Induction of endothelial cell apoptosis by the antivascular agent 5,6-dimethylxanthenone-4-acetic acid

5,6-Dimethylxanthenone-4-acetic acid, synthesised in this laboratory, reduces tumour blood flow, both in mice and in patients on Phase I trial. We used TUNEL (TdT-mediated dUTP nick end labelling) assays to investigate whether apoptosis induction was involved in its antivascular effect. 5,6-Dimethylxanthenone-4-acetic acid induced dose-dependent apoptosis in vitro in HECPP murine endothelial cells in the absence of up-regulation of mRNA for tumour necrosis factor. Selective apoptosis of endothelial cells was detected in vivo in sections of Colon 38 tumours in mice within 30 min of administration of 5,6-Dimethylxanthenone-4-acetic acid (25 mg kg−1). TUNEL staining intensified with time and after 3 h, necrosis of adjacent tumour tissue was observed. Apoptosis of central vessels in splenic white pulp was also detected in tumour-bearing mice but not in mice without tumours. Apoptosis was not observed in liver tissue. No apoptosis was observed with the inactive analogue 8-methylxanthenone-4-acetic acid. Positive TUNEL staining of tumour vascular endothelium was evident in one patient in a Phase I clinical trial, from a breast tumour biopsy taken 3 and 24 h after infusion of 5,6-Dimethylxanthenone-4-acetic acid (3.1 mg m−2). Tumour necrosis and the production of tumour tumour necrosis factor were not observed. No apoptotic staining was seen in tumour biopsies taken from two other patients (doses of 3.7 and 4.9 mg m−2). We conclude that 5,6-Dimethylxanthenone-4-acetic acid can induce vascular endothelial cell apoptosis in some murine and human tumours. The action is rapid and appears to be independent of tumour necrosis factor induction

Identification and characterization of a novel non-structural protein of bluetongue virus

Bluetongue virus (BTV) is the causative agent of a major disease of livestock (bluetongue). For over two decades, it has been widely accepted that the 10 segments of the dsRNA genome of BTV encode for 7 structural and 3 non-structural proteins. The non-structural proteins (NS1, NS2, NS3/NS3a) play different key roles during the viral replication cycle. In this study we show that BTV expresses a fourth non-structural protein (that we designated NS4) encoded by an open reading frame in segment 9 overlapping the open reading frame encoding VP6. NS4 is 77–79 amino acid residues in length and highly conserved among several BTV serotypes/strains. NS4 was expressed early post-infection and localized in the nucleoli of BTV infected cells. By reverse genetics, we showed that NS4 is dispensable for BTV replication in vitro, both in mammalian and insect cells, and does not affect viral virulence in murine models of bluetongue infection. Interestingly, NS4 conferred a replication advantage to BTV-8, but not to BTV-1, in cells in an interferon (IFN)-induced antiviral state. However, the BTV-1 NS4 conferred a replication advantage both to a BTV-8 reassortant containing the entire segment 9 of BTV-1 and to a BTV-8 mutant with the NS4 identical to the homologous BTV-1 protein. Collectively, this study suggests that NS4 plays an important role in virus-host interaction and is one of the mechanisms played, at least by BTV-8, to counteract the antiviral response of the host. In addition, the distinct nucleolar localization of NS4, being expressed by a virus that replicates exclusively in the cytoplasm, offers new avenues to investigate the multiple roles played by the nucleolus in the biology of the cell

Randomised phase II study of ASA404 combined with carboplatin and paclitaxel in previously untreated advanced non-small cell lung cancer

