Day 1 :
Hospital for Sick Children, Canada
Keynote: Cholesterol masking of membrane glycosphingolipids provides a means to evade tumour immunosurveillance and the basis of a living cell transistor
Time : 09:30-10:10
Clifford Lingwood completed his PhD at the University of London in1974, and postdoctoral studies at the Universities of Washington and Toronto. He has been a full professor at the University of Toronto since 1997 and is a senior scientist within the Molecular Medicine program, Research Institute, Hospital for Sick Children, Toronto Dr Lingwood’s research program concerns the biochemistry, chemistry, metabolism and function of glycosphingolipids with a view to the therapy of diseases in which they are involved. He has published more than 200 papers in reputed journals.
Glycosphingolipids (GSLs) and cholesterol accumulate in membrane lipid rafts and play a central role in these foci of signal transduction. However within the GSL/cholesterol complex, an H-bond network is formed which alters the conformation of the GSL carbohydrate from a membrane perpendicular, to a membrane parallel format. This latter GSL conformation is largely unavailable for exogenous ligand binding-“invisible GSLs”.
Due to the increased cholesterol levels typical of cancer cells, we studied the prevalence of cholesterol-masked GSLs in human primary tumour biopsies (prostate, neuroblastoma, colon, breast, testicular, pheochromocytoma, ovarian and ganglioneuroma). We found that such ‘invisible’ GSLs were widely present in these tumour serial cryosections, e.g SSEA1, SSEA3, SSEA4, globoH and Gb3. We propose that such masking can prevent immunosurveillance of tumour–associated GSL antigens and thereby compromise natural tumour immunity to block progression. Moreover, anti GSL Mabs in development or clinical use for treating cancer (F77 for prostate and Unituxin-antiGD2 for pediatric neuroblastoma) were highly subject to such masking, and prior tumour cholesterol extraction with b-methylcyclodextrin, resulted in a remarkable increase in antiGSL tumour staining. This suggests that tumour cholesterol depletion would increase the antineoplastic activity of these therapeutic Mabs.
We also found that the order in which the binding ligands were added was of major significance. Prior Gb3 binding promoted ligand-cholesterol binding and vice versa. This provides the means for amplification and/or diversification of GSL-dependent signal transduction and thus is the first example of a native cell based membrane “transistor”.
I.M.Sechenov First Moscow State Medical University, Russia
Keynote: Antibodies with functionality as a new generation of translational tools to monitor, to predict and to prevent demyelination
Time : 10:10-10:50
Sergey Suchkov graduated from Astrakhan State Medical University and was awarded with MD.In 1985, Suchkov obtained his Ph.D. As a Ph.D. student of the I.M. Sechenov Moscow Medical Academy and Institute of Medical Enzymology, USSR Academy of Medical Sciences, Moscow, Russia. In 2001, Suchkov finished the PostDoc Research Fellowship Program and maintained his Doctor Degree at the National Institute of Immunology, Russia. From 1987 through 1989, Dr. Suchkov was a senior Researcher, Lab of Developmental Immunology, Koltzov Institute of Developmental Biology, USSR Academy of Sciences to deal to developmental immunology. From 1989 through 1995, Dr. Suchkov was being a Head of the Lab of Clinical Immunology and Im-munobiotechnology, Helmholtz Eye Research Institute in Moscow. From 1995 through 2004, Dr. Suchkov was being a Chairman of the Department for Clinical Immunology, Moscow Clinical Research Institute (MONIKI) and the Immunologist-in-Chief of the Moscow Regional Ministry of Health. At present, Dr Sergey Suchkov, M, Ph.D., is Professor in Immunology, Department of Pathology, School for Pharmacy, I.M. Sechenov First Moscow State Medical University, Dean of the Department (Faculty) of The PPPM Development, and the First Vice-President of the University of World Business, Politics and Law and Secretary General, United Cultural Convention (UCC), Cambridge, UK.
Abs against myelin basic protein/MBP endowing with proteolytic activity (Ab-proteases) are of great value to monitor demyelination to illustrate the evolution of multiple sclerosis (MS). Anti-MBPautoAbs from MS patients and mice with EAE exhibited specific proteolytic cleavage of MBP The activity of the MBP-targeted Ab-proteases markedly differs between: (i) MS patients and healthy controls; (ii) different clinical MS courses; (iii) EDSS scales of demyelination to correlate with the disability of MS patients to predict the transformation prior to changes of the clinical course.
