The Hedgehog pathway is one fundamental means of controlling mammalian cell behavior and is used to regulate a wide variety of disparate biological events including tissue patterning, stem cell renewal, and cell proliferation. All Hedgehog signaling relies on the proto-oncogene Smoothened, mutations in which can cause basal cell carcinoma, the most common cancer in North America, and medulloblastoma, the most common solid cancer among children. Tumors have displayed resistance to Smoothened antagonists currently in clinical trials, indicating a need for other inhibitors of the Hedgehog pathway. We have discovered that mammalian Hedgehog signals move Smoothened to an organelle called the primary cilium, and that this movement is necessary for pathway activation. Although it is known that almost all mammalian cells possess a single primary cilium that extends into the extracellular environment, the functions of this organelle are poorly understood. We hypothesize that the primary cilium acts as a cellular antenna, through which Hedgehog signals are transduced. Despite their importance to both development and disease, the molecular mechanisms by which the cilium is generated and by which Smoothened functions remain unclear. To address these gaps in our understanding, we seek to conduct a high-throughput screen to identify novel compounds that can inhibit cilium formation or Smoothened movement to cilia. Using a combination of high-throughput microscopy and high-content analysis, we will visualize and quantitate Smoothened localization to the cilium in cultured cells. We will subject novel compounds that inhibit Smoothened localization to the cilium to secondary assays that reveal whether the compound affects cilium biogenesis, Smoothened trafficking, and Hedgehog pathway activity, among other parameters. This work will produce the chemical tools to pharamacogenetically dissect the molecular mechanisms of ciliogenesis and Smoothened regulation. Additionally, given the central role misregulated Hedgehog signaling plays in diverse human cancers, these compounds may provide the bases for the development of novel chemotherapeutic drugs.
Problems in cell-cell communication can underlie diseases as diverse as birth defects and cancer. One mechanism of cell-cell communication uses Hedgehog proteins, secreted signaling proteins. Defects in Hedgehog signaling can cause birth defects, such as holoprosencephaly, and several types of cancer, including medulloblastoma. Recent evidence has revealed that Hedgehog signaling relies on the primary cilium, a poorly understood organelle present on most cells. To be able to better understand how Hedgehog signaling and the primary cilium function, we propose to conduct a screen for small molecules that perturb these processes. These chemicals may provide the basis for the development of novel therapies for Hedgehog-related cancers.
|Wu, Victoria M; Chen, Steven C; Arkin, Michelle R et al. (2012) Small molecule inhibitors of Smoothened ciliary localization and ciliogenesis. Proc Natl Acad Sci U S A 109:13644-9|