ASA404 (5,6-dimethylxanthenone-4-acetic acid or DMXAA) is a small-molecule tumour-vascular disrupting agent (Tumour-VDA). This randomised phase II study evaluated ASA404 plus standard therapy of carboplatin and paclitaxel in patients with histologically confirmed stage IIIb or IV non-small cell lung cancer (NSCLC) not previously treated with chemotherapy. Patients were randomised to receive ⩽6 cycles of carboplatin area under the plasma concentration–time curve 6 mg ml−1 min and paclitaxel 175 mg m−2 (CP, n=36) or standard therapy plus ASA404 1200 mg m−2 (ASA404-CP, n=37). There was little change in the systemic exposure of either total or free carboplatin or paclitaxel on addition of ASA404. Safety profiles were similar and manageable in both groups, with most adverse effects attributed to standard therapy. Tumour response rate (31 vs 22%), median time to tumour progression (5.4 vs 4.4 months) and median survival (14.0 vs 8.8 months, hazard ratio 0.73, 95% CI 0.39, 1.38) were improved in the ASA404 combination group compared with the standard therapy group. In conclusion, this study establishes the feasibility of combining ASA404 with carboplatin and paclitaxel in patients with previously untreated, advanced NSCLC, demonstrating a manageable safety profile and lack of adverse pharmacokinetic interactions. The results indicate that there may be a benefit associated with ASA404, but this needs to be evaluated in a larger trial

3D Variation in delineation of head and neck organs at risk

<p>Abstract</p> <p>Background</p> <p>Consistent delineation of patient anatomy becomes increasingly important with the growing use of highly conformal and adaptive radiotherapy techniques. This study investigates the magnitude and 3D localization of interobserver variability of organs at risk (OARs) in the head and neck area with application of delineation guidelines, to establish measures to reduce current redundant variability in delineation practice.</p> <p>Methods</p> <p>Interobserver variability among five experienced radiation oncologists was studied in a set of 12 head and neck patient CT scans for the spinal cord, parotid and submandibular glands, thyroid cartilage, and glottic larynx. For all OARs, three endpoints were calculated: the Intraclass Correlation Coefficient (ICC), the Concordance Index (CI) and a 3D measure of variation (3D SD).</p> <p>Results</p> <p>All endpoints showed largest interobserver variability for the glottic larynx (ICC = 0.27, mean CI = 0.37 and 3D SD = 3.9 mm). Better agreement in delineations was observed for the other OARs (range, ICC = 0.32-0.83, mean CI = 0.64-0.71 and 3D SD = 0.9-2.6 mm). Cranial, caudal, and medial regions of the OARs showed largest variations. All endpoints provided support for improvement of delineation practice.</p> <p>Conclusions</p> <p>Variation in delineation is traced to several regional causes. Measures to reduce this variation can be: (1) guideline development, (2) joint delineation review sessions and (3) application of multimodality imaging. Improvement of delineation practice is needed to standardize patient treatments.</p

The antitumour activity of 5,6-dimethylxanthenone-4-acetic acid (DMXAA) in TNF receptor-1 knockout mice

5,6-dimethylxanthenone-4-acetic acid, a novel antivascular anticancer drug, has completed Phase I clinical trial. Its actions in mice include tumour necrosis factor induction, serotonin release, tumour blood flow inhibition, and the induction of tumour haemorrhagic necrosis and regression. We have used mice with a targeted disruption of the tumour necrosis factor receptor-1 gene as recipients for the colon 38 carcinoma to determine the role of tumour necrosis factor signalling in the action of 5,6-dimethylxanthenone-4-acetic acid. The pharmacokinetics of 5,6-dimethylxanthenone-4-acetic acid, as well as the degree of induced plasma and tissue tumour necrosis factor, were similar in tumour necrosis factor receptor-1−/− and wild-type mice. However, the maximum tolerated dose of 5,6-dimethylxanthenone-4-acetic acid was considerably higher in tumour necrosis factor receptor-1−/− mice (>100 mg kg−1) than in wild-type mice (27.5 mg kg−1). The antitumour activity of 5,6-dimethylxanthenone-4-acetic acid (25 mg kg−1) was strongly attenuated in tumour necrosis factor receptor-1−/− mice. However, the reduced toxicity in tumour necrosis factor receptor-1−/− mice allowed the demonstration that at a higher dose (50 mg kg−1), 5,6-dimethylxanthenone-4-acetic acid was curative and comparable in effect to that of a lower dose (25 mg kg−1) in wild-type mice. The 5,6-dimethylxanthenone-4-acetic acid -induced rise in plasma 5-hydroxyindoleacetic acid, used to reflect serotonin production in a vascular response, was larger in colon 38 tumour bearing than in non-tumour bearing tumour necrosis factor receptor-1−/− mice, but in each case the response was smaller than the corresponding response in wild-type mice. The results suggest an important role for tumour necrosis factor in mediating both the host toxicity and antitumour activity of 5,6-dimethylxanthenone-4-acetic acid, but also suggest that tumour necrosis factor can be replaced by other vasoactive factors in its antitumour action, an observation of relevance to current clinical studies