The sequence-specificity of Ab-proteases demonstrates five sites of preferential proteolysis to be located within the immunodominant regions of MBP confirmed by the structural databanks.Two of them falling inside the sequence covering a 81-103 peptide and its 82-98 subsegment as well, with the highest encephalitogenic properties both to act as a specific inducer of EAE and to be attacked by the MBP-targeted Ab-proteases in MS patients with the most severe (progradient) clinical courses.
Sites localized within the frame of 43-68 and 146-170 peptide subsegments whilst being less immunogenic happened to be EAE inducers very rare but were shown to be attacked by Ab-proteases in MS patients with moderate (remission-type) clinical courses.
The activity of Ab-proteases was first registered at the subclinical stages 1-2 years prior to the clinical illness. About 24% of the direct MS-related relatives were seropositive for low-active Ab-proteases from which 38% of the seropositive relatives established were being monitored for 2 years whilst demonstrating a stable growth of the Ab-associated proteolytic activity. Registration in the evolution of highly immunogenic Ab-proteases to attack 81-103 and 82-98 sites pre-dominantly would illustrate either risks of transformation of subclinical stages into clinical ones, or risks of exacerbations to develop.
The activity of Ab-proteases in combination with the sequence-specificity would confirm a high subclinical and predictive (translational) value of the tools as applicable for personalized moni-toring protocols. And close association between the proteolytic sensitivity of MBP and post-translational modifications of the latter may represent one of the key regulatory mechanisms in the epitope generation.
Ab-proteases can be programmed and re-programmed to suit the needs of the body metabolism or could be designed for the development of principally new catalysts with no natural counter-parts. By changing sequence specificity of the Ab-mediated proteolysis one may reach reduction of a density of points of the negative proteolytic effects within the myelin sheath and minimizing scales of demyelination. And, autoAb-mediated proteolysis could thus be applied to isolate from Ig molecules the efficient catalytic domains directed against particular autoimmune epitopes pa-thogenically and clinically relevant (encephalitogenic epitopes).
University of Vienna, Austria
Time : 11:10-11:50
Kristina Djinovic Carugo has completed her PhD at the age of 29 years from Ljubljana University and postdoctoral studies from University of Pavia and EMBL-Heidelberg. She is the head of the Department of Structural and Computational Biology at the University of Vienna, abd director of Laura Bassi Center for Optimised Structural Studies. The main area of her research is structural biology of actin based cystoskeleton with focus on striated muscle Z-disk. She uses an integrative structural biology approach, combining high resolution studies with complementary lower resolution biophysical and biochemical approaches. He has published more than 100 papers in international peer reviewed journals and is serving as a member of scientific advisory boards of repute.
The sarcomere is the minimal contractile unit in the cardiac and skeletal muscle, where actin and myosin filaments slide past each other to generate tension. This molecular machinery is supported by a subset of highly organised cytoskeletal proteins that fulfil architectural, mechanical and signalling functions, including the giant proteins titin, obscurin and nebulin as well as the cross-linking proteins α-actinin and myomesin.
The cross-linking of actin and myosin at the boundaries of their filamentous structures is essential for the muscle integrity and function. In the Z-disks – the lateral boundaries of the sarcomere machinery – the protein α-actinin-2 cross-links antiparallel actin filaments from adjacent sarcomeres, and additionally serves as a binding platform for a number of other Z-disk proteins. In striated muscle cells, the Z-disk represents a highly organized three-dimensional assembly containing a large directory of proteins orchestrated in a multi-protein complex centered on its major component α-actinin, with still poorly understood hierarchy and three-dimensional interaction map. On the way to elucidate the molecular structural architecture of the Z-disk, the hierarchy of its assembly and structure-function relationships, we are studying binary and higher order sub-complexes of α-actinin using biophysical, structural and cell biological approaches.
Here we will present recent data on interaction of muscle α-actinin and filamin C with an adaptor proteins ZASP, myotilin and FATZ-1, forming a fuzzy complex with the latter, and discuss findings in view of muscle Z-disk architecture and assembly.