Identification of novel elements of the Drosophila blisterome sheds light on potential pathological mechanisms of several human diseases.

Main developmental programs are highly conserved among species of the animal kingdom. Improper execution of these programs often leads to progression of various diseases and disorders. Here we focused on Drosophila wing tissue morphogenesis, a fairly complex developmental program, one of the steps of which - apposition of the dorsal and ventral wing sheets during metamorphosis - is mediated by integrins. Disruption of this apposition leads to wing blistering which serves as an easily screenable phenotype for components regulating this process. By means of RNAi-silencing technique and the blister phenotype as readout, we identify numerous novel proteins potentially involved in wing sheet adhesion. Remarkably, our results reveal not only participants of the integrin-mediated machinery, but also components of other cellular processes, e.g. cell cycle, RNA splicing, and vesicular trafficking. With the use of bioinformatics tools, these data are assembled into a large blisterome network. Analysis of human orthologues of the Drosophila blisterome components shows that many disease-related genes may contribute to cell adhesion implementation, providing hints on possible mechanisms of these human pathologies

High capacity in G protein-coupled receptor signaling.

G protein-coupled receptors (GPCRs) constitute a large family of receptors that activate intracellular signaling pathways upon detecting specific extracellular ligands. While many aspects of GPCR signaling have been uncovered through decades of studies, some fundamental properties, like its channel capacity-a measure of how much information a given transmission system can reliably transduce-are still debated. Previous studies concluded that GPCRs in individual cells could transmit around one bit of information about the concentration of the ligands, allowing only for a reliable on or off response. Using muscarinic receptor-induced calcium response measured in individual cells upon repeated stimulation, we show that GPCR signaling systems possess a significantly higher capacity. We estimate the channel capacity of this system to be above two, implying that at least four concentration levels of the agonist can be distinguished reliably. These findings shed light on the basic principles of GPCR signaling

Strategic investment in tuberculosis control in the Republic of Bulgaria

As Bulgaria transitions away from Global Fund grant, robust estimates of the comparative impact of the various response strategies under consideration are needed to ensure sustained effectiveness of the tuberculosis (TB) programme. We tailored an established mathematical model for TB control to the epidemic in Bulgaria to project the likely outcomes of seven intervention scenarios. Under existing programmatic conditions projected forward, the country's targets for achieving TB elimination in the coming decades will not be achieved. No interventions under consideration were predicted to accelerate the baseline projected reduction in epidemiological indicators significantly. Discontinuation of the 'Open Doors' program and activities of non-governmental organisations would result in a marked exacerbation of the epidemic (increasing incidence in 2035 by 6-8% relative to baseline conditions projected forward). Changing to a short course regimen for multidrug-resistant TB (MDR-TB) would substantially decrease MDR-TB mortality (by 21.6% in 2035 relative to baseline conditions projected forward). Changing to ambulatory care for eligible patients would not affect TB burden but would be markedly cost-saving. In conclusion, Bulgaria faces important challenges in transitioning to a primarily domestically-financed TB programme. The country should consider maintaining currently effective programs and shifting towards ambulatory care to ensure program sustainability

A protein kinase Cβ inhibitor attenuates multidrug resistance of neuroblastoma cells

BACKGROUND: The acquisition of drug resistance is a major reason for poor outcome of neuroblastoma. Protein kinase C (PKC) has been suggested to influence drug resistance in cancer cells. The aim of this study was to elucidate whether inhibition of PKCβ isoforms influences drug-resistance of neuroblastoma cells. METHODS: The effect of the PKCβ inhibitor LY379196 on the growth-suppressing effects of different chemotherapeutics on neuroblastoma cells was analyzed with MTT assays. The effect of LY379196 on the accumulation of [(3)H]vincristine was also investigated RESULTS: The PKCβ inhibitor LY379196 suppressed the growth of three neuroblastoma cell lines. LY379196 also augmented the growth-suppressive effect of doxorubicin, etoposide, paclitaxel, and vincristine, but not of carboplatin. The effect was most marked for vincristine and for the cell-line (SK-N-BE(2)) that was least sensitive to vincristine. No effect was observed on the non-resistant IMR-32 cells. Two other PKC inhibitors, Gö6976 and GF109203X, also enhanced the vincristine effect. The PKC inhibitors caused an increased accumulation of [(3)H]vincristine in SK-N-BE(2) cells. CONCLUSIONS: This indicates that inhibition of PKCβ could attenuate multidrug resistance in neuroblastoma cells by augmenting the levels of natural product anticancer drugs in resistant cells

