Mutations in foregut SOX2+ cells induce efficient proliferation via CXCR2 pathway

Identification of the precise molecular pathways involved in oncogene-induced transformation may help us gain a better understanding of tumor initiation and promotion. Here, we demonstrate that SOX2⁺ foregut epithelial cells are prone to oncogenic transformation upon mutagenic insults, such as Kras^G12D and p53 deletion. GFP-based lineage-tracing experiments indicate that SOX2⁺ cells are the cells-of-origin of esophagus and stomach hyperplasia. Our observations indicate distinct roles for oncogenic KRAS mutation and P53 deletion. p53 homozygous deletion is required for the acquisition of an invasive potential, and Kras^G12D expression, but not p53 deletion, suffices for tumor formation. Global gene expression analysis reveals secreting factors upregulated in the hyperplasia induced by oncogenic KRAS and highlights a crucial role for the CXCR2 pathway in driving hyperplasia. Collectively, the array of genetic models presented here demonstrate that stratified epithelial cells are susceptible to oncogenic insults, which may lead to a better understanding of tumor initiation and aid in the design of new cancer therapeutics.     

Loss of Core 1-derived O-Glycans Decreases Breast Cancer Development in Mice

Mucin-type core 1-derived O-glycans, one of the major types of O-glycans, are highly expressed in mammary gland epithelium. Abnormal O-glycans such as Tn antigen are found in over 90% of breast cancers; however, the in vivo role of these aberrant O-glycans in the etiology of breast cancer is unclear. We generated mice with mammary epithelial specific deletion of core 1-derived O-glycans. By crossing with two spontaneous mouse breast cancer models, we determined that loss of core 1-derived O-glycans delays the onset and progression of breast cancer development. Deficiency of core 1 O-glycosylation impaired the localization of Muc1, a major O-glycoprotein, on the apical surfaces of mammary epithelium. Signaling mediated by Muc1, which is critical for breast cancer development, was also defective in the absence of core 1 O-glycans. This study reveals an unexpected role of core 1-derived O-glycans in breast cancer development in mice.     

Vascular-homing peptides for targeted drug delivery and molecular imaging: Meeting the clinical challenges

The vasculature of each organ expresses distinct molecular signatures critically influenced by the pathological status. The heterogeneous profile of the vascular beds has been successfully unveiled by the in vivo phage display, a high-throughput tool for mapping normal, diseased, and tumor vasculature. Specific challenges of this growing field are targeted therapies against cancer and cardiovascular diseases, as well as novel bioimaging diagnostic tools. Tumor vasculature-homing peptides have been extensively evaluated in several preclinical and clinical studies both as targeted-therapy and diagnosis. To date, results from several Phase I and II trials have been reported and many other trials are currently ongoing or recruiting patients. In this review, advances in the identification of novel peptide ligands and their corresponding receptors on tumor endothelium through the in vivo phage display technology are discussed. Emphasis is given to recent findings in the clinical setting of vascular-homing peptides selected by in vivo phage display for the treatment of advanced malignancies and their altered vascular beds.     

A Novel Enhanced Green FluorescentProtein-Expressing NOG Mouse for Analyzingthe Microenvironment of Xenograft Tissues

The interaction between transplanted cells and host tissues is important for the growthand maintenance of transplanted cells. To analyze the mechanisms of these interactions, asystemic fluorescent protein-expressing mouse is a useful recipient. In this study, wegenerated a novel NOG strain, which strongly expresses enhanced green fluorescent protein(EGFP; PgkEGFP-NOG), especially in the liver, kidney, gastrointestinal tract, and testis.Because the host tissues expressed EGFP, xenotransplanted human cancer cells were clearlyidentified as EGFP-negative colonies in PgkEGFP-NOG mice. Immunohistochemical analysisrevealed that EGFP-expressing stromal tissues formed a complicated tumor microenvironmentwithin xenograft tissues. Moreover, a similar microenvironment was observed in human iPScell-derived teratomas. Collectively, these results indicated that a suitablemicroenvironment is essential for the growth and maintenance of xenotransplanted cells andthat PgkEGFP-NOG mice represent a useful animal model for analyzing the mechanisms ofmicroenvironment formation.     

Sindbis virus replication, is insensitive to rapamycin and torin1, and suppresses Akt/mTOR pathway late during infection in HEK cells

Genetically engineered Sindbis viruses (SIN) are excellent oncolytic agents in preclinical models. Several human cancers have aberrant Akt signaling, and kinase inhibitors including rapamycin are currently tested in combination therapies with oncolytic viruses. Therefore, it was of interest to delineate possible cross-regulation between SIN replication and PI3K/Akt/mTOR signaling. Here, using HEK293T cells as host, we report the following key findings: (a) robust SIN replication occurs in the presence of mTOR specific inhibitors, rapamycin and torin1 or Ly294002 – a PI3K inhibitor, suggesting a lack of requirement for PI3K/Akt/mTOR signaling; (b) suppression of phosphorylation of Akt, mTOR and its effectors S6, and 4E-BP1 occurs late during SIN infection: a viral function that may be beneficial in counteracting cellular drug resistance to kinase inhibitors; (c) Ly294002 and SIN act additively to suppress PI3K/Akt/mTOR pathway with little effect on virus release; and (d) SIN replication induces host translational shut off, phosphorylation of eIF2α and apoptosis. This first report on the potent inhibition of Akt/mTOR signaling by SIN replication, bolsters further studies on the development and evaluation of engineered SIN genotypes in vitro and in vivo for unique cytolytic functions.     

Proteomic analysis of reaper 5' untranslated region-interacting factors isolated by tobramycin affinity-selection reveals a role for La antigen in reaper mRNA translation

Translational control is a key step in gene expression regulation during apoptosis. To understand the mechanisms of mRNA translation of a pro-apoptotic gene, reaper (rpr), we adapted the tobramycin-aptamer technique described by Hartmuth et al. (Proc. Natl. Acad. Sci. USA 2002, 99, 16719-16724) for the analysis of proteins interacting with rpr 5' untranslated region (UTR). We assembled ribonucleoprotein complexes in vitro using translation extracts derived from Drosophila embryos and purified the RNA-protein complexes for mas spectrometry analysis. We identified the proteins bound to the 5' UTR of rpr. One of them, the La antigen, was validated by RNA-crosslinking experiments using recombinant protein and by the translation efficiency of reporter mRNAs in Drosophila cells after RNAinterference experiments. Our data provide evidence of the involvement of La antigen in the translation of rpr and set a protocol for purification of tagged-RNA-protein complexes from cytoplasmic extracts.