The FGFR3 receptor tyrosine kinase represents an attractive target for therapy due to its role in several human disorders, including skeletal dysplasias, multiple myeloma, and cervical and bladder carcinomas. By using molecular library screening, we identified a compound named NF449 with inhibitory activity toward FGFR3 signaling. In cultured chondrocytes and murine limb organ culture, NF449 rescued FGFR3-mediated extracellular matrix loss and growth inhibition, which represent two major cellular phenotypes of aberrant FGFR3 signaling in cartilage. Similarly, NF449 antagonized FGFR3 action in the multiple myeloma cell lines OPM2 and KMS11, as evidenced by NF449-mediated reversal of ERK MAPK activation and transcript accumulation of CCL3 and CCL4 chemokines, both of which are induced by FGFR3 activation. In cell-free kinase assays, NF449 inhibited the kinase activity of both wild type and a disease-associated FGFR3 mutant (K650E) in a fashion that appeared non-competitive with ATP. Our data identify NF449 as a novel antagonist of FGFR3 signaling, useful for FGFR3 inhibition alone or in combination with inhibitors that target the ATP binding site.FGFR3 (fibroblast growth factor receptor 3) is a transmembrane tyrosine kinase that serves as a receptor for the members of the fibroblast growth factor (FGF) 2 family and functions in many biological processes, including cell proliferation, differentiation, migration, and survival. To date, activating mutations in FGFR3 have been associated with several human disorders, such as skeletal dysplasias, multiple myeloma, and cervical and bladder carcinomas (1-4).Among the skeletal dysplasias, activating FGFR3 mutations cause achondroplasia, the most common form of human skeletal dysplasia, and thanatophoric dysplasia, the most common form of lethal skeletal dysplasia (1). Long bones of individuals suffering from FGFR3-related skeletal dysplasias show markedly shortened zones of chondrocyte proliferation and differentiation, whereas Fgfr3 knock-out mice and humans without functional FGFR3 demonstrate skeletal overgrowth, together implying the role of FGFR3 as a negative regulator of bone growth (5).Apart from cartilage, ϳ15% of patients suffering from multiple myeloma markedly up-regulate FGFR3 as a consequence of a t(4;14)(p16.3;q32) translocation, with a fraction of patients also harboring activating mutations in FGFR3, identical to those found in the skeletal dysplasias (2, 3). Ectopic expression of FGFR3 enhances multiple myeloma cell proliferation and survival, demonstrating the oncogenic potential of FGFR3 (6).Given its role in human disease, FGFR3 signaling represents an attractive target for therapy. There is no treatment available for achondroplasia at present, thus inciting the development of novel approaches to target FGFR3. The aim of this study was to identify novel inhibitors of FGFR3 signaling in cellular environments relevant to FGFR3-related disorders. We recently established a chondrocyte cell-based reporter assay suitable for identification of inhibitors of ...

Oncogenic activation of the RAS-ERK MAP kinase signaling pathway can lead to uncontrolled proliferation but can also result in apoptosis or premature cellular senescence, both regarded as natural protective barriers to cell immortalization and transformation. In FGFR3-related skeletal dyplasias, oncogenic mutations in the FGFR3 receptor tyrosine kinase cause profound inhibition of cartilage growth resulting in severe dwarfism, although many of the precise mechanisms of FGFR3 action remain unclear. Mutated FGFR3 induces constitutive activation of the ERK pathway in chondrocytes and, remarkably, can also cause both increased proliferation and apoptosis in growing cartilage, depending on the gestational age. Here, we demonstrate that FGFR3 signaling is also capable of inducing premature senescence in chondrocytes, manifested as reversible, ERK-dependent growth arrest accompanied by alteration of cellular shape, loss of the extracellular matrix, upregulation of senescence markers (α-GLUCOSIDASE, FIBRONECTIN, CAVEOLIN 1, LAMIN A, SM22α and TIMP 1), and induction of senescence-associated β-GALACTOSIDASE activity. Our data support a model whereby FGFR3 signaling inhibits cartilage growth via exploiting cellular responses originally designed to eliminate cells harbouring activated oncogenes.

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