Day 1 :
Charles River Laboratories, USA
Keynote: Designing reproductive/juvenile animal studies to reduce animal use, while maximizing human translation
Time : 09:30-10:10
Alan M Hoberman is the Global Director of DART and Juvenile Toxicology for Charles River. He has over 40 years of experience in toxicology and is both a Diplomate of American Board of Toxicology and a Fellow of the Academy of Toxicological Sciences. He is the incoming President of the Teratology Society, the first society dedicated to the study of birth defects. He has published more than 85 peer reviewed papers and co-edited the first book on non-clinical pediatric testing.
The non-clinical studies required to support a new drug application for a large or small molecule drug generally require testing in two species (if appropriate). Approximately 65% of the animals used for these studies are used for the reproductive and developmental toxicity studies (DART). With the introduction of juvenile toxicity studies, usually in one species, the number of animals used has increased significantly. In addition most of these non-clinical studies include extra animals added for the purpose documenting exposure to the drug or the kinetic profile (exposure overtime). Over hundred litters (average about 12 fetuses per litter) and dams can be saved by combining the fertility and early embryonic study (FEED) with the embryo fetal development study (EFD). Low toxicity drugs can be tested in one rather than three studies saving over two hundred dams and litters. Provided sexually mature non-human primates (NHPs) are used in the 90 day study, male and female fertility end points can be added to these studies. In addition the EFD study in NHPs can be combined with the peri-postnatal study (PPND) to eliminate 50% of the NHPs that would be used in a standard program for a biologic or large molecule drug. Micro-sampling (30 µl or less) techniques verses normal sampling techniques (500 µL), can eliminate satellite animals. This can reduce animal usage by another 20% for DART studies and 40% for juvenile toxicity studies. Micro-sampling will also improve the scientific quality of the studies. Specific case studies will be presented.
LSU Health Science Center, USA
Time : 10:10-10:50
Rinku Majumder has completed her PhD in 1999 from Bose Institute, India and Postdoctoral studies from UNC Chapel Hill, School of Medicine in 2003. She is currently an Associate Professor in the Department of Biochemistry & Molecular Biology at LSU Health Science Center, School of Medicine. She has published more than 22 papers in reputed journals and has been serving as a standing study section Member for NIH, AHA grants. She is the Reviewer for reputed journals like Blood, JTH, JBC, Plos One, Biochemistry and Thrombosis Hemostasis.
Background: Current treatment of hemophilia-B consists of infusion of factor IX (FIX) concentrates to substitute for deficient FIX, i.e., replacement therapy. Yet, replacement is only temporary, as infused FIX is cleared rapidly from a patient’s plasma. We found that Protein S (PS) inhibits FIX and importantly, anti-PS antibody increased FIX activity in hemophilia-B plasma, implying that blocking PS activity may achieve longer lasting replacement therapy.
Aims: To assess the effectiveness of anti-PS antibody in reducing clotting time.
Method & Results: We used a modified aPTT assay (clotting initiated with FIX) with FIX-deficient plasma and varied the concentrations of added FIX and anti-PS antibody. The aPTT clotting times in the presence and absence of anti-PS antibody were measured for 2.5 (51 sec+Ab; 69 sec+Ab), 5 (43 sec+Ab; 59 seconds+Ab), 10 (38 sec+Ab; 51 sec+Ab), and 20 nM FIXa (33 sec+Ab; 40 sec+Ab). Results showed that in FIX-deficient plasma, an anti-PS antibody would make added FIXa ~3 fold more active. We also performed a thrombin generation assay with the same FIX-deficient plasma in the presence of 1, 2.5 and 5 nM FIXa and measured peak thrombin formation in the presence of anti-PS antibody. Both the thrombin generation assay and the clotting assay gave similar results, i.e., addition of neutralizing anti-PS antibody made FIX ~3 fold more active. In wild type mice we have found that PS inhibited thrombin generation. Work is underway to assess hemostasis in hemophilia-B mice receiving a low dose of FIX and PS antibodies. Improved hemostasis in this mouse model would move anti-PS antibody to the forefront as a possible adjunct in hemophilia therapy.
Conclusion: Our findings suggest that administration of anti-PS antibodies to hemophilia-B patients may achieve the goal of longer lasting replacement therapy. Antibody blocking PS activity towards FIXa is the most straightforward approach and one most likely to succeed.