Glycoamino Acid Analogues of the Thomsen-Friedenreich Tumor-Associated Carbohydrate Antigen: Synthesis and Evaluation of Novel Antiproliferative Factor Glycopeptides.

Glycoamino acid analogues of the Thomsen-Friedenreich antigen disaccharide, where the 4' and 4″ hydroxyl groups were substituted with fluorine or hydrogen, were synthesized and incorporated into the asialylated antiproliferative factor (-APF), a biologically active form of APF, a glycopeptide found in the urine of patients with interstitial cystitis. Various strategies were employed to incorporate the fluorine atom at the 4-positions of either the galactose or -acetylgalactosamine unit of the disaccharide antigen, based on stereochemistry and reactivity. These glycopeptides were evaluated in antiproliferative assays on both primary normal bladder epithelial cells and T24 bladder carcinoma cells.

Death-associated Protein Kinase-1 Expression and Autophagy in Chronic Lymphocytic Leukemia Are Dependent on Activating Transcription Factor-6 and CCAAT/Enhancer-binding Protein-β.

Expression of DAPK1, a critical regulator of autophagy and apoptosis, is lost in a wide variety of tumors, although the mechanisms are unclear. A transcription factor complex consisting of ATF6 (an endoplasmic reticulum-resident factor) and C/EBP-β is required for the IFN-γ-induced expression of DAPK1 IFN-γ-induced proteolytic processing of ATF6 and phosphorylation of C/EBP-β are obligatory for the formation of this transcriptional complex. We report that defects in this pathway fail to control growth of chronic lymphocytic leukemia (CLL).

Varicella-Zoster Virus Vasculopathy: A Case Report Demonstrating Vasculitis using Black-Blood MRI.

Infections are rare but important causes of stroke. Among these, varicella zoster virus has been known to cause ischemic stroke. During an attack of herpes zoster ophthalmicus, it has been hypothesized that the virus replicates in the trigeminal ganglion and travels via the trigeminal nerve centrally to cause cerebral vasculopathy.

Abnormal Akt signalling in bladder epithelial cell explants from patients with interstitial cystitis/bladder pain syndrome can be induced by antiproliferative factor treatment of normal bladder cells.

Objectives: To determine whether protein kinase B (Akt) signalling and secretion of specific downstream effector proteins are abnormal in specific cell fractions of bladder epithelial cells from patients with interstitial cystitis/bladder pain syndrome (IC/BPS), as explanted bladder epithelial cells from patients with IC/BPS produce a frizzled 8-related glycopeptide antiproliferative factor (APF) that inhibits normal bladder epithelial cell proliferation and expression of several proteins known to be regulated by Akt signalling. A related secondary objective was to determine whether treatment of normal bladder epithelial cells with active synthetic asialo-antiproliferative factor (as-APF) induces similar changes in Akt signalling and specific downstream effector proteins/mRNAs.

Patients And Methods: Cell proteins were extracted into four subcellular fractions from primary bladder epithelial explants of six patients who fulfilled modified National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) criteria for IC/BPS and six age- and gender-matched controls.

Evidence for bladder urothelial pathophysiology in functional bladder disorders.

Understanding of the role of urothelium in regulating bladder function is continuing to evolve. While the urothelium is thought to function primarily as a barrier for preventing injurious substances and microorganisms from gaining access to bladder stroma and upper urinary tract, studies indicate it may also function in cell signaling events relating to voiding function. This review highlights urothelial abnormalities in bladder pain syndrome/interstitial cystitis (BPS/IC), feline interstitial cystitis (FIC), and nonneurogenic idiopathic overactive bladder (OAB).

Monoallelic loss of tumor suppressor GRIM-19 promotes tumorigenesis in mice.

Gene-associated with retinoid-interferon induced mortality-19 (GRIM-19), a STAT3-inhibitory protein, was isolated as a growth-suppressive gene product using a genome-wide expression knockdown screen. We and others have shown a loss of expression and occurrence of mutations in the GRIM-19 gene in a variety of primary human cancers, indicating its potential role as tumor suppressor. To help investigate its role in tumor development in vivo, we generated a genetically modified mouse in which Grim-19 can be conditionally inactivated.

Safety of zoster vaccine in elderly adults following documented herpes zoster.

Background: After completion of the Shingles Prevention Study (SPS; Department of Veterans Affairs Cooperative Studies Program Number 403), SPS participants who had initially received placebo were offered investigational zoster vaccine without charge. This provided an opportunity to determine the relative safety of zoster vaccine in older adults following documented herpes zoster (HZ).

Methods: A total of 13 681 SPS placebo recipients who elected to receive zoster vaccine were followed for serious adverse events (SAE) for 28 days after vaccination.

A synthetic form of frizzled 8-associated antiproliferative factor enhances p53 stability through USP2a and MDM2.

