Symposium: Academic Drug Discovery: Challenges and perspectives

1. Calcium dependent protein kinases and Malaria: Identification of chemical start point for drug discovery Oliver Billker & Katie Chapman Symposium: Academic Drug Discovery: Challenges and perspectives Imperial College London, March 2011

2. Antimalarial drugs Artemisinin Chloroquine • First-line treatment of P. falciparum • Component of a combination. • Reduced efficacy observed in SE Asia. • No longer useful against P. falciparum • Wide spread resistance Important to develop a well supplied pipeline of new antimalarials.

3. Antimalarial drug development • Antimalarials are not economically attractive to the pharmaceutical industry. • Public-private partnerships play a key role. • E.g. Medicines for Malaria Venture, Wellcome Trust, Bill & Melinda Gates Foundation Opportunity and need for academic institutions to make an impact.

4. A paradigm shift in antimalarial screening Prioritise by chemistry, IP, etc. Target basedscreening Libraries of >20.000 starting points for drug development Identify mechanism of induced resistance. Cell based screens Screen against validated targets Genetic tools for target identification at scale (Sanger team) https://www.ebi.ac.uk/chembl/

5. Nature, March 2010 13533 inhibitors of P. falciparum growth IC50 < 1 µM 242 inhibitorsP. falciparum CDPK 1, 4 or 5 2 million GSK compounds

6. Systematic prioritisation of parasite kinase targets in a malaria parasite of rodents • Gene deletion analysis of 65 protein kinases. • Kinomes are highly conserved across Plasmodium species. • >1/3 of parasite protein kinases are redundant in blood stages.

7. Comparative analysis of Plasmodium kinomes Redundant in P. berghei Different in P. falciparum Rita Tewari

8. Calcium dependent protein kinases (CDPKs) • Plant-like kinases with a unique activation mechanism. calmodulin-likedomain kinase domain Billker & Doerig 2010

9. Target selection Plant like  Opportunity for selectivity Family  Opportunity for multi-target inhibitor. CamK CDPK1 and 5: Essential in blood stages. CDPK1 and 4 Essential for transmission. Redundant in P. berghei

10. Predicted CDPK functions CDPK4 CDPK1 CDPK1 CDPK5 CDPK1 CDPK4 CDPK1

11. CDPK4 is required for male gamete formation Guanlyl cyclase PKG Phospho-diesterases cGMP PKG SRPK Map-2 Differential gene expression Male gamete formation Host cell lysis

12. Comparative profiling of recombinant PfCDPK1 and 4 unselective C1 selective C4 selective Katie Chapman, Imperial College Drug Discovery

13. CDPK4 is required for male gamete formation cdpk4

14. The CDPK4 inhibitor 1NM-PP1 blocks parasite transmission to mosquitoes DMSO 1NM-PP1 DMSO 1NM-PP1 Lotta Burström

15. P. berghei CDPK1-GFP is expressed throughout life cycle schizont ookinete oocyst gametocytes sporozoites Sarah Sebastian

16. cdpk1 knock down blocks development… ookinete zygote retort WT ookinete Sarah Sebastian

17. …and prevents transmission of P. berghei to mosquitoes average oocysts/midgut feed n WT CDPK1 fold change 1 20 120 0.05 2,400 2 35 275 0.09 3,000 Sarah Sebastian

18. Target prioritisation: CDPK5 is also essential for blood stage development. Science 238 (2010)

19. Target summary • CDPK1, 4 & 5 are individually essential for parasite development at different life cycle stages. • Absence of redundancy due to different subcellular localisations (lipid modification), expression patterns, substrate preferences. • Pan-CDPK inhibitor could exploit synergies and delay emergence of resistance. Aim: Screen P. falciparum versions of all three targets to explore feasibility of pan-CDPK inhibition