The broad focus of our laboratory is to study the effect of the inhibitory components of the Insulin-like Growth Factor (IGF) system on bone metabolism and cancer. IGFs are potent mitogens that inhibit apoptosis and stimulate osteoblast differentiation. They are thought to play a pivotal role in the maintenance of skeletal mass throughout life. IGFs are produced by many cancers to stimulate growth and inhibit apoptosis. The activities of IGFs on cells depend not only on their local concentration but also on the local concentration of inhibitory and stimulatory IGF binding proteins (IGFBPs) that affect their binding to receptors. IGFBPs are regulated by prostaglandins, growth factors, hormones, and through important signal transduction pathways. The IGFBPs act classically by sequestering IGFs from interaction with specific cell surface receptors but can also act through IGF-independent intracellular mechanisms that are only now being characterized.

Perturbations in IGF system components have been implicated in a number of diseases. Decreased serum or bone levels of stimulatory IGF system components (IGFs, IGFBP-3, IGFBP-5) or increased levels of inhibitory IGFBPs (IGFBP-4 and IGFBP-6) are associated with diseases of aging including osteoporosis, while bone IGF-II levels are increased in patients with osteoarthritis. Reduction in IGFBP-6 levels is associated with prostate, bone, lung, gastric, and desmoid tumor progression suggesting that IGFBP-6 might be a tumor suppressor gene. In addition, overexpression of IGFBP-6 delivered with an adenoviral vector stimulated apoptosis of NSCLC (lung cancer) and rhabdosarcoma cells but not normal cells. Both IGF-dependent and independent mechanisms were involved. Our laboratory has shown that IGFBP-6 is a potent inhibitor of osteoblast differentiation and have begun to determine the underlying molecular mechanisms involved using the yeast two hybrid system and microarray analysis. We found that IGFBP-6 inhibits osteoblast differentiation at least in part by binding to and sequestering a LIM domain protein called Enigma/Lim Mineralization Protein-1 (LMP-1). LMP-1 acts as a transcriptional coactivator to stimulate type I procollagen gene transcription and has been used as a gene therapeutic to stimulate rapid bone formation in a spinal fusion and fracture repair model. We are identifying the amino acid residues required for the IGFBP-6-LMP-1 interaction to develop small molecule reagents that could be used to interfere with this interaction and stimulate bone formation. In addition, we are exploring the use of siRNA technology to reduce IGFBP-6 expression in osteoporotic bone and at the fracture site to stimulate bone formation.

IGFBP-6 expression is transcriptionally regulated by molecules and signal transduction pathways that are associated with osteoblast differentiation and neoplastic progression. The Wnt signaling pathway and Cyclooxygenase (Cox)-2 modulate IGFBP-6 levels. Cox-2 overexpression and specific prostaglandins inhibit IGFBP-6 expression while non-steroidal anti-inflammatory drugs (NSAIDs, inhibitors of Cox-2) stimulate IGFBP-6 expression. Thus increased IGFBP-6 expression might in part mediate the anti-tumorigenic effect of NSAIDs in some tumors. The Wnt signaling pathway (b-catenin) also suppresses IGFBP-6 expression. Upregulation of the Wnt signaling pathway stimulates osteoblast differentiation and is again associated with tumor progression. Our laboratory is studying regulation of IGFBP-6 expression and cytoplasmic movement by these mechanisms in osteoblasts and cancer cells to discover how IGFBP-6 expression might be altered in disease.

To begin to develop gene therapeutics based on suppressing IGFBP-6 or overexpressing bone forming molecules, our laboratory is actively developing osteoblast specific non-viral vectors that could function as potent delivery vehicles for clinical applications. We are developing transgenic mice to determine the degree of osteoblast specificity of our expression vectors. Gene therapy could be developed to treat fractures, bone diseases such as osteoporosis, and cancer. We are working on new non-viral vector designs to enhance gene delivery in vivo using a combination of cell-specific nuclear entry and Sleeping Beauty Transposon technology.