Calcium dependent protein kinases and Malaria: Identification of chemical start point for drug discovery

Symposium: Academic Drug Discovery: Challenges and perspectives

Imperial College London, March 2011

Oliver Billker & Katie Chapman

Antimalarial drugs

• No longer useful against P. falciparum

• Wide spread resistance

ArtemisininChloroquine

• First-line treatment of P. falciparum

• Component of a combination.

• Reduced efficacy observed in SE Asia.

Important to develop a well supplied pipeline of new antimalarials.

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.

A paradigm shift in antimalarial screening

Cell based screens

Libraries of >20.000starting

points for drug

development

https://www.ebi.ac.uk/chembl/

Target based

screening

Genetic tools for target identification at scale(Sanger team)

Identify mechanism of induced resistance.

Screen against validated targets

Prioritise by chemistry, IP, etc.

2 million GSK compounds

13533 inhibitors of P. falciparum growth IC50 < 1 µM

242 inhibitors P. falciparum CDPK 1, 4 or 5

Nature, March 2010

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.

Comparative analysis of Plasmodium kinomes

Different in P. falciparumRedundant in P. bergheiRita Tewari

calmodulin-likedomain

kinase domain

Calcium dependent protein kinases (CDPKs)

• Plant-like kinases with a unique activation mechanism.

Billker & Doerig 2010

Target selection

Redundant in P. berghei

CamK

Plant like Opportunity for selectivityFamily Opportunity for multi-target inhibitor.

CDPK1 and 5:Essential in blood stages.

CDPK1 and 4Essential for transmission.

CDPK5

Predicted CDPK functions

CDPK1

CDPK4

CDPK1

CDPK4

CDPK1

CDPK1

PLC

IP3

PIP2

ER

Ca2+

Ca2+

Ca2+

PLC

C D PK 4

Receptor

Xanthurenic acid Temperature

Host cell lysis

Male gamete formation

Differential gene expression

N

OH

OHCOOH

Guanlyl

cyclasecGMPPKG

Phospho-diesterases

Map-2

CDPK4 is required for male gamete formation

SRPK

PKG

Pic

eata

nnol

Res

vera

tol

PP

2P

P1

Que

rcet

inP

urfa

lcam

ine .

RO

3188

20S

taur

ospo

rine ..

1-N

M P

P1

3-B

R P

P1

1-N

A P

P1

3-M

B P

P1

3-1B

PP

1P

HA

665

752

Mex

iletin

e H

Cl

1/pI

C50

2

3

4

5

6

7

8

9

10

CDPK1CDPK4

Comparative profiling of recombinant PfCDPK1 and 4

unselective C1 selective C4 selective

Katie Chapman, Imperial College Drug Discovery

cdpk4

CDPK4 is required for male gamete formation

DMSO 1NM-PP1

DMSO Rm-1-132 1NM-PP1Ave

rage

num

ber o

f ooc

ysts

0

10

20

30

40

50

DMSO 1NM-PP1

The CDPK4 inhibitor 1NM-PP1 blocks parasite transmission to mosquitoes

Lotta Burström

P. berghei CDPK1-GFP is expressed throughout life cycle

ookinete

gametocytes

oocyst

sporozoitesSarah Sebastian

schizont

cdpk1 knock down blocks development…

WT ookinete

zygote ookineteretort

Sarah Sebastian

1 20 120 0.05 2,400

2 35 275 0.09 3,000

average oocysts/midgutfeed n WT CDPK1 fold change

…and prevents transmission of P. berghei to mosquitoes

Sarah Sebastian

Target prioritisation: CDPK5 is also essential for blood stage development.

Science 238 (2010)

• 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.

Target summary

Aim: Screen P. falciparum versions of all three targets to explore feasibility of pan-CDPK inhibition