High-affinity recombinant phage antibodies to the pan-carcinoma marker epithelial glycoprotein-2 for tumour targeting.

The tumour-associated antigen epithelial glycoprotein-2 (EGP-2) is a promising target for detection and treatment of a variety of human carcinomas. Antibodies to this antigen have been successfully used in patients for imaging of small-cell lung cancer and for adjuvant treatment of minimal residual disease of colon cancer. We describe here the isolation and complete characterization of high-affinity single-chain variable fragments (scFv) to the EGP-2 antigen. First, the binding kinetics of four murine whole antibodies directed to EGP-2 (17-1A, 323/A3, MOC-31 and MOC-161) were determined using surface plasmon resonance (SPR). The MOC-31 antibody has the lowest apparent off-rate, followed by MOC-161 and 323/A3. The V-genes of the two MOC hybridomas were cloned as scFv in a phage display vector and antigen-binding phage were selected by panning on recombinant antigen. The scFvs compete with the original hybridoma antibodies for binding to antigen and specifically bind to human carcinomas in immunohistochemistry. MOC-31 scFv has an off-rate which is better than those of the bivalent 17-1A and 323/A3 whole antibodies, providing it with an essential characteristic for tumour retention in vivo. The availability of these high-affinity anti-EGP-2 antibody fragments and of their encoding V-genes creates a variety of possibilities for their future use as tumour-targeting vehicles

Characterisation and internalisation of recombinant humanised HMFG-1 antibodies against MUC1

The humanised HMFG-1 immunoglobulin has been extensively developed as a clinical immunotherapeutic agent for MUC1 expressing tumours. We have constructed a single-chain Fv (scFv) and Fab fragment from this antibody and shown that both these species retain their specificity for MUC1. The scFv was less stable and less soluble than the Fab. Detailed analyses of the binding kinetics of the whole IgG and Fab fragment show that the affinity for MUC1 synthetic peptides is low (approximately 100 n for the IgG and 10 μ for the Fab), with particularly low but similar dissociation rate constants (0.031–0.095 s−1). Binding to native antigen on the cell surface is over two orders of magnitude better. Confocal immunofluorescence microscopy shows that both the IgG and Fab are internalised rapidly (the IgG is internalised within 15 min) and colocalise to early endosomes. This work provides an appreciation of the binding, internalising and trafficking kinetics, important for the development of future therapeutics based on this antibody

Rise and Fall of an Anti-MUC1 Specific Antibody

So far, human antibodies with good affinity and specificity for MUC1, a transmembrane protein overexpressed on breast cancers and ovarian carcinomas, and thus a promising target for therapy, were very difficult to generate.A human scFv antibody was isolated from an immune library derived from breast cancer patients immunised with MUC1. The anti-MUC1 scFv reacted with tumour cells in more than 80% of 228 tissue sections of mamma carcinoma samples, while showing very low reactivity with a large panel of non-tumour tissues. By mutagenesis and phage display, affinity of scFvs was increased up to 500fold to 5,7×10(-10) M. Half-life in serum was improved from below 1 day to more than 4 weeks and was correlated with the dimerisation tendency of the individual scFvs. The scFv bound to T47D and MCF-7 mammalian cancer cell lines were recloned into the scFv-Fc and IgG format resulting in decrease of affinity of one binder. The IgG variants with the highest affinity were tested in mouse xenograft models using MCF-7 and OVCAR tumour cells. However, the experiments showed no significant decrease in tumour growth or increase in the survival rates. To study the reasons for the failure of the xenograft experiments, ADCC was analysed in vitro using MCF-7 and OVCAR3 target cells, revealing a low ADCC, possibly due to internalisation, as detected for MCF-7 cells.Antibody phage display starting with immune libraries and followed by affinity maturation is a powerful strategy to generate high affinity human antibodies to difficult targets, in this case shown by the creation of a highly specific antibody with subnanomolar affinity to a very small epitope consisting of four amino acids. Despite these "best in class" binding parameters, the therapeutic success of this antibody was prevented by the target biology

The factor structure of the peer-feedback orientation scale (PFOS): toward a measure for assessing students' peer-feedback dispositions

This study reports on the quantitative findings of an exploratory sequential mixed methods study in which the underlying factor structure of students’ peer-feedback orientation (i.e. openness to provide and receive peer-feedback) was investigated. Building on the qualitative findings of a previous study in which the ‘peer-feedback orientation’ concept was introduced, an online survey was developed to collect data among higher education students (N = 148). An exploratory factor analysis produced a five-factor solution including the dimensions: accountability, communicativeness, utility, self-efficacy and receptivity. The practical value of the results lies in having a measure of students’ peer-feedback orientation that provides teachers and researchers with an instrument for understanding students’ dispositions toward receiving and providing peer-feedback

Report on models and guidelines for the collaborative design, development and delivery of SLPs

This document refers to deliverable 6.3 on the development of models and guidelines for the collaborative development and delivery of short learning programs and related mobility. Originally started with an introductory document, based on experience and expertise with joint Erasmus Mundus programmes, the Networked Curriculum Project (NETCU) and the work on the EADTU Mobility Matrix, this document builds further on the experience of the pilots in the ESLP project (WP6) and from other work packages, referring in particular to the comprehensive SLP design guidelines that have been developed (WP4).nonPeerReviewe

Report on collaborative SLPs and related mobility

This report (D6.2) outlines the work carried in Work Package (WP) 6 of this project E-SLP, where 5 actual SLP pilots were designed, implemented and delivered to students. This report builds on the cooperation model described in D6.1, and the work done in WP2 "Concept and role of SLPs in European HE” and WP3 “Institutional policies for SLP’s”. Furthermore D.6.2 describes the methodology used to design each pilot, detailing its target group, study level, contents and programme characteristics. The lessons learned from these pilots will be detailed in D.6.3 “Models and guidelines for the collaborative development and delivery of SLPs and related mobility”, which add the experience level to report D.2.1.nonPeerReviewe