SOST and DKK: Antagonists of LRP Family Signaling as Targets for Treating Bone Disease

The study of rare human genetic disorders has often led to some of the most significant advances in biomedical research. One such example was the body of work that resulted in the identification of the Low Density Lipoprotein-Related Protein (LRP5) as a key regulator of bone mass. Point mutations were identified that encoded forms of LRP5 associated with very high bone mass (HBM). HBM patients live to a normal age and do not appear to have increased susceptibility to carcinogenesis or other disease. Thus, devising methods to mimic the molecular consequences of this mutation to treat bone diseases associated with low bone mass is a promising avenue to pursue. Two groups of agents related to putative LRP5/6 functions are under development. One group, the focus of this paper, is based on antagonizing the functions of putative inhibitors of Wnt signaling, Dickkopf-1 (DKK1), and Sclerostin (SOST). Another group of reagents under development is based on the observation that LRP5 may function to control bone mass by regulating the secretion of serotonin from the enterrochromaffin cells of the duodenum

Mouse Models of Prostate Cancer

The development and optimization of high-throughput screening methods has identified a multitude of genetic changes associated with human disease. The use of immunodeficient and genetically engineered mouse models that mimic the human disease has been crucial in validating the importance of these genetic pathways in prostate cancer. These models provide a platform for finding novel therapies to treat human patients afflicted with prostate cancer as well as those who have debilitating bone metastases. In this paper, we focus on the historical development and phenotypic descriptions of mouse models used to study prostate cancer. We also comment on how closely each model recapitulates human prostate cancer

Analysis of the role of the retinoblastoma susceptibility gene in murine development and tumorigenesis

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 1996.Includes bibliographical references.by Bart O. Williams.Ph.D

Where Wnts Went: The Exploding Field of Lrp5 and Lrp6 Signaling in Bone

Wnt signaling has emerged as a central regulator of skeletal modeling and remodeling. Loss- or gain-of-function mutations in two Wnt co-receptors, Lrp5 and (more recently) Lrp6, have drawn attention to the importance of the Wnt pathway in bone biology. This review summarizes our current understanding of how the Wnt pathway operates on bone and the implications this has for skeletal physiology and drug discovery. Over the past 9 yr, rapid advances have been made in our understanding of the cellular targets for Wnt signaling and of the important regulatory molecules in this metabolic pathway. Both canonical and noncanonical signaling pathways seem to be important for mediating the effects of Wnt in bone. A rapidly expanding catalog of genetically engineered mice has been used to establish the importance of downstream effector molecules (such as β-catenin) in the Wnt pathway, as well as the critical role of endogenous inhibitors of Wnt signaling (such as Dkk1 and sclerostin) in bone metabolism. Indeed, regulation of sclerostin in osteocytes is emerging as an important final pathway for regulating bone anabolism in response to diverse trophic stimuli, from mechnotransduction to the anabolic actions of PTH. From the outset, it had been assumed that the effects of Wnt signaling in bone were caused by direct actions in osteoblast precursors, osteoblasts, and osteocytes. However, startling recent findings have challenged this view and suggest that a key target, at least in mice, is the duodenal enterochromaffin cell. There, Wnt signaling transduced by Lrp5 regulates serotonin synthesis, which acts in an endocrine fashion to regulate bone cell metabolism. It will take time to reconcile this new information with the considerable body of information we already have regarding the actions of Wnt in bone. The Wnt pathway has rapidly emerged as a therapeutic target for drug discovery. Neutralizing antibodies and small-molecule inhibitors of endogenous Wnt inhibitors have shown early promise as bone anabolic agents. However, given the central role of the Wnt pathway in regulating growth and development in extraskeletal tissues, as well as our still rudimentary understanding of how this signaling cascade actually affects bone metabolism, considerable work will be needed to ensure the safety of these new therapies

Biomechanical Evaluation of fracture Fixation Constructs using a Variable-angle Locked Periprosthetic Femur Plate System

