No abstract

R e s e a R c h a R t i c l e3 4 0 7 jci.org Volume 124 Number 8 August 2014 IntroductionThe intestine is made up of repetitive units that consist of a differentiated compartment (the villus) and a proliferative compartment (the crypt). Intestinal stem cells are located in the crypt (1, 2) and produce rapidly proliferating daughter cells, the transit amplifying cells, which subsequently differentiate into 2 main epithelial lineages. The absorptive lineage is composed of all enterocytes, while the secretory lineage is composed of goblet cells (secreting protective mucins), enteroendocrine cells (secreting hormones like serotonin or secretin), and Paneth cells (3, 4). Whether transit amplifying cells differentiate along an absorptive or a secretory lineage is regulated by the Notch pathway (5). Engagement of Notch receptors by Notch ligands, such as Delta or Jagged, induces proteolytic cleavage of the receptor by γ-secretase. The cleaved NOTCH1 receptor (NICD1) translocates into the nucleus, resulting in the formation of an active transcriptional complex composed of RBPJκ (also known as CSL or CBF1) and NICD1. Notch signal activation induces hairy/enhancer of split (Hes) gene expression. Ablation of Notch signaling via genetic deletion of Rbpj results in secretory cell expansion (6). Conversely, in transgenic mice overexpressing NICD1, goblet cells are absent, and the proliferative compartment is expanded (7).The Wnt signaling pathway is a key regulator of intestinal stem cell homeostasis (8), and 2 of the target genes induced by Wnt signaling, c-MYC and c-JUN, encode transcription factors that have oncogenic potential (9, 10). Consequently, aberrant activation of the adenomatous polyposis coli/β-catenin/T cell factor (APC/ β-catenin/TCF) pathway is an initiating event in the majority of human colorectal cancers (11).c-MYC is a transcription factor with key functions in cell differentiation and cancer development (12)(13)(14). In the intestine, c-MYC is required for the altered proliferation and differentiation induced by APC inactivation (15-18). c-MYC is a highly labile protein, and at least 2 ubiquitin ligases, SKP2 and FBW7, can target it for proteasomal degradation (19-21). c-MYC ubiquitination is antagonized by the deubiquitinase USP28, which "piggybacks" on FBW7 and stabilizes c-MYC protein (22). Thus, an E3 ubiquitin ligase and a deubiquitinase, FBW7 and USP28, are together recruited to substrates (22), and a cycle of deubiquitination and ubiquitination controls c-MYC stability.Genomic data from human cancers suggest that most colorectal cancer mutations converge on c-MYC misregulation (23). Due to its key role in tumorigenesis, much recent research has been directed to finding ways to target c-MYC function (24-29). Dominant-negative approaches targeting c-MYC function impair intestinal tumor formation, and c-Myc heterozygous mice show reduced tumor development in the Apc min/+ model (16,17). Transgenic expression of a dominant-negative allele of Myc, OmoMyc, has provided proof of principle that targeting ...

Scaffold modification based on Wang's pioneering MDM2-p53 inhibitors led to novel, chemically stable spiro-oxindole compounds bearing a spiro[3H-indole-3,2'-pyrrolidin]-2(1H)-one scaffold that are not prone to epimerization as observed for the initial spiro[3H-indole-3,3'-pyrrolidin]-2(1H)-one scaffold. Further structure-based optimization inspired by natural product architectures led to a complex fused ring system ideally suited to bind to the MDM2 protein and to interrupt its protein-protein interaction (PPI) with TP53. The compounds are highly selective and show in vivo efficacy in a SJSA-1 xenograft model even when given as a single dose as demonstrated for 4-[(3S,3'S,3'aS,5'R,6'aS)-6-chloro-3'-(3-chloro-2-fluorophenyl)-1'-(cyclopropylmethyl)-2-oxo-1,2,3',3'a,4',5',6',6'a-octahydro-1'H-spiro[indole-3,2'-pyrrolo[3,2-b]pyrrole]-5'-yl]benzoic acid (BI-0252).

ApoER2 and very low density lipoprotein (VLDL) receptor transmit the Reelin signal into target cells of the central nervous system. To a certain extent, both receptors can compensate for each other, and only the loss of both receptors results in the reeler phenotype, which is characterized by a gross defect in the architecture of laminated brain structures. Nevertheless, both receptors also have specific distinct functions, as corroborated by analyses of the subtle phenotypes displayed in mice lacking either ApoER2 or VLDL receptor. The differences in their function(s), however, have not been defined at the cellular level. Here, using a panel of chimeric receptors, we demonstrate that endocytosis of Reelin and the fate of the individual receptors upon stimulation are linked to their specific sorting to raft versus non-raft domains of the plasma membrane. VLDL receptor residing in the non-raft domain endocytoses and destines Reelin for degradation via the clathrin-coated pit/clathrin-coated vesicle/endosome pathway without being degraded to a significant extent. Binding of Reelin to ApoER2, a resident of rafts, leads to the production of specific receptor fragments with specific functions of their own and to degradation of ApoER2 via lysosomes. These features contribute to a receptor-specific fine tuning of the Reelin signal, leading to a novel model that emphasizes negative feedback loops specifically mediated by ApoER2 and VLDL receptor, respectively.