Frizzled 8-associated Antiproliferative Factor (APF) is a sialoglycopeptide urinary biomarker of interstitial cystitis/painful bladder syndrome (IC/PBS), a chronic condition of unknown etiology with variable symptoms that generally include pelvic and/or perineal pain, urinary frequency, and urgency. We previously reported that native human APF suppresses the proliferation of normal bladder epithelial cells through a mechanism that involves increased levels of p53. The goal of this study was to delineate the regulatory mechanism whereby p53 expression is regulated by APF.

Integration analysis of quantitative proteomics and transcriptomics data identifies potential targets of frizzled-8 protein-related antiproliferative factor in vivo.

Unlabelled: What's known on the subject? and What does the study add? Interstitial cystitis (IC) is a prevalent and debilitating pelvic disorder generally accompanied by chronic pain combined with chronic urinating problems. Over one million Americans are affected, especially middle-aged women. However, its aetiology or mechanism remains unclear.

Quantitative proteomics identifies a beta-catenin network as an element of the signaling response to Frizzled-8 protein-related antiproliferative factor.

Antiproliferative factor (APF), a Frizzled-8 protein-related sialoglycopeptide involved in the pathogenesis of interstitial cystitis, potently inhibits proliferation of normal urothelial cells as well as certain cancer cells. To elucidate the molecular mechanisms of the growth-inhibitory effect of APF, we performed stable isotope labeling by amino acids in cell culture analysis of T24 bladder cancer cells treated with and without APF. Among over 2000 proteins identified, 54 were significantly up-regulated and 48 were down-regulated by APF treatment.

Antiproliferative factor decreases Akt phosphorylation and alters gene expression via CKAP4 in T24 bladder carcinoma cells.

Background: Urinary bladder cancer is a common malignancy worldwide, and outcomes for patients with advanced bladder cancer remain poor. Antiproliferative factor (APF) is a potent glycopeptide inhibitor of epithelial cell proliferation that was discovered in the urine of patients with interstitial cystitis, a disorder with bladder epithelial thinning and ulceration. APF mediates its antiproliferative activity in primary normal bladder epithelial cells via cytoskeletal associated protein 4 (CKAP4).

An hTERT-immortalized human urothelial cell line that responds to anti-proliferative factor.

Studies of the urothelium, the specialized epithelial lining of the urinary bladder, are critical for understanding diseases affecting the lower urinary tract, including interstitial cystitis, urinary tract infections and cancer. However, our understanding of urothelial pathophysiology has been hampered by a lack of appropriate model systems. Here, we describe the isolation and characterization of a non-transformed urothelial cell line (TRT-HU1), originally explanted from normal tissue and immortalized with hTERT, the catalytic subunit of telomerase.

Structure-Activity Studies on Antiproliferative Factor (APF) Glycooctapeptide Derivatives.

Antiproliferative factor (APF), a sialylated glycopeptide secreted by explanted bladder epithelial cells from interstitial cystitis/painful bladder syndrome (IC/PBS) patients, and its unsialylated analogue (as-APF) significantly decrease proliferation of bladder epithelial cells and/or certain carcinoma cell lines in vitro. We recently reported a structure-activity relationship profile for the peptide portion of as-APF and revealed that truncation of the C-terminal alanine did not significantly affect antiproliferative activity. To better understand the structural basis for the maintenance of activity of this truncated eight amino acid as-APF (as-APF8), we synthesized several amino acid-substituted derivatives and studied their ability to inhibit bladder epithelial cell proliferation in vitro as well as their solution conformations by CD and NMR spectroscopy.

Palmitoylation of cytoskeleton associated protein 4 by DHHC2 regulates antiproliferative factor-mediated signaling.

Previously, we identified cytoskeleton-associated protein 4 (CKAP4) as a major substrate of the palmitoyl acyltransferase, DHHC2, using a novel proteomic method called palmitoyl-cysteine identification, capture and analysis (PICA). CKAP4 is a reversibly palmitoylated and phosphorylated protein that links the ER to the cytoskeleton. It is also a high-affinity receptor for antiproliferative factor (APF), a small sialoglycopeptide secreted from bladder epithelial cells of patients with interstitial cystitis (IC).

Heparin-binding epidermal growth factor-like growth factor functionally antagonizes interstitial cystitis antiproliferative factor via mitogen-activated protein kinase pathway activation.

Objective: To delineate the mechanism underlying the potential functional relationship between interstitial cystitis antiproliferative factor (APF) and heparin-binding epidermal growth factor-like growth factor (HB-EGF), as APF has previously been shown to decrease the proliferation rate of normal bladder epithelial cells and the amount of HB-EGF produced by these cells.

Materials And Methods: APF-responsive T24 transitional carcinoma bladder cells were treated with high-pressure liquid chromatography-purified native APF with or without HB-EGF to determine the involvement of signalling pathways and proliferation by Western blot analysis, p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (Erk)/MAPK assays, and 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) assay.

Results: Cyclic stretch induced the secretion of HB-EGF from T24 cells overexpressing the HB-EGF precursor, resulting in enhanced proliferation.

Structure-activity relationship studies for the peptide portion of the bladder epithelial cell antiproliferative factor from interstitial cystitis patients.

