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The formation of the skeleton requires inductive signals that are balanced with their antagonists in a highly regulated negative feedback system. Inappropriate or excessive expression of BMPs (bone morphogenetic proteins) or their antagonists results in genetic disorders affecting the skeleton, such as fibrodysplasia ossificans progressiva. BMP signaling mediated through binding to its receptors is a critical step in the induction of abnormal ossification. Therefore, we hypothesized that engineering more effective inhibitors of this BMP-signaling process may lead to the development of therapies for such conditions. BMP4-induced heterotopic ossification was used as a model for testing the ability of the BMP antagonist Noggin to block de novo bone formation, either by local or systemic delivery. Since Noggin naturally acts locally, a Noggin mutein, hNOGΔB2, was engineered and was shown to circulate systemically, and its ability to block heterotopic ossification was tested in a mouse model with use of adenovirus-mediated somatic cell gene transfer. BMP4-induced heterotopic ossification can be prevented in vivo either by local delivery of wild-type Noggin or after somatic cell gene transfer of a Noggin mutein, hNOGΔB2. Furthermore, the data in the present study provide proof of concept that a naturally occurring factor can be engineered for systemic delivery toward a desirable pharmacological outcome. Clinical Relevance: Blocking bone formation is clinically relevant to disorders of heterotopic ossification in humans, such as fibrodysplasia ossificans progressiva. Furthermore, development of BMP antagonists as therapeutic agents may provide modalities for the treatment of other pathologic conditions that arise from aberrant expression of BMPs and/or from a lack of their antagonists. [ABSTRACT FROM AUTHOR]

Background: The activity of bone morphogenetic proteins (BMPs) is regulated extracellularly by several families of secreted, negatively-acting factors. These BMP antagonists participate in the control of a diverse range of embryonic processes, such as establishment of the dorsal-ventral axis, neural induction, and formation of joints in the developing skeletal system. The ongoing process of neurogenesis in the adult brain also requires inhibition of BMP ligand activity. To date, the three-dimensional structures of these antagonists as well as the nature of their interaction with ligand have remained unknown. Toward that end, we have determined the crystal structure of the antagonist Noggin bound to BMP-7. Methods: The complex of the two homodimeric proteins was preformed, isolated by size exclusion chromatography, and crystallized at neutral pH. To probe the molecular interface of the complex and to quantitate the activity of a human mutant form, variant Noggin proteins were produced and their binding affinities were measured in vitro. The correlation between binding affinity and biological activity was examined with Noggin-soaked beads implanted in the developing chick limb bud. Results and Conclusions: The structure of the complex reveals that Noggin inhibits BMP signaling by blocking the binding sites of both types of receptors (Type I and Type II), mimicking their modes of binding. The affinity of Noggin variants for BMP-7 correlated well with the inhibition of BMP-induced chondrogenesis in the chick limb bud, confirming that Noggin acts by sequestering the ligand in an inactive state. Interestingly, the scaffold of Noggin was found to contain a cystine knot topology and protein fold similar to that of BMPs, indicating that ligand and antagonist may have evolved from a common ancestral gene. Clinical Relevance: Mutations in the human Noggin locus (NOG) are associated with three similar yet distinct skeletal dysplasias: proximal symphalangism (SYM1),... [ABSTRACT FROM AUTHOR]