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Background: Effective patient–provider communication enables shared decision-making and leads to improved patient satisfaction, treatment adherence and health outcomes. The ideal time for d...

The deubiquitylating enzyme USP18 is a major negative regulator of the interferon (IFN) signalling cascade. IFN pathways contribute to resistance to conventional chemotherapy, radiotherapy, and immunotherapy and are often deregulated in cancer. USP18 is the predominant human protease that cleaves interferon-stimulated gene ISG15, a ubiquitin-like protein tightly regulated in the context of innate immunity, from its modified substrate proteins in vivo. In this study, using advanced proteomic techniques, we have expanded the USP18-dependent ISGylome and proteome in a chronic myeloid leukaemia (CML)-derived cell line (HAP1) treated with type I IFN. Novel ISGylation targets were characterised that modulate the sensing of innate ligands, antigen presentation and secretion of cytokines. Consequently, CML USP18-deficient cells are more antigenic, driving increased activation of cytotoxic T lymphocytes (CTLs) and are more susceptible to irradiation. Our results suggest USP18 as a pharmacological target in cancer immunotherapy and radiotherapy.

The primary goal of this study was to examine attachment style effects on self-disclosure, trust, satisfaction, and uncertainty levels of individuals in long-distance romantic relationships (LDRRs) who utilise technologically mediated communication channels as a primary means of daily interaction with their romantic partners. Participants (N = 211) from 27 countries who maintained committed romantic relationships (M = 3 years) with geographical separation (M = 1042 miles, Mdn = 160 miles) were surveyed. Using structural equation modelling (SEM), 7 of the 8 hypotheses were supported, showing that attachment style and self-disclosure predict relational uncertainty, trust, and relational satisfaction. In examining the distinctions between attachment styles (i.e., secure, preoccupied, fearful-avoidant, and dismissive) three models were developed and confirmed with good fit. The implications of the study suggest how both attachment style and self-disclosure influence the overall success of LDRRs.

The primary goal of this study was to establish a model of privacy control, stemming from Communication Privacy Management (CPM) theory, between the parent and the emerging adult child (N = 851) to predict the likelihood to reveal private information. Using Structural Equation Modeling (SEM), the study confirmed that the CPM criteria variables (culture, gender, context, motivation, and risk–benefit ratio) exert an influence on revealing private information. Three psychosocial outcome variables were also examined: rumination, stress, and well-being. Specifically, emerging adults reporting a communicatively open family relational culture, low levels of relational quality, and low levels of relational risk were more likely to reveal private information to their parents, while personal characteristics, perceived reciprocity, characteristics of the confidant, and stigma risks did not significantly affect the likelihood to reveal private information. The implications of the study suggest that predictors of priv...

Introduction In tertiary centres both EMR and ESD sometimes in combination with hybrid techniques are used in the treatment of polpys>20 mm. EMR allows for removal of polyps confined to the mucosa often in a piecemeal fashion while ESD aims for enbloc resection for lesions extending into the submucosa. The most serious complications include bleeding and perforation. Over a 3 year period at a tertiary centre we aimed to assess complication and recurrence rates between these two modalities. Method Data was collected retrospectively collected over a 3 year period. All EMR/ESD were performed by three dedicated consultant endoscopists with over 10 years experience in performing level III and IV endoscopic resection. The endoscopy Unisoft database was interrogated to select all patients having EMR/ESD including screen-detected lesions. Lesions Results 128 polyps were removed endoscopically from 125 patients (80 males) 64%. Mean age was 70 and mean polyp size 38 mm (range 20–101 mm). For low-grade dysplatic lesions histology confirmed 83 (64%) tubulo-villous adenomas (TVA) and 21 (16%) Tubulo-adenomas (TA). There were 11 (8.5%) adenocarcinomas, 7 (5.4%) and high-grade dysplastic lesions. All adenocarcinomas were eventually referred for surgical management. One large high-grade dysplastic lesion was referred for Trans-anal microscopic surgery (TEMS) after submucosal fibrosis and bleeding encountered during ESD. There were 6 (4.6%) inflammatory/hyperplastic lesions. Procedural bleeding was encountered in six patients, one patient had delayed bleeding. One patient had post-polypectomy pain syndrome. There were no perforations. No difference in complication rates was found between EMR and ESD. 12 month recurrence rate was 15/113 (13%). This was related to increasing size of polyps. All recurrences were managed endoscopically. LOS was between 0–24 hours in over 97% of patients. Conclusion In high volume centres EMR/ESD is safe with low complication and recurrence rates. Disclosure of Interest None Declared

This study explored how participants discursively rendered voluntary kin relationships sensical and legitimate. Interpretive analyses of 110 interviews revealed four main types of voluntary kin: (i) substitute family, (ii) supplemental family, (iii) convenience family, and (iv) extended family. These types were rendered sensical and legitimated by drawing on the discourse of the traditional family. Except for the extended family, three of four voluntary kin family types were justified by an attributed deficit in the blood and legal family. Because voluntary kin relationships are not based on the traditional criteria of association by blood or law, members experience them as potentially challenging, requiring discursive work to render them sensical and legitimate to others.