We performed comprehensive structure-activity relationship (SAR) studies on the peptide portion of antiproliferative factor (APF), a sialylated frizzled-8 related glycopeptide that inhibits normal bladder epithelial and urothelial carcinoma cell proliferation. Glycopeptide derivatives were synthesized by solid-phase methods using standard Fmoc chemistry and purified by RP-HPLC; all intermediate and final products were verified by HPLC-MS and NMR analyses. Antiproliferative activity of each derivative was determined by inhibition of (3)H-thymidine incorporation in primary normal human bladder epithelial cells.

Urine markers do not predict biopsy findings or presence of bladder ulcers in interstitial cystitis/painful bladder syndrome.

Purpose: We tested for associations between urine markers, bladder biopsy features and bladder ulcers in interstitial cystitis/painful bladder syndrome.

Materials And Methods: Subjects were 72 patients with interstitial cystitis/painful bladder syndrome undergoing bladder distention and biopsy. Urine was collected before the procedure.

Identification of CKAP4/p63 as a major substrate of the palmitoyl acyltransferase DHHC2, a putative tumor suppressor, using a novel proteomics method.

Protein palmitoylation is the post-translational addition of the 16-carbon fatty acid palmitate to specific cysteine residues by a labile thioester linkage. Palmitoylation is mediated by a family of at least 23 palmitoyl acyltransferases (PATs) characterized by an Asp-His-His-Cys (DHHC) motif. Many palmitoylated proteins have been identified, but PAT-substrate relationships are mostly unknown.

Toxicokinetic study of recombinant human heparin-binding epidermal growth factor-like growth factor (rhHB-EGF) in female Sprague Dawley rats.

Purpose: To determine the toxicity and pharmacokinetics of recombinant heparin-binding epidermal growth factor-like growth factor in female Sprague Dawley rats following intra-bladder and intravenous administration.

Materials And Methods: rhHB-EGF was administered once daily for 6 or 27 days at doses of 3, 10, or 30 microg/kg. 125I-rhHB-EGF was administered on day 7 or 28 for pharmacokinetic analysis.

p53 mediates interstitial cystitis antiproliferative factor (APF)-induced growth inhibition of human urothelial cells.

Antiproliferative factor (APF) is a sialoglycopeptide elevated in the urine of patients with interstitial cystitis, a urinary bladder disorder of unknown etiology that is characterized by chronic pelvic pain. The present study was directed toward uncovering a pathway through which APF signals. Treatment of human urothelial cells with native APF resulted in growth inhibition accompanied by blockade of cell cycle transit and increased p53.

Changes in urine markers and symptoms after bladder distention for interstitial cystitis.

Purpose: We evaluated changes in urine markers and symptom scores after bladder distention in patients with interstitial cystitis.

Materials And Methods: Study subjects were 33 new patients who had undergone no prior interstitial cystitis treatment. Urine specimens were taken before and 1 month after bladder distention.

CKAP4/p63 is a receptor for the frizzled-8 protein-related antiproliferative factor from interstitial cystitis patients.

Antiproliferative factor (APF) is a low molecular weight sialoglycopeptide that is secreted by bladder cells from interstitial cystitis patients and is a potent inhibitor of both normal bladder epithelial and bladder carcinoma cell proliferation. We hypothesized that APF may produce its antiproliferative effects by binding to a transmembrane receptor. This study demonstrates that cytoskeleton-associated protein 4/p63 (CKAP4/p63), a type II transmembrane receptor, binds with high affinity to APF.

Do the National Institute of Diabetes and Digestive and Kidney Diseases cystoscopic criteria associate with other clinical and objective features of interstitial cystitis?

Purpose: We compared urine markers, bladder biopsy findings and clinical features of patients with symptoms of interstitial cystitis (IC) who did or did not meet the cystoscopic criteria defined by the National Institute of Diabetes and Digestive and Kidney Diseases.

Materials And Methods: Urine markers and symptom questionnaires were measured before and 1 month after bladder distention for IC. Bladder biopsies were taken at the time of distention.

An antiproliferative factor from interstitial cystitis patients is a frizzled 8 protein-related sialoglycopeptide.

Approximately 1 million people in the United States suffer from interstitial cystitis, a chronic painful urinary bladder disorder characterized by thinning or ulceration of the bladder epithelial lining; its etiology is unknown. We have identified a glycosylated frizzled-related peptide inhibitor of cell proliferation that is secreted specifically by bladder epithelial cells from patients with this disorder. This antiproliferative factor (APF) profoundly inhibits bladder cell proliferation by means of regulation of cell adhesion protein and growth factor production.

The use of urine proteomic and metabonomic patterns for the diagnosis of interstitial cystitis and bacterial cystitis.

The advent of systems biology approaches that have stemmed from the sequencing of the human genome has led to the search for new methods to diagnose diseases. While much effort has been focused on the identification of disease-specific biomarkers, recent efforts are underway toward the use of proteomic and metabonomic patterns to indicate disease. We have developed and contrasted the use of both proteomic and metabonomic patterns in urine for the detection of interstitial cystitis (IC).