BackgroundIn the United States there are more than 230,000 total hip replacements annually, and periprosthetic femoral fractures occur in 0.1–4.5% of those patients. The majority of these fractures occur at the tip of the stem (Vancouver type B1). The purpose of this study was to compare the biomechanically stability and strength of three fixation constructs and identify the most desirable construct.MethodsFifteen medium adult synthetic femurs were implanted with a hip prosthesis and were osteotomized in an oblique plane at the level of the implant tip to simulate a Vancouver type B1 periprosthetic fracture. Fractures were fixed with a non-contact bridging periprosthetic proximal femur plate (Zimmer, Inc., Warsaw, IN). Three proximal fixation methods were used: Group 1, bicortical screws; Group 2, unicortical screws and one cerclage cable; and Group 3, three cerclage cables. Distally, all groups had bicortical screws. Biomechanical testing was performed using an axial-torsional testing machine in three different loading modalities (axial compression, lateral bending, and torsional/sagittal bending), next in axial cyclic loading to 10,000 cycles, again in the three loading modalities, and finally to failure in torsional/sagittal bending.ResultsGroup 1 had significantly greater load to failure and was significantly stiffer in torsional/sagittal bending than Groups 2 and 3. After cyclic loading, Group 2 had significantly greater axial stiffness than Groups 1 and 3. There was no difference between the three groups in lateral bending stiffness. The average energy absorbed during cyclic loading was significantly lower in Group 2 than in Groups 1 and 3.ConclusionsBicortical screw placement achieved the highest load to failure and the highest torsional/sagittal bending stiffness. Additional unicortical screws improved axial stiffness when using cable fixation. Lateral bending was not influenced by differences in proximal fixation.Clinical RelevanceTo treat periprosthetic fractures, bicortical screw placement should be attempted to maximize load to failure and torsional/sagittal bending stiffness

A critical role for Apc in hematopoietic stem and progenitor cell survival

The adenomatous polyposis coli (Apc) tumor suppressor is involved in the initiation and progression of colorectal cancer via regulation of the Wnt signaling cascade. In addition, Apc plays an important role in multiple cellular functions, including cell migration and adhesion, spindle assembly, and chromosome segregation. However, its role during adult hematopoiesis is unknown. We show that conditional inactivation of Apc in vivo dramatically increases apoptosis and enhances cell cycle entry of hematopoietic stem cells (HSCs)/ hematopoietic progenitor cells (HPCs), leading to their rapid disappearance and bone marrow failure. The defect in HSCs/HPCs caused by Apc ablation is cell autonomous. In addition, we found that loss of Apc leads to exhaustion of the myeloid progenitor pool (common myeloid progenitor, granulocyte-monocyte progenitor, and megakaryocyte-erythroid progenitor), as well as the lymphoid-primed multipotent progenitor pool. Down-regulation of the genes encoding Cdkn1a, Cdkn1b, and Mcl1 occurs after acute Apc excision in candidate HSC populations. Together, our data demonstrate that Apc is essential for HSC and HPC maintenance and survival

Accelerated and increased joint damage in young mice with global inactivation of mitogen-inducible gene 6 after ligament and meniscus injury

Abstract Introduction Ligament and meniscal damage can cause joint disease. Arthritic joints contain increased amounts of epidermal growth factor receptor (EGFR) protein, and polymorphisms in EGFR are associated with arthritis risk. The role of endogenous EGFR regulation during joint disease due to ligament and meniscal trauma is unknown. Mitogen-inducible gene 6 (MIG-6) can reduce EGFR phosphorylation and downstream signaling. We examined the effect of EGFR modulation by MIG-6 on joint disease development after ligament and meniscus injury. Methods Knee ligament transection and meniscus removal were performed surgically on mice homozygous for a global inactivating mutation in MIG-6 (Mig-6 −/− ) and in wild-type (WT) animals. Results Two weeks after surgery, Mig-6 −/− mice had bone erosion as well as greater fibrous tissue area and serum RANKL concentration than WT mice. Four weeks after surgery, Mig-6 −/− mice had less cartilage and increased cell proliferation relative to contralateral control and WT knees. Increased apoptotic cells and growth outside the articulating region occurred in Mig-6 −/− mice. Tibia trabecular bone mineral density (BMD) and the number of trabeculae were lower in surgically treated knees relative to the respective control knees for both groups. BMD, as well as trabecular thickness and number, were lower in surgically treated knees from Mig-6 −/− mice relative to WT surgically treated knees. Phosphorylated EGFR staining in surgically treated knees decreased for WT mice and increased for Mig-6 −/− mice. Fewer inflammatory cells were present in the knees of WT mice. Conclusion Mig-6 −/− mice have rapid and increased joint damage after ligament and meniscal trauma. Mig-6 modification could lessen degenerative disease development after this type of injury.http://deepblue.lib.umich.edu/bitstream/2027.42/109507/1/13075_2013_Article_4169.pd

