Evaluation of Phage Display Discovered Peptides as Ligands for Prostate-Specific Membrane Antigen (PSMA)

The aim of this study was to identify potential ligands of PSMA suitable for further development as novel PSMA-targeted peptides using phage display technology. The human PSMA protein was immobilized as a target followed by incubation with a 15-mer phage display random peptide library. After one round of prescreening and two rounds of screening, high-stringency screening at the third round of panning was performed to identify the highest affinity binders. Phages which had a specific binding activity to PSMA in human prostate cancer cells were isolated and the DNA corresponding to the 15-mers were sequenced to provide three consensus sequences: GDHSPFT, SHFSVGS and EVPRLSLLAVFL as well as other sequences that did not display consensus. Two of the peptide sequences deduced from DNA sequencing of binding phages, SHSFSVGSGDHSPFT and GRFLTGGTGRLLRIS were labeled with 5-carboxyfluorescein and shown to bind and co-internalize with PSMA on human prostate cancer cells by fluorescence microscopy. The high stringency requirements yielded peptides with affinities KD∼1 μM or greater which are suitable starting points for affinity maturation. While these values were less than anticipated, the high stringency did yield peptide sequences that apparently bound to different surfaces on PSMA. These peptide sequences could be the basis for further development of peptides for prostate cancer tumor imaging and therapy. © 2013 Shen et al

Teachers as Educational Innovators in Inquiry-Based Science Teaching and Learning

This chapter describes inquiry-based science teaching and learning (IBST/L) pilots designed by teachers during a professional development programme. There is research-based evidence that IBSL/T may promote students’ learning and their motivation to learn science, and therefore it is beneficial to familiarise the teachers with this approach. Building on teachers’ existing expertise in designing their teaching, the programme introduced theoretical aspects of the IBST/L approach and its research-based benefits for students’ motivation, interest and science learning. The course aimed to support teachers as educational innovators in the process of designing and testing IBST/L pilots, during which they collaboratively reflected on and revised their existing practices. The data of this piece of research consists of the teachers’ poster presentations of their IBST/L pilots and a video recording of the reflection session. The content analysis revealed that the pilots’ structure seemed traditional but encompassed some IBST/L features. It is concluded that teacher educators need to understand teachers’ views of IBST/L in order to more effectively support planning and reflection.This chapter describes inquiry-based science teaching and learning (IBST/L) pilots designed by teachers during a professional development programme. There is research-based evidence that IBSL/T may promote students’ learning and their motivation to learn science, and therefore it is beneficial to familiarise the teachers with this approach. Building on teachers’ existing expertise in designing their teaching, the programme introduced theoretical aspects of the IBST/L approach and its research-based benefits for students’ motivation, interest and science learning. The course aimed to support teachers as educational innovators in the process of designing and testing IBST/L pilots, during which they collaboratively reflected on and revised their existing practices. The data of this piece of research consists of the teachers’ poster presentations of their IBST/L pilots and a video recording of the reflection session. The content analysis revealed that the pilots’ structure seemed traditional but encompassed some IBST/L features. It is concluded that teacher educators need to understand teachers’ views of IBST/L in order to more effectively support planning and reflection.Peer reviewe

Reverse logistics, stakeholders' influence, organizational slack, and managers' posture

Reverse logistics (RL) has strategic importance. However, little is known concerning what motivates firms to adopt RL systems. Drawing on stakeholder theory formulations, organizational slack, and the manager's strategic stance concept, this article develops a model that proposes external, internal, and individual factors that affect the implementation of RL programs. Our framework considers three major explicative variables: the attributes of the stakeholder (power, legitimacy and urgency), organizational slack for RL programs, and the manager's strategic posture. The study draws on a sample of 118 Spanish companies and uses a probit model to determine the influence of these factors on the probability of firms to implement RL systems. The study finds that customers, employees, and the government salience in terms of RL activities and manager's progressive posture have a significant influence on the final decision of implanting RL programs. Conversely, the study finds that shareholder salience negatively impacts the decision.Fundación BBVA and the project “Estrategias en las empresas europeas internacionales y medioambiente: análisis de recursos humanos, producción y negocio” for supporting this research. They also acknowledge the help of Prof. Dr. Carlos Larrinaga in providing access to financial statements. They are indebted as well to the Spanish Minister of Education and Science and the Comunidad Autónoma de Madrid which have provided financial support under the research projects entitled “Diseño e implantación de las estrategias medioambientales de aprovisionamiento, fabricación y comercialización: relación con el rendimiento organizativo” (sec2001-1578-c02- 01), “La dirección de operaciones y las teorías de stakeholders y de recursos y capacidades. Implicaciones estratégicas en la gestión del ciclo de vida de los equipos informáticos” (SEJ04- 07877-C02-02),. “Efectos de los distintos tipos de relaciones entre las empresas y sus mercados: prácticas de marketing, orientación al mercado y resultados económicos”, and “ Innovación versus imitacón: un test de la capacidad innovadora de las empresas de la Comunidad de Madrid “.Publicad