Wig-1, a novel regulator of N-Myc mRNA and N-Myc-driven tumor growth

Wig-1 is a transcriptional target of the p53 tumor suppressor and encodes an mRNA stability-regulating protein. We show here that Wig-1 knockdown causes a dramatic inhibition of N-Myc expression and triggers differentiation in neuroblastoma cells carrying amplified N-Myc. Transient Wig-1 knockdown significantly delays development of N-Myc-driven tumors in mice. We also show that N-Myc expression is induced upon moderate p53-activating stress, suggesting a role of the p53-Wig-1-N-Myc axis in promoting cell cycle re-entry upon p53-induced cell cycle arrest and DNA repair. Moreover, our findings raise possibilities for the improved treatment of poor prognosis neuroblastomas that carry amplified N-Myc

Valproic acid inhibits adhesion of vincristine- and cisplatin-resistant neuroblastoma tumour cells to endothelium

Drug resistance to chemotherapy is often associated with increased malignancy in neuroblastoma (NB). In pursuit of alternative treatments for chemoresistant tumour cells, we tested the response of multidrug-resistant SKNSH and of vincristine (VCR)-, doxorubicin (DOX)-, or cisplatin (CDDP)-resistant UKF-NB-2, UKF-NB-3 or UKF-NB-6 NB tumour cell lines to valproic acid (VPA), a differentiation inducer currently in clinical trials. Drug resistance caused elevated NB adhesion (UKF-NB-2VCR, UKF-NB-2DOX, UKF-NB-2CDDP, UKF-NB-3VCR, UKF-NB-3CDDP, UKF-NB-6VCR, UKF-NB-6CDDP) to an endothelial cell monolayer, accompanied by downregulation of the adhesion receptor neural cell adhesion molecule (NCAM). Based on the UKF-NB-3 model, N-myc proteins were enhanced in UKF-NB-3VCR and UKF-NB-3CDDP, compared to the drug naïve controls. p73 was diminished, whereas the p73 isoform deltaNp73 was upregulated in UKF-NB-3VCR and UKF-NB-3CDDP. Valproic acid blocked adhesion of UKF-NB-3VCR and UKF-NB-3CDDP, but not of UKF-NB-3DOX, and induced the upregulation of NCAM surface expression, NCAM protein content and NCAM coding mRNA. Valproic acid diminished N-myc and enhanced p73 protein level, coupled with downregulation of deltaNp73 in UKF-NB-3VCR and UKF-NB-3CDDP. Valproic acid also reverted enhanced adhesion properties of drug-resistant UKF-NB-2, UKF-NB-6 and SKNSH cells, and therefore may provide an alternative approach to the treatment of drug-resistant NB by blocking invasive processes

Neuroblastoma Cell Lines Contain Pluripotent Tumor Initiating Cells That Are Susceptible to a Targeted Oncolytic Virus