Mice with a Heterozygous Lrp6 Deletion Have Impaired Fracture Healing

Bone fracture non-unions, the failure of a fracture to heal, occur in 10%–20% of fractures and are a costly and debilitating clinical problem. The Wnt/β-catenin pathway is critical in bone development and fracture healing. Polymorphisms of linking low-density lipoprotein receptor-related protein 6 (LRP6), a Wnt-binding receptor, have been associated with decreased bone mineral density and fragility fractures, although this remains controversial. Mice with a homozygous deletion of Lrp6 have severe skeletal abnormalities and are not viable, whereas mice with a heterozygous deletion have a combinatory effect with Lrp5 to decrease bone mineral density. As fracture healing closely models embryonic skeletal development, we investigated the process of fracture healing in mice heterozygous for Lrp6 (Lrp6+/−) and hypothesized that the heterozygous deletion of Lrp6 would impair fracture healing. Mid-diaphyseal femur fractures were induced in Lrp6+/− mice and wild-type controls (Lrp6+/+). Fractures were analyzed using micro-computed tomography (μCT) scans, biomechanical testing, and histological analysis. Lrp6+/− mice had significantly decreased stiffness and strength at 28 days post fracture (PF) and significantly decreased BV/TV, total density, immature bone density, and mature area within the callus on day-14 and -21 PF; they had significantly increased empty callus area at days 14 and 21 PF. Our results demonstrate that the heterozygous deletion of Lrp6 impairs fracture healing, which suggests that Lrp6 has a role in fracture healing

The Wnt Co-Receptor Lrp6 Is Required for Normal Mouse Mammary Gland Development

Canonical Wnt signals are transduced through a Frizzled receptor and either the LRP5 or LRP6 co-receptor; such signals play central roles during development and in disease. We have previously shown that Lrp5 is required for ductal stem cell activity and that loss of Lrp5 delays normal mammary development and Wnt1-induced tumorigenesis. Here we show that canonical Wnt signals through the Lrp6 co-receptor are also required for normal mouse mammary gland development. Loss of Lrp6 compromises Wnt/β-catenin signaling and interferes with mammary placode, fat pad, and branching development during embryogenesis. Heterozygosity for an inactivating mutation in Lrp6 is associated with a reduced number of terminal end buds and branches during postnatal development. While Lrp6 is expressed in both the basal and luminal mammary epithelium during embryogenesis, Lrp6 expression later becomes restricted to cells residing in the basal epithelial layer. Interestingly, these cells also express mammary stem cell markers. In humans, increased Lrp6 expression is associated with basal-like breast cancer. Taken together, our results suggest both overlapping and specific functions for Lrp5 and Lrp6 in the mammary gland

Lrp5 and Lrp6 exert overlapping functions in osteoblasts during postnatal bone acquisition

The canonical Wnt signaling pathway is critical for skeletal development and maintenance, but the precise roles of the individual Wnt co-receptors, Lrp5 and Lrp6, that enable Wnt signals to be transmitted in osteoblasts remain controversial. In these studies, we used Cre-loxP recombination, in which Cre-expression is driven by the human osteocalcin promoter, to determine the individual contributions of Lrp5 and Lrp6 in postnatal bone acquisition and osteoblast function. Mice selectively lacking either Lrp5 or Lrp6 in mature osteoblasts were born at the expected Mendelian frequency but demonstrated significant reductions in whole-body bone mineral density. Bone architecture measured by microCT revealed that Lrp6 mutant mice failed to accumulate normal amounts of trabecular bone. By contrast, Lrp5 mutants had normal trabecular bone volume at 8 weeks of age, but with age, these mice also exhibited trabecular bone loss. Both mutants also exhibited significant alterations in cortical bone structure. In vitro differentiation was impaired in both Lrp5 and Lrp6 null osteoblasts as indexed by alkaline phosphatase and Alizarin red staining, but the defect was more pronounced in Lrp6 mutant cells. Mice lacking both Wnt co-receptors developed severe osteopenia similar to that observed previously in mice lacking β-catenin in osteoblasts. Likewise, calvarial cells doubly deficient for Lrp5 and Lrp6 failed to form osteoblasts when cultured in osteogenic media, but instead attained a chondrocyte-like phenotype. These results indicate that expression of both Lrp5 and Lrp6 are required within mature osteoblasts for normal postnatal bone development