Showing papers by "Philip A. Beachy published in 2011"

Hedgehog/Wnt feedback supports regenerative proliferation of epithelial stem cells in bladder

Abstract: Epithelial integrity in metazoan organs is maintained through the regulated proliferation and differentiation of organ-specific stem and progenitor cells. Although the epithelia of organs such as the intestine regenerate constantly and thus remain continuously proliferative, other organs, such as the mammalian urinary bladder, shift from near-quiescence to a highly proliferative state in response to epithelial injury. The cellular and molecular mechanisms underlying this injury-induced mode of regenerative response are poorly defined. Here we show in mice that the proliferative response to bacterial infection or chemical injury within the bladder is regulated by signal feedback between basal cells of the urothelium and the stromal cells that underlie them. We demonstrate that these basal cells include stem cells capable of regenerating all cell types within the urothelium, and are marked by expression of the secreted protein signal Sonic hedgehog (Shh). On injury, Shh expression in these basal cells increases and elicits increased stromal expression of Wnt protein signals, which in turn stimulate the proliferation of both urothelial and stromal cells. The heightened activity of this signal feedback circuit and the associated increase in cell proliferation appear to be required for restoration of urothelial function and, in the case of bacterial injury, may help clear and prevent further spread of infection. Our findings provide a conceptual framework for injury-induced epithelial regeneration in endodermal organs, and may provide a basis for understanding the roles of signalling pathways in cancer growth and metastasis.

Hedgehog-responsive candidate cell of origin for diffuse intrinsic pontine glioma

Abstract: Diffuse intrinsic pontine gliomas (DIPGs) are highly aggressive tumors of childhood that are almost universally fatal. Our understanding of this devastating cancer is limited by a dearth of available tissue for study and by the lack of a faithful animal model. Intriguingly, DIPGs are restricted to the ventral pons and occur during a narrow window of middle childhood, suggesting dysregulation of a postnatal neurodevelopmental process. Here, we report the identification of a previously undescribed population of immunophenotypic neural precursor cells in the human and murine brainstem whose temporal and spatial distributions correlate closely with the incidence of DIPG and highlight a candidate cell of origin. Using early postmortem DIPG tumor tissue, we have established in vitro and xenograft models and find that the Hedgehog (Hh) signaling pathway implicated in many developmental and oncogenic processes is active in DIPG tumor cells. Modulation of Hh pathway activity has functional consequences for DIPG self-renewal capacity in neurosphere culture. The Hh pathway also appears to be active in normal ventral pontine precursor-like cells of the mouse, and unregulated pathway activity results in hypertrophy of the ventral pons. Together, these findings provide a foundation for understanding the cellular and molecular origins of DIPG, and suggest that the Hh pathway represents a potential therapeutic target in this devastating pediatric tumor.

Structure of the protein core of the glypican Dally-like and localization of a region important for hedgehog signaling

Abstract: Glypicans are heparan sulfate proteoglycans that modulate the signaling of multiple growth factors active during animal development, and loss of glypican function is associated with widespread developmental abnormalities. Glypicans consist of a conserved, approximately 45-kDa N-terminal protein core region followed by a stalk region that is tethered to the cell membrane by a glycosyl-phosphatidylinositol anchor. The stalk regions are predicted to be random coil but contain a variable number of attachment sites for heparan sulfate chains. Both the N-terminal protein core and the heparan sulfate attachments are important for glypican function. We report here the 2.4-A crystal structure of the N-terminal protein core region of the Drosophila glypican Dally-like (Dlp). This structure reveals an elongated, α-helical fold for glypican core regions that does not appear homologous to any known structure. The Dlp core protein is required for normal responsiveness to Hedgehog (Hh) signals, and we identify a localized region on the Dlp surface important for mediating its function in Hh signaling. Purified Dlp protein core does not, however, interact appreciably with either Hh or an Hh:Ihog complex.

Hedgehogs, Flies, Wnts and MYCs: The Time Has Come for Many Things in Medulloblastoma