BACKGROUND AND STUDY AIMS It is well recognized that myocardial ischemia can occur during endoscopic retrograde cholangiopancreatography (ERCP). Acute arrhythmias and ST segment changes have been reported by a number of authors, but the longer-term sequelae with regard to permanent myocardial damage are not known. The aim of this study was to determine the presence or absence of significant injury to the heart muscle. PATIENTS AND METHODS Sixty-two patients undergoing therapeutic ERCP were assessed clinically and with electrocardiography (ECG) for the presence of ischemic heart disease before the procedure. Extensive intraprocedural monitoring was carried out, postprocedural ECGs were recorded, and serum troponin T levels were measured. The ECGs were evaluated blindly by a single cardiologist. RESULTS In 61 of the 62 patients, no changes were observed between the ECGs before and after the procedure. One patient had postprocedural T wave inversion but a normal troponin T level, excluding myocardial damage. One patient with mild renal insufficiency and treated heart failure had borderline troponin T elevation (0.05 microg/l) but no ECG changes. No complications of ERCP occurred. CONCLUSIONS Whilst ECG and rhythm changes indicating transient myocardial ischemia do occur during ERCP, there is no evidence that myocardial damage takes place as a consequence of this.

The first decade of the new millennium saw an upsurge in global financing for health. When the world took stock of progress on the Millennium Development Goals in mid-2010 the one addressing maternal health showed the least progress. Did maternal health miss the boat? In mid-2010 the Secretary-General of the United Nations launched a “Global Strategy for Women’s and Children’s Health”, also known as the “Every Woman Every Child” initiative. Has the tide now turned in favour of maternal health? The authors try to answer this question by first examining whether maternal health really missed out with respect to increased global funding and why this may have occurred. They then assess whether the new initiative will make a difference by comparing several elements of the approach taken by HIV/AIDS activist to that of maternal health activists. They suggest that real progress requires international financing, thus pledges must become robust and reliable commitments. They conclude that the absence of an organisational structure in the current initiative means the global maternal health financing revolution will probably not happen.

We have constructed clones encoding N-terminal fragments of human DNA topoisomerase IIalpha. We show that the N-terminal domain (approximately 50 kDa) has an intrinsic ATPase activity that can be stimulated by DNA. The enzyme obeys Michaelis-Menten kinetics showing a approximately 6-fold increase in kcat in the presence of DNA. Cross-linking studies indicate that the N-terminal domain is a dimer in the absence and presence of nucleotides. Using site-directed mutagenesis, we have identified the catalytic residue for ATP hydrolysis as Glu86. Phosphorylation of the N-terminal domain with protein kinase C does not affect the ATPase activity. The ATPase domain of human topoisomerase IIalpha shows significant differences from its counterpart in DNA gyrase and we discuss the mechanistic implications of these data.