Although disease remission can frequently be achieved for patients with neuroblastoma, relapse is common. The cancer stem cell theory suggests that rare tumorigenic cells, resistant to conventional therapy, are responsible for relapse. If true for neuroblastoma, improved cure rates may only be achieved via identification and therapeutic targeting of the neuroblastoma tumor initiating cell. Based on cues from normal stem cells, evidence for tumor populating progenitor cells has been found in a variety of cancers.Four of eight human neuroblastoma cell lines formed tumorspheres in neural stem cell media, and all contained some cells that expressed neurogenic stem cell markers including CD133, ABCG2, and nestin. Three lines tested could be induced into multi-lineage differentiation. LA-N-5 spheres were further studied and showed a verapamil-sensitive side population, relative resistance to doxorubicin, and CD133+ cells showed increased sphere formation and tumorigenicity. Oncolytic viruses, engineered to be clinically safe by genetic mutation, are emerging as next generation anticancer therapeutics. Because oncolytic viruses circumvent typical drug-resistance mechanisms, they may represent an effective therapy for chemotherapy-resistant tumor initiating cells. A Nestin-targeted oncolytic herpes simplex virus efficiently replicated within and killed neuroblastoma tumor initiating cells preventing their ability to form tumors in athymic nude mice.These results suggest that human neuroblastoma contains tumor initiating cells that may be effectively targeted by an oncolytic virus

HDAC1 Inactivation Induces Mitotic Defect and Caspase-Independent Autophagic Cell Death in Liver Cancer

Histone deacetylases (HDACs) are known to play a central role in the regulation of several cellular properties interlinked with the development and progression of cancer. Recently, HDAC1 has been reported to be overexpressed in hepatocellular carcinoma (HCC), but its biological roles in hepatocarcinogenesis remain to be elucidated. In this study, we demonstrated overexpression of HDAC1 in a subset of human HCCs and liver cancer cell lines. HDAC1 inactivation resulted in regression of tumor cell growth and activation of caspase-independent autophagic cell death, via LC3B-II activation pathway in Hep3B cells. In cell cycle regulation, HDAC1 inactivation selectively induced both p21WAF1/Cip1 and p27Kip1 expressions, and simultaneously suppressed the expression of cyclin D1 and CDK2. Consequently, HDAC1 inactivation led to the hypophosphorylation of pRb in G1/S transition, and thereby inactivated E2F/DP1 transcription activity. In addition, we demonstrated that HDAC1 suppresses p21WAF1/Cip1 transcriptional activity through Sp1-binding sites in the p21WAF1/Cip1 promoter. Furthermore, sustained suppression of HDAC1 attenuated in vitro colony formation and in vivo tumor growth in a mouse xenograft model. Taken together, we suggest the aberrant regulation of HDAC1 in HCC and its epigenetic regulation of gene transcription of autophagy and cell cycle components. Overexpression of HDAC1 may play a pivotal role through the systemic regulation of mitotic effectors in the development of HCC, providing a particularly relevant potential target in cancer therapy

Comparative analysis of xanafide cytotoxicity in breast cancer cell lines

Xanafide, a DNA-intercalating agent and topoisomerase II inhibitor, has previously demonstrated comparable cytotoxicity to the parent drug amonafide (NSC 308847). The current study was conducted to investigate further the anti-proliferative effects of xanafide in human breast cancer cell lines, in vitro and in vivo. The in vitro activity of xanafide against MCF-7, MDA-MB-231, SKBR-3 and T47D cell lines was compared to that of paclitaxel, docetaxel, gemcitabine, vinorelbine and doxorubicin. In MCF-7, xanafide demonstrated comparable total growth inhibition (TGI) concentrations to the taxanes and lower TGI values than gemcitabine, vinorelbine and doxorubicin. MCF-7 (oestrogen receptor (ER)+/p53 wild-type) was the most sensitive cell line to xanafide. MDA-MB-231 and SKBR-3 exhibited similar sensitivity to xanafide. T47 D (ER+/p53 mutated), showed no response to this agent. The in vivo activity of xanafide was further compared to that of docetaxel in MCF-7 and MDA-MB-231 cell lines using the hollow fibre assay. Xanafide was slightly more potent than docetaxel, at its highest dose in MCF-7 cell line, whereas docetaxel was more effective than xanafide in MDA-MB-231 cell line. Our results show that there is no relationship between sensitivity of these cell lines to xanafide and cellular levels of both isoforms of topoisomerase II and suggest that ER and p53 status and their crosstalk may predict the responsiveness or resistance of breast cancer patients to xanafide