Role of TMPRSS2-ERG Gene Fusion in Negative Regulation of PSMA Expression

Prostate specific membrane antigen (PSMA) is overexpressed in prostatic adenocarcinoma (CaP), and its expression is negatively regulated by androgen stimulation. However, it is still unclear which factors are involved in this downregulation. TMPRSS2-ERG fusion is the most common known gene rearrangement in prostate carcinoma. Androgen stimulation can increase expression of the TMPRSS2-ERG fusion in fusion positive prostate cancer cells. The purpose of this investigation is to determine whether PSMA expression can be regulated by the TMPRSS2-ERG gene fusion. We employed two PSMA positive cell lines: VCaP cells, which harbor TMPRSS2-ERG fusion, and LNCaP cells, which lack the fusion. After 24 hours of androgen treatment, TMPRSS2-ERG mRNA level was increased in VCaP cells. PSMA mRNA level was dramatically decreased in VCaP cells, while it only has moderate change in LNCaP cells. Treatment with the androgen antagonist flutamide partially restored PSMA expression in androgen-treated VCaP cells. Knocking down ERG by siRNA in VCaP cells enhances PSMA expression both in the presence and absence of synthetic androgen R1881. Overexpressing TMPRSS2-ERG fusions in LNCaP cells downregulated PSMA both in the presence or absence of R1881, while overexpressing wild type ERG did not. Using PSMA-based luciferase reporter assays, we found TMPRSS2-ERG fusion can inhibit PSMA activity at the transcriptional level. Our data indicated that downregulation of PSMA in androgen-treated VCaP cells appears partially mediated by TMPRSS2-ERG gene fusion

ALCAM Regulates Motility, Invasiveness, and Adherens Junction Formation in Uveal Melanoma Cells

ALCAM, a member of the immunoglobulin superfamily, has been implicated in numerous developmental events and has been repeatedly identified as a marker for cancer metastasis. Previous studies addressing ALCAM’s role in cancer have, however, yielded conflicting results. Depending on the tumor cell type, ALCAM expression has been reported to be both positively and negatively correlated with cancer progression and metastasis in the literature. To better understand how ALCAM might regulate cancer cell behavior, we utilized a panel of defined uveal melanoma cell lines with high or low ALCAM levels, and directly tested the effects of manipulating these levels on cell motility, invasiveness, and adhesion using multiple assays. ALCAM expression was stably silenced by shRNA knockdown in a high-ALCAM cell line (MUM-2B); the resulting cells displayed reduced motility in gap-closure assays and a reduction in invasiveness as measured by a transwell migration assay. Immunostaining revealed that the silenced cells were defective in the formation of adherens junctions, at which ALCAM colocalizes with N-cadherin and ß-catenin in native cells. Additionally, we stably overexpressed ALCAM in a low-ALCAM cell line (MUM-2C); intriguingly, these cells did not exhibit any increase in motility or invasiveness, indicating that ALCAM is necessary but not sufficient to promote metastasis-associated cell behaviors. In these ALCAM-overexpressing cells, however, recruitment of ß-catenin and N-cadherin to adherens junctions was enhanced. These data confirm a previously suggested role for ALCAM in the regulation of adherens junctions, and also suggest a mechanism by which ALCAM might differentially enhance or decrease invasiveness, depending on the type of cadherin adhesion complexes present in tissues surrounding the primary tumor, and on the cadherin status of the tumor cells themselves

ALCAM (CD166) Expression and Serum Levels in Pancreatic Cancer

BACKGROUND: This study was conducted to evaluate the expression of the activated leukocyte cell adhesion molecule (ALCAM) in pancreatic cancer (PAC) and to determine whether or not the ectodomain shedding of ALCAM (s-ALCAM) could serve as a biomarker in the peripheral blood of PAC patients. MATERIAL AND METHODS: Tissue specimens and blood sera of patients with PAC (n = 264 and n = 116, respectively) and the sera of 115 patients with chronic pancreatitis (CP) were analyzed via ALCAM immunohistochemistry and s-ALCAM ELISA tests. Results were correlated with clinical, histopathological, and patient survival data (Chi-square test, Kaplan-Meier analysis, log-rank test, respectively). RESULTS: ALCAM was expressed in the majority of PAC lesions. Immunohistochemistry and serum ELISA tests revealed no association between ALCAM expression in primary tumors or s-ALCAM and clinical or histopathological data. Neither ALCAM nor s-ALCAM showed a significant impact regarding overall survival (p = 0.261 and p = 0.660, respectively). S-ALCAM serum levels were significantly elevated compared to the sera of CP patients (p<0.001). The sensitivity of s-ALCAM in detecting PAC was 58.6% at a specificity of 73.9% (AUC = 0.69). CONCLUSIONS: ALCAM is expressed in the majority of PAC lesions, but statistical analysis revealed no association with clinical or pathological data. Although significantly elevated in patients with PAC, the sensitivity and specificity of the s-ALCAM serum quantification test was low. Therefore, its potential as a novel diagnostic marker for PAC remains elusive and further investigations are required

Pan-cancer analysis of whole genomes

Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale(1-3). Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4-5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter(4); identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation(5,6); analyses timings and patterns of tumour evolution(7); describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity(8,9); and evaluates a range of more-specialized features of cancer genomes(8,10-18).Peer reviewe