C om putational

C hem istry forC rop

Protection R esearch
K laus-Jürgen Schleifer

C om putationalC hem istry & B iology

B A SF SE,Ludw igshafen

B ASF ata G lance

B ASF – The C hem icalC om pany

„ The w orld’s leading chem ical

com pany
„ O ffers intelligentsystem solutions
and high-value products
foralm ostallindustries
„ Sales 2007:€57,951 m illion
„ Incom e from operations
(EBIT)2007:€7,316 m illion
„ Em ployees
atyear-end 2007:95,175
13.09.2008 Istanbul,Som m erSchool,KJS 2

1
 B ASF´s Portfolio

C hem icals

Functional
Solutions
Plastics

Agricultural
Solutions
Perform ance
Products

O il& G as
13.09.2008 Istanbul,Som m erSchool,KJS 3

B ASF´s Portfolio

C hem icals

Functional
Solutions
Plastics

Agricultural
Solutions
Perform ance
Products

O il& G as
13.09.2008 Istanbul,Som m erSchool,KJS 4

2
 A griculturalSolutions

C rop protection

Fungicides Insecticides H erbicides O thers

againstharm ful againstharm ful againstw eeds e.g.grow th
diseases insectpests regulators

13.09.2008 Istanbul,Som m erSchool,KJS 5

D emA ands
ctive of a edi
ingr newentr
Act ive Ingr
esear ch edient
forcropfor
C rpr
opotPr
ectotectis
ion ona m ultidim ensionaltask!

Excellentefficacy
(betterthan currentm arket N egligible residues in food
standards)

Ecologically harm less N o adverse effects on w ild life

13.09.2008 Istanbul,Som m erSchool,KJS 6

3
 H ow do w e find new A ctive Ingredients?

O rganism -based com pound library M echanism -based

hits

A ctive – butw hatis optim isation Targetactivity – and

its m ode ofaction ? in the greenhouse?

greenhouse-screening H T-screening
new lead-structures &
developm entproducts
13.09.2008 Istanbul,Som m erSchool,KJS 7

R & D Process

R esearch D evelopm ent

Lead Lead Product

Identification O ptim ization D evelopm ent R egistration

Lead D evelopm ent D ossier Product

C andidate
0 1 2 3 4 5 6 7 8 9 10
Years

Î R & D costper productaround $ 250 m illion (industry average)

13.09.2008 Istanbul,Som m erSchool,KJS 8

4
 R & D Process

M olecularM odelling
Support

Lead Lead Product

Identification O ptim ization D evelopm ent R egistration

Lead D evelopm ent D ossier Product

C andidate
0 1 2 3 4 5 6 7 8 9 10
Years

Î R & D costper productaround $ 250 m illion (industry average)

13.09.2008 Istanbul,Som m erSchool,KJS 9

Issues addressed by M olecular M odelling

and C hem oinform atics

Excellentefficacy
(betterthan currentm arket N egligible residues in food
standards)

Ecologically harm less N o adverse effects on w ild life

13.09.2008 Istanbul,Som m erSchool,KJS 10

5
 Issues addressed by M olecular M odelling
and C hem oinform atics

ƒ chem oinform atics tools forphys-chem

property calculation
 selection rules forscreening libraries
 absorption & distribution effects
ƒ m olecularm odelling m ethods for
Excellentefficacy activity prediction
(betterthan currentm arket  rationalise SAR
 synthesis priorisation
standards)

Ecologically harm less N o adverse effects on w ild life

13.09.2008 Istanbul,Som m erSchool,KJS 11

Issues addressed by M olecular M odelling

and C hem oinform atics

ƒ chem oinform atics tools forphys-chem

property calculation
 selection rules forscreening libraries
 absorption & distribution effects
ƒ m olecularm odelling m ethods for
Excellentefficacy activity prediction
(betterthan currentm arket  rationalise SAR
 synthesis priorisation
standards)

Ecologically harm less N o adverse effects on w ild life

13.09.2008 Istanbul,Som m erSchool,KJS 12

6
 M olecularM odelling Techniques
Tw o M ain C ategories

Perform experim ents D e novo design

no protein 3D -structure, protein 3D -structure,

no active ligands no active ligands

no protein 3D -structure, protein 3D -structure,

active ligands active ligands
Pharm acophores D ocking & scoring
Free-energy –
3D -Q SA R calculation
Ligand-B ased Structure-B ased
D rug D esign D rug D esign
13.09.2008 Istanbul,Som m erSchool,KJS 13

M olecularM odelling Techniques

Tw o M ain C ategories

Perform experim ents D e novo design

no protein 3D -structure, protein 3D -structure,

no active ligands no active ligands

no protein 3D -structure, protein 3D -structure,

active ligands active ligands
Pharm acophores D ocking & scoring
Free-energy –
3D -Q SA R calculation
Ligand-B ased Structure-B ased
D rug D esign D rug D esign
13.09.2008 Istanbul,Som m erSchool,KJS 14

