1. Jennifer M. Finefield Robert M. Williams, Advisor Colorado State University December 6, 2011 Selective HDAC Inhibitor Novel Drug Delivery Method Biosynthetic Studies

2. Efforts Toward the Synthesis of a Selective Histone Deacetylase Inhibitor

3. DNA Packaging For DNA to fit within the nucleus, it must be condensed DNA is packaged into chromatin To begin packaging, DNA is wound around histones www.med.unc.edu www.christopher_vidal.com

4. Gene Expression: Dynamic Wrapping and Unwrapping of DNA Histone Acetyltransferase (HAT) readies DNA for transcription Histone Deacetylase (HDAC) returns DNA to the inactive state HDAC inhibitors prevent removal of acetyl residues X www.med.unc.edu www.christopher_vidal.com

5. Transcriptional Control HDAC inhibitors mimic the natural substrate Deacetylation is prevented, eventually leading to cell death

6. Zn-Dependent Histone Deacetylase Enzymes HDAC enzymes are divided into different classes Within each class, there are different isoforms Many known HDAC inhibitors display very little selectivity for class or isoform Marks, P. A., et al., Advances in Cancer Research, 2005, 137

7. HDAC Inhibitors and Enhancing Selectivity Many known HDAC inhibitors display very little selectivity for class or isoform Wiest, O. et al., J. Med. Chem. 2004, 47, 3409; Methot, J. L., et al., Bioorg. Med. Chem. Lett. 2008, 18, 973

8. Improving Selectivity of HDACi: Targeting HDAC1 and HDAC2 Moradei, O. M., et al., J. Med. Chem. 2007, 50, 5543

9. Improving Selectivity of HDACi: Targeting HDAC3 Finefield, J. M.; Williams, R. M.; Wiest, O.; Bradner, J. Unpublished results

10. Improving Selectivity of HDACi: Targeting HDAC3 Finefield, J. M.; Williams, R. M.; Wiest, O.; Bradner, J. Unpublished results Ser118, present in HDAC1/2 (green) » Tyr118 in HDAC3 (blue)

11. Key Disconnections

12. Thiazoline-Pyridine Synthesis Bowers, A., et al., Org. Lett., 2009, 11, 1301

13. Synthesis of the Amide Isostere Bowers, A., et al., J. Am. Chem. Soc., 2009, 131, 2900

14. Alternate Route

15. 2-Thiophenyl Biaryl Synthesis

16. Final Steps and Future Direction ______________________________________________________________________________________________________________________________

17. Design and Synthesis of a Novel Drug Delivery Method Specifically Targeted to Multiple Myeloma Cells

18. Multiple Myeloma MM is a plasma cell malignancy that can lead to bone destruction, anaemia, hypercalcaemia, and renal insufficiency MM is associated with older age (median age 66 years) and is found to occur more often in men than women Cause of MM remains unknown Current treatments include a single high-dose of melphalan, velcade, and various combination treatments Trialx.com, Mahindra, A., et al., Blood Reviews 2010, 24, S5; Barlogie, B., et al., Blood 2004, 103, 20

19. Tumor Specific Oligonucleotide (MB8226) UAGGCUACGUACUUAAGCG

20. The Trojan Horse Nakatani, K. et al., J. Am. Chem. Soc. 2000, 122, 2172

21. Naphthyridine Modified MM Drugs Currently undergoing clinical trials to be used as a combination treatment for multiple myeloma Given either as a high-dose treatment or as part of a combination for the treatment of multiple myeloma

22. Naphthyridine Modified Vorinostat Brown, E. V., J. Org. Chem. 1965, 30, 1607; Yoshida, M. et al., Synthesis 2008, 1099; Gediya, L. K. et al. J. Med. Chem. 2005, 48, 5047

23. Naphthyridine Modified Vorinostat Mai, A. et al. OPPI Briefs 2001, 33, 391

24. Naphthyridine Modified Melphalan Nakatani, K. et al., Bioorg. Med. Chem. 2003, 11, 2347; Gullbo, J. et al., Oncol. Res. 2003, 14, 113

25. Preliminary Test Results

27. Studies on the Biosynthesis of Reverse Prenylated Indole Secondary Metabolites from Aspergillus versicolor and Aspergillus sp. MF297-2

28. Reverse Prenylated Indole Secondary Metabolites 1969-2006

29. Proposed Biosynthesis of the Bicyclo[2.2.2]diazaoctane Ring System Porter and Sammes Diels-Alder Proposal (1970) Enzymatic Diels-Alder Reaction Porter, A. E. A. et al., Chem. Commun. 1970, 1103; Williams, R. M., Chem. Pharm. Bull. 2002, 50, 711.

