1. Future Treatment of Oxidative-Stress Related Disorders with Nanotechnology Dr. Beverly A Rzigalinski Virginia College of Osteopathic Medicine Virginia Polytechnic & State University Blacksburg, VA

2. Cerium Oxide Cerium - Rare earth element Cerium atom surrounded by oxide lattice Cerium has multiple valence states Oxide lattice can have numerous “defects” Micro sized particles used in coatings, particularly catalytic converters Redox Reactions NOX

3. What is Nano - Cerium Oxide? Nanoscale materials behave differently from their micro-sized counterparts Valence states of Cerium may change more quickly Oxygen vacancies or “defects” increase dramatically at the nano scale Better redox?

4. Days Average Life Span Control Nano-Treated A Single Dose of Engineered Cerium Oxide Nanoparticles Increases Brain Cell Longevity

5. 27 DIV Organotypic brain cell culture 27 DIV Organotypic brain cell Culture treated with 10 nM Ceria Nanoparticles at 10 DIV 123 DIV Organotypic brain cell Culture treated with 10 nM Ceria Nanoparticles at 10 DIV

6. 400 200 300 100 Time (Seconds ) 100  M Glutamate 68 DIV Nano-Treated - 28 DIV Control 28 DIV Nano-Treated [Ca 2+ ]i (nM) Nanoparticle-Treated Cultures Maintain Normal Neuronal Signaling Throughout Their Extended Lifespan

7. What’s Going On?

8. O2O2 Regeneration:  Lattice Rearrangements?  Reaction with H2O?  Adsorption of OH or H+?  Other cellular reactants? O 2--x? Ce +4 Free Radical O2O2 Ce +3

9. Cerium Oxide Nanoparticles Scavenge Hydroxyl Radicals Generated by the Fenton Reaction Fenton Reaction +10 nm Ceria +7 nm Ceria +Fe-doped Ceria

10. Cerium Oxide Nanoparticles Scavenge Superoxide Radicals Irradiated riboflavin +7 nm Ceria +10 nm Ceria +Fe-doped Ceria

11. Activity (Units) Catalase SODGlutathione Peroxidase CeO 2 -Nanoparticles Have Activity SOD, GSH, And Catalase-Like Activity 10 nm 7 nm

12. Cerium Oxide Nanoparticles Protect Cells from UV-Mediated Oxidative Stress

14. Brain Injury

15. Injured Cells/mg protein (x 10 6 ) Uninjured 6.5 mm Injury (Moderate) 7.5 mm Injury (Severe) Cerium Oxide Nanoparticles Protect Organotypic Brain Cell Cultures From In Vitro Trauma (10 nM, 10 DIV ) n=16, in triplicate

16. Nano-Treated 1 hr Post Injury Injured Cells/mg protein (x 10 6 ) Uninjured 6.5 mm Injury 7.5 mm Injury Cerium Oxide Nanoparticles Protect Organotypic Brain Cell Cultures From In Vitro Trauma _ _ _ _ 4 1 2 3 Control

17. No Response  [Ca 2+ ] i 100-265 nM  [Ca 2+ ] i >265 nM No Response  [Ca 2+ ] i 100-265 nM  [Ca 2+ ] i >265 nM No Response  [Ca 2+ ] i 100-265 nM  [Ca 2+ ] i >265 nM Percentage Of Neurons Examined 0 20 40 60 80 100 120 Uninjured 24 Hr post Moderate Injury 24 Hr post Moderate Injury Nano-Treated Nanoparticles Preserve Normal Glutamate Signaling in Injured Neurons

18. AD Models

19.  Peptide + 10 nm Ceria Ceria Alone Ceria + Albumin Cerium Oxide Nanoparticles Block Amyloid-beta –Induced Free Radical Production

20. Treatment Groups Total injured cells(x10 4 ) 0 20 40 60 80 100 Control 10CeONP 100CeONP Aβ Aβ+10 CeONP Aβ+100CeONP Cerium Oxide Nanoparticles Protect Neurons from Aβ (1-42) -Induced Toxicity

21. Aβ+10nm Cerium Oxide NP (10 nM)Aβ+100nm Cerium Oxide NP ( 100 nM) Control AA

22. Drosophila Models Longevity & Parkinson’s Disease

23. Days 020406080 Percent Survival 0 20 40 60 80 100 10nM CeO2 Control Cerium Oxide Nanoparticles Increase the Life Span Of Female Drosophila and Maintain Normal Geotaxis with Aging Geotactic Response

24. Post-paraquat Time (in hours) 1357244872 Percent Survival 0 20 40 60 80 100 120 Control 1  M CeO 2 NP Cerium Oxide Nanoparticles Decrease Paraquat Toxicity In Middle-Aged Female Drosophila 5 flies lived out Normal lifespan

25. What About Mammalian Models? Mice – 90 nmols total, tail vein injection Accumulation in brain, heart, lung No metabolism? Rats – Up to 150  g injected. No toxicity noted No adverse tissue pathology

26. A New Generation of Nanopharmaceuticals Cerium Oxide - A promising agent to treat diseases associated with oxidative stress. More ground to cover….. –How much free radical scavenging is too much? –Controlled free radical scavenging? –Toxicity profile

27. NanoNeuroLab at VCOM Undergraduate Students Eric Amateis Matt Kurnick Julie Karafakis Clinical Dr. Jill Cramer Post Doctoral Dr. Shaadi Elswaifi Dr. Igor Danelison Research Associates Neeraj Singh Courie Cohen Kevin Hockey Chine Logan Collaborators Dr. Kathleen Meehan Dr. Rick Davis Dr. David Cox Nanophase Inc. Advanced Powder Tech.