Abstracts / Immunobiology 217 (2012) 1129–1222 1149 T cell costimulatory receptor. CD46 costimulation of CD4 + T cells drives the activation and differentiation of pro-inflammatory IFNg- secreting cells of the Th1 subset, but in the presence of high levels of the T cell growth factor IL2, CD46 ligation also leads to the develop- ment of a subset of anti-inflammatory Th1 T cells, which secrete the immunoregulatory cytokine IL10. We ran microRNA microarrays to study the expression of microRNAs in T cells costimulated by either CD28 or CD46. We found that CD46 costimulation leads to more rapid changes in expression of microRNAs linked to T cell activa- tion, indicating that CD46 gives a more potent costimulatory signal than CD28. We also found that proliferation of T cells, and upreg- ulation of protein markers of activation, including the high affinity alpha chain of the IL2 receptor, are greater after CD46 costimulation compared to CD28. In addition, we ran microRNA expression arrays from the FACS-sorted T cell populations, which arise after CD46 costimulation. We found several microRNAs, which are differen- tially expressed in the regulatory IL10-secreting Th1 cells compared to the pro-inflammatory IFNg-secreting Th1 T cells. The use of an antisense locked nucleic acid inhibitor of one of these microRNAs, miR-150, led to a decrease in the ratio of IL10 to IFNg secreted from cells after CD46 costimulation, suggesting that miR-150 may positively regulate IL10 secretion after CD46 costimulation. http://dx.doi.org/10.1016/j.imbio.2012.08.057 57 CFHR1 gene variants and their pathophysiological relevance Agustin Tortajada 1,2 , Maria Alba 2,3 , Sheila Pinto 1,2 , Margarita Lopez-Trascasa 2,4 , Pilar Sanchez-Corral 2,3 , Claire L. Harris 5 , San- tiago Rodriguez de Cordoba 1,2 1 Centro de Investigaciones Biologicas, Consejo Superior de Investiga- ciones Cientificas, Madrid, Spain 2 Centro de Investigacion Biomedica en Enfermedades Raras, Madrid, Spain 3 Unidad de Investigacion, Hospital Universitario La Paz, Madrid, Spain 4 Servicio de Imnunologia, Hospital Universitario La Paz, Madrid, Spain 5 Institute of Infection and Immunity, School of Medicine, Cardiff Uni- versity, Cardiff, UK Factor H related 1 (FHR-1) is the most abundant FHR protein (50 mg/ml) and that showing the highest sequence similarity with factor H. Analysis of the CFHR1 gene in our population has revealed a relatively long list of genetic variants, including single nucleotide mutations or polymorphisms, two major allotypes and several genomic rearrangements, most of them associated with pathol- ogy. Of particular interest are the two major FHR-1*A and FHR-1*B allotypes because of their differential association with AMD and aHUS and a genomic rearrangement resulting in a novel FHR-1 protein with an internal duplication of SCRs1-4, which is asso- ciated with C3-GN. To determine whether these genetic variants have functional consequences we have purified the FHR-1 proteins from the plasma of appropriate donors and determined their inter- action with C3b/iC3b/C3dg using ELISA and SPR technologies. We have also determined their capacity to compete the factor I-cofactor and convertase decay accelerating activities of factor H and have analysed their interaction with other FHR proteins and heparin. The results of these experiments and the potential mechanisms that justify their association with pathology will be described and discussed. http://dx.doi.org/10.1016/j.imbio.2012.08.058 58 CFHR5 nephropathy in a family without Cypriot ancestry Nicholas Medjeral-Thomas 1 , Talat H. Malik 1 , Tibor Toth 2 , Terry Cook 1 , Charlie Tomson 2 , Matthew C. Pickering 1 1 Centre for Complement and Inflammation Research, Imperial College, London, UK 2 Southmead Hospital, Bristol, UK C3 glomerulopathy describes glomerular pathology associated with isolated deposition of complement C3. Distinct entities within this classification include dense deposit disease and C3 glomeru- lonephritis. We have described familial C3 glomerulonephritis in association with two genomic rearrangements within the comple- ment factor H-related (CFHR) locus. In one family the condition segregated with a hybrid CFHR3-1 gene. In another study we described families from Cyprus in which the disease segregated with an internal duplication within the CFHR5 gene (CFHR5 nephropathy). This mutation resulted in an abnormal CFHR5 pro- tein which contained duplication of the first two short consensus repeat domains. Here we demonstrate that this abnormal pro- tein is associated with familial C3 glomerulonephritis in a family without Cypriot ancestry. The mutation is close to but distinct from that which we characterised in Cypriot CFHR5 nephropa- thy. The demonstration that an identical abnormal CFHR5 protein is associated with familial C3 glomerulonephritis in Cypriots and non-Cypriots provides strong genetic evidence that the change is causative. http://dx.doi.org/10.1016/j.imbio.2012.08.059 59 Characterization of a human bispecific antibody against CD20/CD55 for the treatment of burkitt lymphoma Paolo Macor 1 , Nelly Mezzaroba 1 , Luca De Maso 1 , Chiara Garrovo 2 , Stefania Biffi 2 , Daniele Sblattero 3 , Roberto Marzari 1 , Francesco Tedesco 1 1 Department of Life Sciences, University of Trieste, Trieste, Italy 2 Optical Imaging Laboratory, CBM, Area Science Park, Trieste, Italy 3 Department of Medical Sciences, University of Eastern Piedmont, Novara, Italy The effect of the antibodies in the treatment of cancer is strongly influenced by the expression of complement regulatory proteins on tumor cells. We have previously developed two minibodies neutralizing CD55 and CD59 that increased the effect of Ritux- imab in the treatment of a Hu/SCID model of Non-Hodgkin’s lymphoma. Here we have developed two bispecific molecules (CD20/55 or CD20/59) using our anti-CD55 and anti-CD59 minian- tibody together with the anti-CD20 (Rituximab) specificity. BsAbs (MB20/55 and MB20/59) were produced using an innovative sin- gle vector, purified by affinity chromatography and characterized using western-blot, ELISA and FACS analysis. BsAbs were com- pared to a minibody (MB20) with the same specificity of Rituximab for CD20, produced and purified in the same conditions of BsAbs. Complement-dependent cytotoxicity assay revealed that MB20/55 or MB20/59 had a killing effect 4 times higher that of MB20, using BJAB or MEC1 as Burkitt Lymphoma and Chronic Lymphocitic Leukemia cell lines respectively. BsAb mixture induced up to 75% of tumor cell cytotoxicity while the cell death caused by MB20 was below do not reach 10%. In vivo pharmacokinetic of BsAbs labeled with Cy5.5 was evaluated by time-domain optical imaging technol- ogy in a Hu-SCID model of Burkitt lymphoma. BsAbs distribution profile mimics the data obtained studying the pharmacokinetics