Eukaryotic topoisomerase II enzymes are essential for efficient chromosome DNA segregation in both mitosis and meiosis (10, 19), and this makes them attractive targets for cytotoxic agents (3, 8). All topoisomerase II enzymes catalyze the passage of one DNA double helix through another via a transient double-stranded break in DNA. The topoisomerase II reaction requires the binding of the enzyme as a dimer and the creation of a 4-bp staggered break in the DNA via the formation of a covalent bond between each enzyme monomer and the 5′-DNA ends of a G (gate) segment of DNA. Another DNA segment, the T (transported) segment, is then captured by an ATP-operated clamp and passed through the broken gate strand, which is then religated (2). Topoisomerase II is inhibited by a variety of antitumor drugs. For example, doxorubicin, m-AMSA (amsacrine), epipodophyllotoxins, and mitoxantrone all interfere with the breakage and religation of the G segment of DNA, forming structures which favor DNA strand breakage often referred to as “cleavable complexes.” In the absence of antitumor agents, such structures are usually short-lived. The presence of antitumor agents induces a large number of cleavable complexes, which if unresolved ultimately lead to cell death (8). ICRF-159, a bisdioxopiperazine derivative which “locks” the ATP-operated clamp of the enzyme (18), and merbarone (7), a thiobarbiturate derivative which acts via an as yet unknown mechanism, also inhibit DNA topoisomerases and are cytotoxic agents. In contrast to what is found for many other eukaryotes, there are two isoforms of human topoisomerase II, topoisomerase IIα and topoisomerase IIβ. The α-isozyme form has a monomeric molecular mass of 170 kDa and is encoded by a gene on chromosome 17q21-22 (21), whereas the β isoform has a molecular mass of 180 kDa and is encoded by a gene on chromosome 3p24 (12). Although it is known that both human isoenzymes can be inhibited by antitumor agents such as etoposide, m-AMSA, and merbarone in vitro (6), the extent to which inhibition of either topoisomerase IIα or IIβ is cytotoxic in vivo is unclear. Topoisomerase IIα is known to be preferentially expressed during mitosis, whereas topoisomerase IIβ shows little variation in levels during the cell cycle (26). One would speculate from these data that topoisomerase IIα is the major target of cytotoxic agents. However, drug-resistant cell lines have shown altered levels of either or both topoisomerase isoforms, suggesting some drug selectivity for α or β isoforms (11, 24, 25), and there have been some in vitro studies suggesting that α and β isoforms respond differently to different topoisomerase inhibitors (7, 15). The exact nature of such selectivity has, however, been difficult to determine due to the problems associated with the isolation and separation of the two isoforms for both in vivo and in vitro studies. Saccharomyces cerevisiae has a single form of topoisomerase II which has been frequently used as a eukaryotic model in functional studies and in the study of antitumor agents (17, 23). An S. cerevisiae mutant temperature sensitive for topoisomerase II in combination with yeast/human hybrid topoisomerases has been used as a model to study the relative sensitivities of human α and β topoisomerase II enzymes to a variety of topoisomerases II inhibitors both in vitro and in vivo (4). Sensitivities to the antitopoisomerase drugs were estimated following a short contact inhibition assay (15) based on viable-cell counts. Such methods are highly labor intensive and can have quite large margins of error. We have previously shown that a topoisomerase II deletion strain of S. cerevisiae can be fully complemented by either of the two human topoisomerase II isozymes expressed from full-length cDNAs (13). Using this system we are now able to distinguish from one another the effects of different chemical classes of topoisomerase inhibitor on yeast topoisomerase II and on either of the full-length human α or β isoenzymes. In addition, this study uses a rapid throughput microwell assay, which is shown to be comparable to cell counting, to monitor the effects of drugs on yeast growth. Using this novel assay to generate data, we present a study which uses full-length human cDNAs expressed in a drug-permeable S. cerevisiae topoisomerase II deletion strain to determine the relative sensitivities of two human topoisomerases to a variety of anti-tumor drugs.

We have purified human topoisomerase IIalpha from HeLa cells and studied its ATPase reaction. The ATPase activity is stimulated by DNA and shows apparent Michaelis-Menten kinetics. Although the ATPase activity of human topoisomerase IIalpha is lower than that of Saccharomyces cerevisiae, it is more active in decatenation, implying more efficient coupling of the ATPase to DNA strand passage under these conditions. Using plasmid pBR322 as the DNA cofactor, the reaction shows hyperstimulation by DNA at a base pair to enzyme dimer ratio of 100-200:1. When DNA fragments are used as the cofactor, the reaction requiresapproximately 100 base pairs to stimulate the activity and fragments of approximately 300 base pairs show hyperstimulation. This behavior can be rationalized in terms of the enzyme requiring fragments that can bind to both the DNA gate and the ATP-operated clamp in order for the ATPase reaction to be stimulated. Hyperstimulation is a consequence of the saturation of DNA with enzyme. The mechanistic implications of these results are discussed.

The base excision repair (BER) pathway is essential for the removal of DNA bases damaged by alkylation or oxidation. A key step in BER is the processing of an apurinic/apyrimidinic (AP) site intermediate by an AP endonuclease. The major AP endonuclease in human cells (APE1, also termed HAP1 and Ref-1) accounts for >95% of the total AP endonuclease activity, and is essential for the protection of cells against the toxic effects of several classes of DNA damaging agents. Moreover, APE1 overexpression has been linked to radio- and chemo-resistance in human tumors. Using a newly developed high-throughput screen, several chemical inhibitors of APE1 have been isolated. Amongst these, CRT0044876 was identified as a potent and selective APE1 inhibitor. CRT0044876 inhibits the AP endonuclease, 3'-phosphodiesterase and 3'-phosphatase activities of APE1 at low micromolar concentrations, and is a specific inhibitor of the exonuclease III family of enzymes to which APE1 belongs. At non-cytotoxic concentrations, CRT0044876 potentiates the cytotoxicity of several DNA base-targeting compounds. This enhancement of cytotoxicity is associated with an accumulation of unrepaired AP sites. In silico modeling studies suggest that CRT0044876 binds to the active site of APE1. These studies provide both a novel reagent for probing APE1 function in human cells, and a rational basis for the development of APE1-targeting drugs for antitumor therapy.

