1. Discovery of Neuro-regenerative Natural Products from Marine Algae and Cyanobacteria Amanda Cordes, Dr. Doug Goeger, Dr. William Gerwick

2. Background At present, there are an estimated 243,000 people living with spinal cord injuries in the United States 53% of new injuries occur within the 16-30 age group

3. Goals Identify a small molecule from algae/cyanobacteria which exhibits activity in stimulating nerve regeneration

4. Many algae compounds are similar to retinoic acid and eicosanoids A marine sponge compound, Lembehyne A, promotes neurite outgrowth Known outgrowth promoters stop cell cycle progression, as do many of the lab’s characterized compounds

5. Neuro 2a Neuroblastoma Cells: A mouse cancer cell line deriving from neurons Neuron: Cell with capability of transmitting electric signals, found in nervous system Neurite: Long, branching outgrowth from a neuron Differentiate: Cells mature, adopt distinctive functions, less likely to divide http://cancerweb.ncl.ac.uk First – Defining Some Terms

6. Neuro 2a Cells with Neurites

7. Experimental Protocol Neuro 2a cells are cultured in serum supplemented medium Cells then exposed to novel marine extracts, observed in 24 hr increments Neurite outgrowth compared against untreated control cells and ones treated with Lactacystin and with 8-Br- cAMP, known outgrowth promoters

8. Neurite Outgrowth Controls Treated Control

9. Choosing a Concentration Concentration (μg/ml) 3010310.30.1 F917419 G959064 H979695662111 I9796 6121 1454 Fraction Values are % lethality

10. 52 pure compounds 27 crude fractions One worth mentioning…..

11. …Symploca www.sitesatlas.com

12. Symploca 1454 Fractions 1454 A  30 μg/mL:0.53%  10 μg/mL:0.27% 1454 B  30 μg/mL:.32%  10 μg/mL:1.2% 1454 C  30 μg/mL:Toxic  10 μg/mL:.35%

13. Cont’d 1454 D  1 μg/mL:0.55%  0.3 μg/mL:1.5% 1454 E  10 μg/mL:1.9%  3 μg/mL:.88% 1454 F  3 μg/mL:4.0%  1 μg/mL:.48%

14. Cont’d 1454 G  1 μg/mL:3.1%  0.3 μg/mL:4.0% 1454 H  0.3 μg/mL:5.2%  0.1 μg/mL:0.9% 1454 I  0.3 μg/mL:3.2%  0.1 μg/mL:1.5%

15. Sub fractions H-1  0.003 μg/mL:4.5%  0.001 μg/mL:0.8% H-2  0.1 μg/mL:1.1%  0.03 μg/mL:1.0%

16. Baseline 1 H-1 Peak Baseline 2

17. Assay Results H-1 Peak  0.03 μg/mL:1.0%  0.01 μg/mL:1.6% Baseline 1  0.003 μg/mL:1.8%  0.001 μg/mL:0.5% Baseline 2  0.003 μg/mL:2.0%  0.001 μg/mL:0.6%

18. Possible Synergisms? 0.03 μg/mL 0.003 μg/mL

19. …..perhaps…. H-1 Peak and Baseline 1  2.3% H-1 Peak and Baseline 2  4.3%  6.5%  4.3%

20. …..results, conclusions, and future plans… Further investigation of Symploca compounds Continued screening of crude fractions and newly purified compounds

21. Acknowledgements Rebecca Medina Gerwick Lab Kevin Ahern & HHMI

22. References Blanco et al., Cell Biol. Int. 2001, 25, 909 Fenteany et al., J. Biol. Chem.1998, 273, 8545 Aoki et al., Biochem. Biophys. Res. Com. 2001, 289, 558