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Cell Cultures:
From the Bench to the Bed

9:00 Comments by Session Chairperson
Suzette Priola, Ph.D

9:05 Exploring Pathogenesis in Cell Culture
Suzette A. Priola, Ph.D., Senior Investigator, Chief, TSE/Prion Molecular
Biology Section, Laboratory of Persistent and Viral Diseases, Rocky
Mountain Laboratories, NIH
The early events that occur during TSE infection are largely unknown. Using a tissue culture system that enables us to exclusively track abnormal prion protein (PrPSc) during acute exposure of cells to TSE infectivity, we have studied the initial interaction between PrPSc and the cell. Our data suggest that early events during TSE infection may be largely strain and cell-type independent. Understanding the acute stages of TSE infection may provide new targets and strategies for TSE prevention and/or therapeutics as well as advance our knowledge of basic TSE pathogenesis.

9:30 Talk Title to Be Announced
Larisa Cervenakova, M.D., Ph.D., Senior Scientist, Transmissible Diseases Department, Holland Laboratory, American Red Cross

9:55 Clearance and Prevention of Prion Infection in Cell Culture by Anti-PrP Antibodies
Thomas M. Wisniewski, M.D., Professor, Neurology, Pathology and
Psychiatry, New York University School of Medicine

Treatment, Removal, or Inactivation

11:05 Anti-TSE Immunization in a Hamster Model
Giorgio Poli, Professor, DVM, Veterinary Pathology, University of Milan
Vaccination has been shown to be effective in mouse models for neurodegenerative conditions characterized by protein misfolding, such as Alzheimer’s disease (AD) and Transmissible Spongiform Encephalopathies (TSEs) and many different immunogens and strategies of intervention have been proposed. Here we show that the immunization of hamster with a synthetic oligopeptides PrP 119-146, corresponding to the central part of hamster prion protein, prolonged the survival time (>23 days) in animals challenged with the 263K strain of scrapie agent. The immunized hamster mounted a specific, but weak, antibody response, and developed also a specific cell-mediated response (as shown by proliferation assay on splenocytes). Moreover, the results obtained with RT-Real Time PCR showed an over-expression of mRNA for GFAP and pro-inflammatory cytokines (TNF-a, IL-1β) in brain of infected controls compared to immunized animals and healthy controls, as well as an decrease in IL-10 expression. Increased survival after challenge was also associated with reduction of cerebral lesions, glial fibrillary acid protein (GFAP) and PrP deposition (both in brain and spleen). Our results indicate that a humoral and cell mediated immune response can slow down PrPres deposition and decrease neuroinflammation; nevertheless, it is conceivable that the vaccination can activate in peripheral organs specific different and interacting mechanisms, as observed in other scrapie mouse models.

11:30 Application of PRDT Affinity Binding Technology to a Reduction of Risk of TSE
Transmission in the Blood and Biopharmaceutical Industry
Luisa Gregori, Ph.D., Assistant Director MNV Laboratory and Assistant Professor University of Maryland, BREF, University of Maryland and VA Medical Center
Transmissible spongiform encephalopathy (TSE) diseases are neurodegenerative illnesses that can be transmitted by blood transfusion. Precautionary measures against the spread of TSE through the blood supply have been implemented, but alone those measures are not sufficient. Pathogen Removal and Diagnostic Technologies, Inc. (PRDT) developed a new strategy based on the use of ligands derived from a combinatorial chemistry approach that specifically adsorb the TSE agent. After screening several million compounds, PRDT identified a ligand that when coupled in resin format reduced the TSE causative agent from blood to the limit of detection of a bioassay. This resin was incorporated into a prion reduction filter, the P-CAPTTM filter, manufactured by MacoPharma. The product has already demonstrated utility in reducing infectivity > 3log¹⁰ (brain derived infectivity in red blood cell concentrate - RBC) and > 1.2log¹⁰ (endogenous whole blood derived infectivity).

11:55 Differential Resistance of Prions Strains to Inactivation
Kurt Giles, Ph.D., Assistant Adjunct Professor, Institute for Neurodegenerative Diseases, University of California San Francisco
Understanding prion inactivation is essential for avoiding iatrogenic transmission of CJD via surgical instruments, and ensuring the safety of the food supply. Using a range of transgenic mouse models sensitive to naturally occurring and rodent-adapted prion strains, we have determined the difference in resistance to inactivation for a range of prion strains. By applying a set of rigorous statistical approaches we have produced the most comprehensive study of prion inactivation.