7
 3D -Q SA R M odels:
C oM SIA-M ethodology*

„ find the com m on features thatare im portantin binding to the

biologically relevantreceptor
(in the absence ofstructuralinform ation aboutthe receptor)
„ correlate 3D -structuralproperties w ith biologicalactivity

considered properties:
z electrostatic
z donor/acceptor targetactivity
(IC 50-values)
z steric
z hydrophobic

„ use the resulting m odelto predictthe activity ofnew analogues

*K lebe etal.,J.M ed.C hem .,1994,37,p.4130-4146
13.09.2008 Istanbul,Som m erSchool,KJS 15

Exam ple 1:Strobilurines -

Fungicides from Fungi

D efensive chem icals isolated from fungi

O

O O O O
O
O

K iefernzapfenrübling B uchenschleim rübling

(Strobilurus tenacellus) Strobilurin A (O udem ansiella m ucida) O udem ansin A

M ode ofaction
H+ H+ H+
Strobilurines block the fungal
C ytb *
energy production by inhibition UQ
pool
C ytc
A TP
I II IV Synthase
C ytc1
ofthe com plex IIIofthe 2e-
III
respiratory chain.
N AD H Succinate 1/2 O 2 AD P H+
N AD + Fum arate
H 2O ATP

13.09.2008 Istanbul,Som m erSchool,KJS 16

8
 3D -Q SA R M odelforStrobilurines

Input:
3D -conform ation
C hem ically diverse ligands
w ith differentactivity levels generation

structuralalignm ent

120 strobilurine analogues -8 -9

very
high
high
actact
ivit
ivi
y(
tIyC(50
IC <5010
-10 < 10) )
broad activity range (10 < IC 50 < 10-5)

13.09.2008 Istanbul,Som m erSchool,KJS 17

3D -Q SA R M odelforStrobilurines

Input:
3D -conform ation
C hem ically diverse ligands
w ith differentactivity levels generation

structuralalignm ent

calculate
property fields

property field:steric dem and

13.09.2008 Istanbul,Som m erSchool,KJS 18

9
 3D -Q SA R M odelforStrobilurines

Input:
3D -conform ation
C hem ically diverse ligands
w ith differentactivity levels generation

structuralalignm ent

calculate
property fields
C pd pIC 50 S1 S2 ... Sn E1 E2 ... En ...

2
7.2

8.5
correlate property
3 5.3 fields w ith activity
...

120 9.8

− log(IC 50 ) = y + a ⋅ S1 + b⋅ S2 + ...+ h ⋅ Sn + k ⋅ E1 + m ⋅ E2 + ...+ z⋅ E n + ...

13.09.2008 Istanbul,Som m erSchool,KJS 19

3D -Q SA R M odelforStrobilurines

Input:
3D -conform ation
C hem ically diverse ligands
w ith differentactivity levels generation

structuralalignm ent

calculate
property fields

correlate property
fields w ith activity
Training set(120 cpds):
reproduction ofexperim entaldata r2 = 0.95
3D -Q SAR m odel
leave-one-outcross-validation q2 = 0.79
13.09.2008 Istanbul,Som m erSchool,KJS 20

10
 3D -Q SA R M odelforStrobilurines

Input:
3D -conform ation
C hem ically diverse ligands
w ith differentactivity levels generation

structuralalignm ent

calculate
property fields

correlate property
fields w ith activity

Prediction ofindependenttestset: r2pred = 0.78

3D -Q SAR m odel
(32 com pounds)
13.09.2008 Istanbul,Som m erSchool,KJS 21

IC 50 Predictions via D ocking & Scoring?

PD B code:1SQ B & 1SQ P

13.09.2008 Istanbul,Som m erSchool,KJS 22

11
 IC 50 Predictions via D ocking & Scoring?

D ocking & Scoring 3D -Q SAR m odel

•docking of32 com pounds into 1SQ P •32 com pounds predicted w ith
strobilurine C oM SIA m odel
•10 com pounds w ithoutplausible poses
•r2pred = 0.78
•poorquantitative correlation w ith
IC 50 values
13.09.2008 Istanbul,Som m erSchool,KJS 23

Exam ple 2:Protox Inhibitors as

H erbicides

Protoporphyrinogen IX Prot
Bopor
iologi
phyr
calef
in IfXect

N N N N
H H H
Protox*
H H H
N N N N
*Protoporphyrinogen
O xidase

CO O H CO O H CO O H CO O H

Fe²+ M g²+

H em e bronzing C hlnecr
and orophyl
osis l
ofleaftissue

13.09.2008 Istanbul,Som m erSchool,KJS 24

12
 Scaffolds ofProtox Inhibitors

N het-5 N het-6 N

R O Xn Xn Xn

het-5 N het-6 N het C

Xn Xn Xn

O ,S

het N het N ,C N
H
Xn Xn
O ,S
H 2N

biochem icalassay:pIC 50-values from 3 to > 11

13.09.2008 Istanbul,Som m erSchool,KJS 25

Pharm acophore H ypothesis

Substrate tem plate based on C SD and Q M

b c
N N
H H
H H A /b B /c
N N
a d

Cl
O
B a d
N
A O
O Cl
O

13.09.2008 Istanbul,Som m erSchool,KJS

C SD code SO C LU R 26

13
 R esulting Alignm ent

F O
O Cl F
Cl Cl
N
F N N H
F O
N N O Br
FF N S O
O F HN
F O

318 protox-inhibitors
13.09.2008 Istanbul,Som m erSchool,KJS 27

Statistics derived from 3D -Q SA R M odel

# cpds. PC r² q²LO O (L50% O ) SD EP LO O (L50% O )

317 6 0.87 0.69 0.78
317 6 0.96 0.90 0.43
318 6 0.97 0.95 (0.91) 0.30 (0.41)

