in Nf1-null osteoblasts when given in conjunction with rhBMP-2. Outcome measures included alkaline phosphatase activity, matrix mineralization, and osteogenic gene expression. In summary, JNK inhibitors represent a class of potentially useful adjunctive agents for orthopedic medicine, particularly in the context of NF1.