Abstract: Medulloblastoma epitomizes the war against childhood cancer. Although this small round blue-cell tumor accounts for only 20% of all childhood brain tumors, or just fewer than 600 new cases each year in the United States, medulloblastoma has nevertheless become a paradigm for biologic study, clinical investigation, and assessment of late effects. Craniospinal irradiation historically became the mainstay of therapy for this WHO grade 4 neoplasm because of its propensity to disseminate throughout the neuraxis via the subarachnoid space. By the 1970s, 36 Gy of craniospinal radiotherapy resulted in 5-year survival of greater than 50% for better-risk patients. With cooperative group trials in the 1980s, adjuvant chemotherapy was introduced to medulloblastoma treatment protocols (SIOP I [International Society of Paediatric Oncology], CCG 942 [Children’s Cancer Group], and POG 7909 [Pediatric Oncology Group]), leading to improved survival and subsequent reduction of radiotherapy dosage for averagerisk patients in the 1990s and early part of the last decade. A decrease in craniospinal radiotherapy from 36 Gy to 23.4 Gy resulted in better neurocognitive and physical outcomes for average-risk patients, highlighting the importance of developing appropriately riskadapted therapy. Although prognosis has improved over many years, a significant fraction of children still do not respond to standard therapy. Perhaps equally disturbing, survivors are left with a multitude of sequelae attributable to chemotherapy and radiation therapy, including cognitive impairment, hearing loss, neuropathies, seizures, endocrinopathies, strokes, and second malignancies. Thus, there is an overwhelming need to refine strategies that will minimize unnecessarily aggressive therapy for patients with low-risk disease and maximize efficacious treatment for patients with high-risk disease. Such refinement can come only with better disease or risk classification. In an era of genomic profiling, risk stratification for medulloblastoma disappointingly remains strictly clinical, based solely on age, resection extent, Chang metastasis staging, and more recently anaplasia. Since the mid 1990s, a two-tiered risk stratification system has been employed for the purposes of North American cooperative group trials. Patients older than 3 years with less than 1.5 cm of residual tumor and nonmetastatic disease are considered average risk, whereas those younger than 3 years with more than 1.5 cm of residual tumor, evidence of tumor dissemination, or diffuse anaplasia are considered high risk. Multiple histopathologic, cytogenetic, and molecular factors have been found to influence prognosis, but to date, none have been incorporated into the risk-stratification system. Histopathologically, the desmoplastic variant has long been recognized as connoting better prognosis, whereas the large-cell anaplastic variant more recently has been found to be associated with reduced survival. Moreover, isochromosome 17q, loss of heterozygosity at chromosome 17p, and high-level chromosomal gain at 8q24 (locus of c-MYC) have all been associated with large-cell anaplastic variant medulloblastoma and poor prognosis. Monosomy 6 has been associated with favorable prognosis. In addition, a number of molecular markers have been identified over the past decade and have laid the groundwork for a risk-stratification system with a biologic basis. Favorable prognosis has been described for children with high TrkC expression or nucleopositivity of -catenin, an indicator of activated Wnt signaling—a pathway now known to affect embryogenesis and cancer. Poor prognosis is indicated by overexpression of calbindin-D, elevated ErbB2 expression, and elevated expression of c-MYC, a member of the MYC family of transcription factors that stimulates cell proliferation. In recent years, large-scale genomic and gene expression profile studies have begun to parse the molecular heterogeneity of medulloblastomas into clear subtypes with associated risk stratification. Initial gene expression microarray studies used a supervised approach, grouping samples on the basis of histology, metastatic status, and survival to identify differentially expressed genes in these clinical and histologic groups. More recently, unsupervised microarray studies in medulloblastoma by Kool et al (62 samples) and Thompson et al (46 samples) each identified up to five subgroups of medulloblastoma, including a subgroup characterized by activation of the Wnt signaling pathway, associated with excellent prognosis, and a subgroup characterized by activation of the Hedgehog (Hh) signaling pathway occurring primarily in infants and adults and associated with desmoplastic histology. The Hh signaling pathway is involved in organogenesis, limb formation, brain patterning, and multiple forms of cancer. There are several Hh family ligands, the best known of which is Sonic Hedgehog (SHh) for its multiple developmental roles, including in cerebellar development. The study by Kool et al also identified a molecular subtype associated with a high rate of metastatic disease, anaplastic/ large-cell histology, and poor prognosis. In this issue of Journal of Clinical Oncology, four separate studies dovetail with previous work to crystallize a molecular classification of medulloblastoma and codify a new risk-stratification model on the basis of the molecular biology of these disease subtypes. Ellison et al JOURNAL OF CLINICAL ONCOLOGY E D I T O R I A L S VOLUME 29 NUMBER 11 APRIL 1

Method of detection using antibodies that specifically bind hedgehog-derived polypeptides

Abstract: The present invention provides two novel polypeptides, referred to as the “N” and “C” fragments of hedgehog, or N-terminal and C-terminal fragments, respectively, which are derived after specific cleavage at a G′CF site recognized by the autoproteolytic domain in the native protein. Also included are sterol-modified hedgehog polypeptides and functional fragments thereof. Methods of identifying compositions which affect hedgehog activity based on inhibition of cholesterol modification of hedgehog protein are described.