1. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Fighting Infection in Patients with Cystic Fibrosis Bill Schwan UW-La Crosse

2. UW-L Chemistry Microbiology Biology MYCO-PHYTE DISCOVERY GROUP a UW-La Crosse interdisciplinary initiative Chemistry Microbiology Biology University of Wisconsin–La Crosse

3. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse General procedure: collect  screen  isolate  screen  elucidate  synthesize  derivatize  screen  screen  screen myco / phyto crude extract X inactive–stop active partition & bioassay initial bioassay (other screens?) RELATIVELY RAPID (and inexpensive) MUCH SLOWER (and costly) more bio (and tox) screens synthesis analogs protect IP pre-clinical studies clinical trials MYCOPHYTE DISCOVERY GROUP active–go inactive–stop Isolate active cmpd(s) purify solve structure search literature protect IP

4. UW-L Chemistry Microbiology Biology MYCOPHYTE DISCOVERY the multidisciplinary team—these people and more! Prof William Schwan Microbiology, UW-L Prof Tom Volk Biology, UW-L Prof Marc Rott Microbiology, UW-L Prof Aaron Monte Chemistry, UW-L The WiSys Team: Mindy, Bill Maliyakal, Bethany, Lisa Jonathan Fritz, JD, MS, BS Whyte, Hirschboeck, & Dudek, S.C. Dr. Joe Toce Chemistry, UW-L Prof James Cook & M. S. Kabir Chemistry, UW-M Dr. Xueting Liu Chemistry, UW-L University of Wisconsin–La Crosse

5. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse progress / collaborative opportunities MYCOPHYTE DISCOVERY MD-1 Gm+, VRE, MRSA, TB solved: 2 active compounds noyes underway— MD-2Gm– rodssolved: 2-AQnounderwaynounderway— MD-3 Gm+, potent VRE, MRSA partial solution: complex probably———— MD-4TBsolved: grifolinno———— MD-5 potent, broad antifungal, etc purification underway ————— MD-6broad, potent purification underway ————— MD-7 N. sicca / gonorrhoeae purification underway ————— —————— —————— PROJECTACTIVITYSTRUCTURENOVEL?SYNTHESIS NOVEL ANALOGS PRE-CLINICALCLINICAL + ~300 wild mushroom species with wide variety of activities… + ongoing collaborations w/ WI Native Americans / herbal remedies… COULD SHARE CRUDE EXTRACT LIBRARY TO SCREEN FOR ADDL ACTIVITIES AND HERE!NEED HELP HERE *

6. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Screened fungal fruiting body library against bacteria that colonize the lungs of Cystic Fibrosis patients Goal: Find new antibiotic Project MD-2/F250

7. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP www.nlm.nih.gov What is Cystic Fibrosis?

8. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP What is Cystic Fibrosis? Homozygous recessive genetic disease afflicting Caucasian individuals Average lifespan of a CF patient is only 32 years Characterized by chronic bronchial infections that kill more than 90% of CF patients Antibiotic resistance in the bacteria a big problem

9. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Screened a library of 260 fungal fruiting bodies Used disk diffusion assays for screening Target organisms were Pseudomonas aeruginosa and Acinetobacter baumannii Best hit was against fungus labeled F10/F250 Expanded the assay to include Burkholderia cepacia and Stenotrophomonas maltophilia Screening Protocol

10. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Disk Diffusion Assays Against Glucose Nonfermenting Gram-negative Rods

11. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Purification of the Active Drug First, thin-layer chromatography Second, HPLC separation of active peak Elucidation of structure of active drug using Mass Spectrometry NMR Spectroscopy

12. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Structure of Active Natural Product Drug

13. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP 2-AQ MICs Against Glucose Nonfermenting Gram-negative Rods

14. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP What To Do Next? Synthesize 2-AQ Make analogs of 2-AQ Proceed with pre-clinical testing of 2-AQ More cytotoxicity testing in vitro Safety testing in mice/rats Efficacy testing in mice/rats

15. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Possible 2-AQ Analogs

16. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Synthetic Route #1 Merck Patent Approach

17. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Synthetic Route #2 Wittig-Style Approach

18. UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse MYCOPHYTE DISCOVERY GROUP Acknowledgements Mycophyte Discovery Co-Founders: Aaron Monte, Marc Rott, Joe Toce, Tom Volk UW-L Helpers: Craig Dunek, Kathleen Engelbrecht, Mike Gebhardt, Aaron McCoy, Ron McKelvey, Maria Drieta, Will Fiers, Nathan Connell Others: David Lewis, Jim Cook, Maliyakal John Monetary Assistance: UW-L, UW-System ARG and ARG-WiTAG grants

19. MYCOPHYTE DISCOVERY become a player in WI biotech? be the first startup in Western WI? …can it be done? UW-L Chemistry Microbiology Biology University of Wisconsin–La Crosse business opportunities?