Prediction ofan externaldata set(n=20)w ith pIC 50 from 6 to 10

r²pred = 0.95 SD EP = 0.24

13.09.2008 Istanbul,Som m erSchool,KJS 28

14
 Sound SA R ?

O
F Cl F
O Cl
Cl N
N H
N O
N Br
F3C N O O
S HN
O O
F3C N

R
N N
H H
H H
N N

13.09.2008 Istanbul,Som m erSchool,KJS 29

R eaction Path ofProto-O xidation

C am adro etal.,Biochem J.,1991,277,p.17

H H

N N N N N N
H H H H
H H H H H H
N N -H - N N -H - N N
-H ⊕ -H ⊕

CO O H CO O H CO OH CO O H CO O H CO O H
-H -

H H H

N N N N N N
H H H H
H
H H H H H H ⊕
-H ⊕
N
-H ⊕
N N N N N
+H⊕

H H H

CO O H CO O H CO OH CO O H CO O H CO O H

13.09.2008 Istanbul,Som m erSchool,KJS 30

15
 A re Protox Inhibitors Transition State
A nalogues?

ƒ Q M -optim ization ofsubstrate and interm ediates 1,2 & 4

ƒ alignm entw ith protox inhibitor
 interm ediate 4 fits best

F Cl
O

N N
F S
N O
F
F

13.09.2008 Istanbul,Som m erSchool,KJS 31

A fter the 3D -Q SA R -M odelw as com pleted:

The Protox Structure!

F
F
F

Br N
N
F

Cl O
O

IN H

pdb-code:1SEZ
13.09.2008 Istanbul,Som m erSchool,KJS 32

16
 D ocking oftw o Protox Inhibitors

F
F
F

Br N
N
F

F F
Cl N
F
Cl O
S
O N
O O
F N
F N
O
O
N S
F

Cl N
F F

13.09.2008 Istanbul,Som m erSchool,KJS 33

A lignm entH ypothesis vs.D ocking Pose

F F

O N

F N
O

Cl N

binding site entrance

13.09.2008 Istanbul,Som m erSchool,KJS 34

17
 A lignm entH ypothesis vs.D ocking Pose

F F

O N

F N
O

Cl N

binding site entrance

13.09.2008 Istanbul,Som m erSchool,KJS 35

Is M odelling justnice to have ?

Strobilurines:Pyraclostrobin

N O
Cl N
O N
O
O

IC 50 = 7.4 10-10

nontreated

13.09.2008 Istanbul,Som m erSchool,KJS 36

18
 IntegralPartofthe Innovation C hain!

Protox-Inhibitors:Saflufenacil

F
F
N O
F O
O O
N
N S
H N
O
F Cl

IC 50 = 4.2 10-9

F
F
N O
nontreated K ixorTM F
N
O
O O
N S
H N
O
F Cl

13.09.2008 Istanbul,Som m erSchool,KJS IC 50 = 4.2 10-9 37

Sum m ary & C onclusion

¾ M olecularm odelling is a w ell-integrated discipline to support

lead identification and optim isation in agrochem icalresearch.
¾ 3D -Q SAR m odelling is the m ethod ofchoice forlead optim isation
in the absence ofprotein-structures.
¾ C onsistentalignm enthypothesis is required forhigh-quality m odels.
¾ R igorous validation is essentialto assess theirpredictive pow er.
¾ U ncertainties aboutbioactive conform ation & differentligand binding m odes.

Str
Prot
obi
ox-
lur
Mine-
odel
M:odel:
¾ Al
Lead
ignmst
enthypot
ructure or
hesi
iginat
s gui
esded
fromby
a subst
naturalpr
rate oductf
structur
rom
e. fungi.
¾ R esul
obust
ting
3Dal
-Qignm
SARentgi
m odelt
vesoreason
predictnew
to transi
act
tii
on-
ve stat
robi
e-l
anal
urine
ogue
anal
hypot
ogues.
hesis.
¾ 3D
X-r-
ay
Q SAR
struct
mur
odelper
e offersfor
the
m spossi
betteri
bilitny t
far
orm
getact
odelr
ivi
e-
tyint
pr
er
edi
prct
et
ion
ation.
than docking scores.
13.09.2008 Istanbul,Som m erSchool,KJS 38

19
Thank you foryour
attention!

20