1. The effect of copper chelators on Zn(-) Superoxide Dismutase Lucy Brennan Dr. Joe Beckman Dr. Kari Trumbull

2. Amyotrophic Lateral Sclerosis  Motor neuron disease characterized by progressive paralysis  10% of cases are familial  Mutations in SOD account for up to 7% of fALS  Motor neuron disease characterized by progressive paralysis  10% of cases are familial  Mutations in SOD account for up to 7% of fALS

3. Superoxide dismutase  (Cu 2+ Zn)SOD+O 2 - --> (Cu 1+ Zn)SOD+O 2 (Cu 1+ Zn)SOD +O 2 - +2H + --> (Cu 2+ Zn)SOD+H 2 O 2  Mutations linked to structural instability  Zn(-)SOD has a toxic gain of function believed to underlie fALS  (Cu 1+ )SOD +O 2 --> (Cu 2+ )SOD+ 0 2 -  (Cu 2+ Zn)SOD+O 2 - --> (Cu 1+ Zn)SOD+O 2 (Cu 1+ Zn)SOD +O 2 - +2H + --> (Cu 2+ Zn)SOD+H 2 O 2  Mutations linked to structural instability  Zn(-)SOD has a toxic gain of function believed to underlie fALS  (Cu 1+ )SOD +O 2 --> (Cu 2+ )SOD+ 0 2 -

4. Copper chelators  Dimercaptrol (BAL)  treatment for heavy metal toxicity and Wilson’s disease  Sulfhydral complex  Dimercaptrol (BAL)  treatment for heavy metal toxicity and Wilson’s disease  Sulfhydral complex

5. Goals  Validation of WST assay  To find a Cu-chelator that effectively removes copper from SOD  Modifying the Cu-chelator into a potential drug for ALS

6. WST assay

7. Superoxide detection  An assay including potassium superoxide and WST-1 was performed  Addition of KO 2 - was added to the assay  WST has been reduced

8. Xanthine assay  WST-1 + Xanthine and Xanthine oxidase  An increased absorbance was noted, indicating that WST-1 is reacting with superoxide

9. Addition of BAL with Zn(-)SOD

10. Implications  Validation of WST assay  Limitations  Insight into activity of Zn(-)SOD  Possible chelation therapy for ALS  Viable test for such compounds

11. Acknowledgements  HHMI  Kevin Ahern  Dr. Joe Beckman  Dr. Kari Trumbull  HHMI  Kevin Ahern  Dr. Joe Beckman  Dr. Kari Trumbull