A potential region of drug-DNA interaction in the A subunit of DNA gyrase has previously been identified from crystallographic studies. The local amino acid sequence has been compared with similar regions in yeast topoisomerase II and human topoisomerase IIalpha. Three non- conserved, potentially solvent-accessible residues at positions 762, 763 and 766 in human topoisomerase IIalpha lie between well-conserved regions. The corresponding residues in GyrA (83, 84 and 87) have a high frequency of mutation in quinolone-resistant bacteria. Mutations in human topoisomerase IIalpha have been generated in an attempt to engineer ciprofloxacin sensitivity into this enzyme: M762S, S763A and M766D (each mutated to the identical amino acid present in gyrase), along with an M762S/S763A double mutant and a triple mutant. These enzymes were introduced into a temperature-sensitive yeast strain, deficient in topoisomerase II, for in vivo studies, and were overproduced for in vitro studies. The M766D mutation renders the enzyme incapable of supporting the temperature-sensitive strain at a non-permissive temperature. However, both M766D and the triple mutant enzymes can be overproduced and are fully active in vitro. The double mutant was impaired in its ability to cleave DNA and had reduced catalytic activity. The triple mutation confers a three-fold increase in sensitivity to ciprofloxacin in vitro and similar sensitivities to a range of other quinolones. The activity of the quinolone CP-115,953, a bacterial and eukaryotic topoisomerase II poison, was unaffected by any of these mutations. Mutations in this region were found to increase the sensitivity of the enzyme to the DNA intercalating anti-tumour agents m-AMSA and ellipticine, but confer resistance to the non-intercalating agents etoposide, teniposide and merbarone, an effect that was maximal in the triple mutant. We have therefore shown the importance of this region in determining the sensitivity of topoisomerase II to drugs and have engineered increased sensitivity to quinolones.

New synthetic routes to a series of tetra- and pentacyclic acridines related in structure to marine natural products are reported. The novel water-soluble agent dihydroindolizino[7,6,5-kl]acridinium chloride 14 has inhibitory activity in a panel of non-small-cell lung and breast tumor cell lines exceeding that of m-AMSA. The salt inhibited the release of minicircle products of kDNA confirming that disorganization of topoisomerase II partly underlies the activity of the compound. COMPARE analysis of the NCI mean graph profile of compound 14 at the GI(50) level corroborates this conclusion with Pearson correlation coefficients (>0.6) to clinical agents of the topoisomerase II class: however, this correlation was not seen at the LC(50) level. The inhibitory action of 14 on Saccharomyces cerevisiae transfected with human topoisomerase II isoforms showed a 3-fold selectivity against the IIalpha isoform over the IIbeta isoform. Unlike m-AMSA, 14 is not susceptible to P-glycoprotein-mediated drug efflux and retains activity in lung cells with derived resistance to the topoisomerase II inhibitor etoposide.

We have recently shown that a carboxyl-terminal sequence of parathyroid hormone-related protein, PTHrP[107–139], is a potent direct inhibitor of osteoclastic bone resorption. We now report that this bone resorption inhibitory activity, which we have called osteostatin, is entirely contained within the highly conserved pentapeptide PTHrP[107–l 11]. Our results indicate that processing at residue 106 and a free amino terminus is required for full activity of the peptide. The retroinverted peptide is considerably less potent than the parent peptide. The retention of full biological activity in such a short fragment was unexpected. This data provides the basis for the development of further analogs with potential therapeutic use in disorders associated with increased osteoclastic bone resorption.