End-stage heart failure in congenitally corrected transposition of the great arteries:a multicentre study

BACKGROUND AND AIMS: For patients with congenitally corrected transposition of the great arteries (ccTGA), factors associated with progression to end-stage congestive heart failure (CHF) remain largely unclear. METHODS: This multicentre, retrospective cohort study included adults with ccTGA seen at a congenital heart disease centre. Clinical data from initial and most recent visits were obtained. The composite primary outcome was mechanical circulatory support, heart transplantation, or death. RESULTS: From 558 patients (48% female, age at first visit 36 ± 14.2 years, median follow-up 8.7 years), the event rate of the primary outcome was 15.4 per 1000 person-years (11 mechanical circulatory support implantations, 12 transplantations, and 52 deaths). Patients experiencing the primary outcome were older and more likely to have a history of atrial arrhythmia. The primary outcome was highest in those with both moderate/severe right ventricular (RV) dysfunction and tricuspid regurgitation (n = 110, 31 events) and uncommon in those with mild/less RV dysfunction and tricuspid regurgitation (n = 181, 13 events, P &lt; .001). Outcomes were not different based on anatomic complexity and history of tricuspid valve surgery or of subpulmonic obstruction. New CHF admission or ventricular arrhythmia was associated with the primary outcome. Individuals who underwent childhood surgery had more adverse outcomes than age- and sex-matched controls. Multivariable Cox regression analysis identified older age, prior CHF admission, and severe RV dysfunction as independent predictors for the primary outcome. CONCLUSIONS: Patients with ccTGA have variable deterioration to end-stage heart failure or death over time, commonly between their fifth and sixth decades. Predictors include arrhythmic and CHF events and severe RV dysfunction but not anatomy or need for tricuspid valve surgery.</p

Immunophenotyping of Circulating T Helper Cells Argues for Multiple Functions and Plasticity of T Cells In Vivo in Humans - Possible Role in Asthma

BACKGROUND: The immune process driving eosinophilic and non-eosinophilic asthma is likely driven by different subsets of T helper (Th) cells. Recently, in vitro studies and animal studies suggest that Th cell subsets displays plasticity by changing their transcription factor or by expressing multiple transcription factors. Our aim was to determine whether individuals with asthma and elevated circulating eosinophils express signs of different regulatory immune mechanisms compared with asthmatics with low blood eosinophils and non-asthmatic control subjects. In addition, determine the relationship between eosinophilia and circulating Th cell subsets. METHODOLOGY/PRINCIPAL FINDINGS: Participants were selected from a random epidemiological cohort, the West Sweden Asthma Study. Immunophenotypes of fresh peripheral blood cells obtained from stable asthmatics, with and without elevated eosinophilic inflammation (EOS high and EOS low respectively) and control subjects, were determined by flow cytometry. No differences in the number of Th1 (T-bet), Th2 (GATA-3), Th17 (RORγt) or Treg (FOXP3) cells were observed between the groups when analysing each subset separately. However, in all groups, each of the Th subsets showed expression of additional canonical transcription factors T-bet, GATA-3, RORγt and FOXP3. Furthermore, by in vitro stimulation with anti-CD3/anti-CD28 there was a significant increase of single expressing GATA-3(+) and co-expressing T-bet(+)GATA-3(+) cells in the EOS high asthmatics in comparison with control subjects. In addition, T-bet(-)GATA-3(+)RORγt(+)FOXP3(+) were decreased in comparison to the EOS low asthmatics. Finally, in a group of control subjects we found that the majority of proliferating Th cells (CD4(+)CD25(+)Ki67(+)) expressed three or four transcription factors. CONCLUSIONS: The ability of human Th cells to express several regulatory transcription factors suggests that these cells may display plasticity in vivo