30. Proposed Biosynthesis of the Bicyclo[2.2.2]diazaoctane Ring System Enzyme Controlled Stereoselectivity

31. Reverse Prenylated Indole Secondary Metabolites 2007-2011

32. Isolation of the Notoamides: New Addition to the Stephacidin Family 2007: Aspergillus sp. MF297-2 Kato, H. et al., Angew. Chem. Int. Ed. 2007, 46, 2254

33. Isolation of the Notoamides: New Addition to the Stephacidin Family 2008: Aspergillus versicolor NRRL 35600 Greshock, T. J. et al., Angew. Chem. Int. Ed. 2008, 47, 3573

34. Antipodal Natural Products Tsukamoto, S. et al., Org. Lett. 2009, 11, 1297; Greshock, T. J. et al., Angew. Chem. Int. Ed. 2008, 47, 3573

35. Isolation of the Notoamides: New Addition to the Stephacidin Family 2008-2010: Aspergillus sp. MF297-2 Tsukamoto, S. et al: JACS, 2009, 131, 3834; JNP 2008, 71, 2064; OL 2009, 11, 1297; JNP 2010, 73, 1438

36. Isolation of Notoamide E: A Potential Biosynthetic Precursor 2008-2010: Aspergillus sp. MF297-2 Tsukamoto, S. et al., J. Am. Chem. Soc. 2009, 131, 3834

37. Proposed Biosynthetic Pathway: Notoamide E Greshock, T. J. et al., Angew. Chem. Int. Ed. 2008, 47, 3573

38. Synthesis of [ 13 C] 2 -Notoamide E Tsukamoto, S. et al., JACS 2009, 131, 3834; Grubbs, A. W. et al., TL 2005, 46, 9013; Grubbs, A. W. et al., ACIE 2007, 46, 2257

39. Synthesis of [ 13 C] 2 -Notoamide E Tsukamoto, S. et al., J. Am. Chem. Soc. 2009, 131, 3834

40. [ 13 C] 2 -Notoamide E Incorporation Study with Aspergillus sp. MF297-2 No labeled bicyclo[2.2.2]diazaoctane containing metabolites were produced Tsukamoto, S. et al., J. Am. Chem. Soc. 2009, 131, 3834

41. [ 13 C] 2 -Notoamide E Incorporation Study with Aspergillus versicolor Finefield, J. M.; Williams, R. M. et al., Tetrahedron Lett. 2011, 52, 1987

42. Possible Precursors Leading to Stephacidin A

43. Synthesis of Deoxybrevianamide E and 6-Hydroxydeoxybrevianamide E Kato, H.; Nakamura, Y.; Finefield, J. M.; Umaoka, H.; Nakahara, T.; Williams, R. M.; Tsukamoto, S., TL 2011, 52, 6923

44. Synthesis of Ketopremalbrancheamide

45. Biosynthetic Breakthrough: Characterization of the (  )-Notoamide Biosynthetic Gene Cluster Ding, Y.; de Wet, J. R.; Cavalcoli, J.; Li, S.; Greshock, T. J.; Miller, K. A.; Finefield, J. M.; Sunderhaus, J. D.; McAfoos, T. J.; Tsukamoto, S.; Williams, R. M.; Sherman, D. H., JACS 2010, 132, 12733

46. Biosynthetic Breakthrough: Identification of Two Prenyltransferases Ding, Y.; de Wet, J. R.; Cavalcoli, J.; Li, S.; Greshock, T. J.; Miller, K. A.; Finefield, J. M.; Sunderhaus, J. D.; McAfoos, T. J.; Tsukamoto, S.; Williams, R. M.; Sherman, D. H., JACS 2010, 132, 12733

47. Early Steps in the Biosynthetic Pathway

48. Feeding Study with [ 13 C] 2 -[ 15 N]-6- Hydroxydeoxybrevianamide E No incorporation into advanced metabolites Finefield, J. M.; Williams, R. M. et al., JOC 2011, 76, 5954; Finefield, J. M.; Williams, R. M.; Tsukamoto, S. et al., TL 2011, 52, 6923

49. Possible Enantio-diverging Pathways from Notoamide S

50. Notoamide S Incorporation Study with Aspergillus versicolor Unlabeled synthesis of notoamide S: McAfoos, T. J. et al., Heterocycles 2010, 82, 461 Results from feeding study: Finefield, J. M.; Tsukamoto, S.; Williams, R. M. et al., unpublished results

51. Notoamide S Incorporation Study with Aspergillus sp. MF297-2 Tsukamoto, S. et al., unpublished results

52. Notoamide S: Additional Biosynthetic Insight

53. Notoamide T: Precursor Incorporation Studies ________________________________________________________________________________________________________________________________ Finefield, J. M., Tsukamoto, S.; Williams, R. M. et al., unpublished results

54. Notoamide S: Additional Biosynthetic Insight

55. Biosynthetic Precursor Incorporation Study ________________________________________________________________________________________________________________________________ Finefield, J. M.; Tsukamoto, S.; Williams, R. M. et al., Org. Lett. 2011, 13, 3802

56. Characterization of the (  )-Notoamide Biosynthetic Gene Cluster (  )-Notoamide Biosynthetic Gene Cluster (+)-Notoamide Biosynthetic Gene Cluster Li, S.; Sherman, D. H. et al., unpublished results

57. Current Postulated Biogenesis of the Notoamides and Stephacidins

58. Summary

59. Acknowledgements Prof. Robert M. Williams Williams Research Group Prof. David H. Sherman Sherman Research Group Prof. Sachiko Tsukamoto Tsukamoto Research Group Dr. James Berenson