Introduction The purpose of ESD and Hybrid ESD (circumferential excision and snaring) is to obtain en bloc specimen. Margins are checked for residual tissue and APC applied if appropriate Methods Single endoscopist using ESD and Hybrid ESD (H-ESD) technique was retrospectively audited from April 2004 to August 2012. Service evaluation data of 38 patients with large sessile polyps who underwent ESD and Hybrid ESD was reviewed from a cohort of 224 colonoscopies referred for large polyp EMR. All procedures were intended as ESD. NICE recommendations for case selection were followed in 92% cases. Due to challenges in submucosal dissection of the large lesions, piecemeal resection was done after circumferential cutting. First follow-up endoscopy was performed at 3–6 months and the second at 12–14 months. Results Mean age was 70 with 16 males and 22 females. Mean size of polyp in ESD group was 26mm. Range 15–50mm. Mean size in the H-ESD group was 49mm. Range 20–100mm. Complete resections were achieved in 17 (44%) out of 38 cases. Due to piecemeal resection pathologists were not able to confirm adequacy of excision in 12 cases. In 9 cases resection was reported incomplete on index procedure. ESD performed in 13 (34%) cases. Complete resection achieved in 6. Out of 7 incomplete resections in the ESD group, 3 were reported by pathologists as lesion extending to the lateral margin hence incomplete excision. Histology did not comment on margin clearance in 3 ESD. 1 ESD was a sub mucosal lipoma on histology. This was an unintentional ESD for lipoma. Histology: ESD group: TVA with LGD 7, TVA with HGD 5. H-ESD was performed in 25 (65.7%) cases. Complete resection achieved in 11 cases, incomplete resection in 7 and lateral margin clearance not confirmed in 7 H-ESD cases due to piecemeal resection. Histology: H-ESD group: TVA with LGD 16, TVA with HGD 6 and adenocarcinoma in 2 cases-one’s lateral and deep margins were clear and the other was incomplete and referred to MDT In 4 ESD and 7 H-ESD cases there was minor bleeding controlled endoscopically at the time. 1 delayed post- H-ESD bleeding required 11 days of hospital stay and 2 units of blood transfusion. 1 retroperitoneal perforation and 1 case of serosal cut managed conservatively with clips and antibiotics. APC performed in 16 (42%) out of 38. Recurrence was identified in 6 H-ESD cases (15.7%). 5 local recurrences detected at 3 months and 1 local recurrence detected at 24 months. In 13 ESD cases no perforation or recurrence upto 14 months was noted despite 5 histologically incomplete dissections. Conclusion ESD in bowel is challenging and has a long learning curve. These procedures should be performed by trained endoscopists in accredited units and a national registry should be maintained. Disclosure of Interest None Declared.

The antiviral activity of podophyllotoxin against herpes simplex type 1 virus (HSV-1) grown in Vero cells was studied by a simple microtitration assay. Antiviral effects were induced at similar concentrations as direct cellular toxicity, as characterised by a time-dependent loss of cell monolayer. Podophyllotoxin-mediated toxicity arises from cytoplasmic microtubular, and hence cytoskeletal, decay. Some degree of selectivity was seen for inhibition of virus replication over direct cellular toxicity. Podophyllotoxin acted against an early viral process, as an antiviral effect was still seen if drug was removed 2 h after infection. Similar effects were seen with colchicine, a classical tubulin-binding compound, but not with bromovinyldeoxyuridine. Podophyllotoxin was capable of inducing a cytoprotective effect in Vero cells, as pre-treatment of cells abrogated virus growth for up to 90 min after removal of drug. This is coincident with the repolymerisation of cellular microtubules and re-formation of the cytoskeleton. We conclude that HSV-1 relies upon a functional cellular cytoskeleton for efficient completion of an early replicative event. Such a process may be the transport of viral material to the nucleus or inhibition of the formation of intranuclear viral 'replication factories', bodies containing cytoskeletal fragments constructed after viral infection.

Humoral hypercalcemia of malignancy is a common complication of lung and certain other cancers. The hypercalcemia results from the actions of tumor factors on bone and kidney. We report here the isolation of full-length complementary DNA clones of a putative hypercalcemia factor, and the expression from the cloned DNA of the active protein in mammalian cells. The clones encode a prepro peptide of 36 amino acids and a mature protein of 141 amino acids that has significant homology with parathyroid hormone in the amino-terminal region. This previously unrecognized hormone may be important in normal as well as abnormal calcium metabolism.

Psychiatric hospital records were used to ascertain whether acting-out behavior increases during the full moon phase. Acting-out behavior is identified as that which was dangerous to self or others to the extent that either seclusion or restraint was deemed necessary by qualified mental health professionals to prevent physical harm. Records in the seclusion/restraint log book of an Alabama hospital for 1982, 1983, and 1984 were used for the study. To ensure confidentiality no identification of subjects was made. This study, like many previous similar studies shows no significant increases during the full moon phase.