FRAGMENT- BASED DRUG DESIGN Yemane Mengistu Michigan State University January 30, 2008.
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Transcript of FRAGMENT- BASED DRUG DESIGN Yemane Mengistu Michigan State University January 30, 2008.
FRAGMENT- BASED DRUG DESIGN
Yemane MengistuMichigan State UniversityJanuary 30, 2008
Annual Research and Development Expense
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1980
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1996
2000
2004
Exp
ense
($
Bill
ions
)
40
80
120
160
Ann
ual N
ME
App
rova
ls
R & D Investment
NME ( New Medical Entities )Source : Pharma, FDA, Lehman :
Drug Discovery Process
Target moleculeeg. Enzymes
High-throughputScreening (HTS)
Combinatorial Chemistry Natural products
Lead &Drug
optimization
N
NR2
R3
R4
N
N
X N
H
R1
R1R1
- R2 : diverse substituentX= O, S
NHHO
HO
R2
R1
OH
Toxicity Clinical trials
Creating a Library
NH
O
ON
O
OHOH
HO H
O
H
HN
OH
OHOH
O
R
O
NH
HO
HO
HO
O
Tunicamycins
O
Thymidinyl nucleoside
A Natural Product
Sun, D., Lee, R.E. Tetrahedron Lett. 2005, 46, 8497-8501
NH
O
R
NO
N
R2O
R1
O
HNR3
OH
O
Thymidinyl nucleoside library
Creating a Library Using Ugi Chemistry
Creating a Library Using Ugi Chemistry
R1 OH
O
R3 H
O
R2 NH2R4 NC R1 N
NR4
O
O
H
R2
R3
Walters, W.P., Stahl, M.T., Murcko, M.A. Drug Discovery Today 1998, 3,160-178
Creating a Thymidinyl Ugi Library
Sun, D., Lee, R.E. Tetrahedron Lett. 2005, 46, 8497-8501
Si
Et
Cl
5'-azidonucleoside
N
NH
NH
O
O
R
NO
O
N3
=SiEt
Et
SnCl2/HSPh/NEt3
R1CHO
R2COOHR3NC
NH
O
O
R
NO
O
N
R2O
R1
O
HNR3
1.HF/pyridine
NH
O
O
R
NO
N
R2
O
R1
O
HNR3
R=Me, thymidinyl,R=H, 2'-deoxyuridinyl
NH
O
O
R
NO
O
H2N
OH
Et
Thymidinyl Ugi Library
CHO
CHO
CHONC
Cl
Cl
HOCHO
S CHOH3C
O CHO
CHO
CHO
HO2C
CO2H
CO2H
CO2HCO2H
S
I
CO2Me
OBr COOH
CO2H
HO2CCO2H
MeO CO2H
OHO2C
COOHF
MeOO
NC
NC
NC
NONC
NN
NNC
S NCO O
Sun, D.; Lee, R.E. Tetrahedron Lett. 2005, 46, 8497-8501
NH
O
R
NO
N
R2O
R1
O
HNR3
OH
O
High-throughput Screening (HTS)
A process of assaying a large number of compounds against biological targets.
Up to 100,000 compounds can be analyzed in a day. Robots can usually prepare and analyze many plates simultaneously.
http://www.metprog.org.uk/images/manufacturing_icon.jpg
What types of compounds become leads from an
HTS?
HTS Drug like (Rule of 5) Lead-likeness
Molecular weight 500
# Hydrogen Bond acceptors 10
Sum of N and O
# Hydrogen Bond Donors 5
Sum of NH and OH
ClogP<5
Molecular weight ~300
Fewer Hydrogen Bond Acceptors
Sum of N and O
ClogP<3
Low to high affinity for the target receptor
Lead like behaviorDrug like behavior
≤
≤
≤
Lipophilicity Lipophilicity
Congreve, M. et al. Drug Disco. Today.2003,8, 876-877Lipinski, C.A. et al. Adv.Drug Deli.Rev.1997,23,3-25.
Lipinski’s Rules (Pfizer)
0
100
200
300
400
500
600
Potency
Rel
ativ
e M
olec
ula
r M
ass
1µM 10 nM1mM
HTShits
Drugs
Lead optimization
Drug Candidates
HTS Library
A Typical Drug Discovery Cascade
Incr
ease
d ris
k of
fai
lure
HTS
HTS Hits
Hits actives
Lead series
Drug candidates
Drug
1,000,000
2000
1200
50-200
10
1
Opera ,T.I. J Comput. Aided Mol Des 2002, 16, 323-334
HTS
1996 1999 2003 2004
Compounds screened 100,000 430,000 615,000 1,050,000
Average lead potency 3,000 nM 400 nM 10 nM 10 nM
Screen success 20% 50% 58% 65%
Leads per target 1.0 1.7 1.9 2.0
GlaxoSmithKline’s HTS Scoreboard
Chemical engineering news, 2004, 82 ,23-32
Fragnomics: Fragment Based Drug Design
An approach that uses small and relatively simple molecules to make lead compounds
Fragments Lead
Target Merge andExpand
PotentialMedicinal
compounds
Look for affinity
~Enzymes, etc
MW of Average HTS Hits and Fragments
0
100
200
300
400
500
600
Potency
Rel
ativ
e M
olec
ula
r M
ass
1µM 10 nM1mM
Fragments
HTShits
Drugs
Lead optimiza
tion
Drug Candidates
Rees D.C, Congreve M, Murray C.W, Carr R .Nat. Rev. Drug Discov. 2004, 3:660
Conventional HTS approach Fragment based drug design
Fragnomics: Fragment Based Drug Design
Erlason D.A, McDowell RS, O’Brien T. J Med Chem. 2004, 47:3463-82Lewis, W.G.et al Angew. Chem. Int. Ed. Engl. 2002, 41,1053-1057
Kd>100µM
Fragnomics: Fragment Based Drug Design
Kd = 6.5
HN
N O
NH
H2N
O
OO
HN
NH
HN
O
O
OHO
NH
HN
O
O
Swayze, E.E, et al.J.Med.Chem. 2003, 46, 4232-4235
Conventional HTS approach Fragment based drug design
Kd > 100M
What Qualifies Compounds to be Fragments?
HN
N O
NH
H2N
O
OO
HN
NH
HN
O
O
Congreve, M. et al. Drug Discov.Today 2003,8, 876-877
Molecular Weight Mr ~300 Da H-bond donors (HBD) <3 H-bond acceptors (HBA) <3
OHO
NH
HN
O
O
What Qualifies Compounds to be Fragments?
N
NH2
O
Mr =200HBD= 2HBA=3
N
NH2
O
Cl
NH
N
O
FO
IC50 = 1.3mM
IC50 = 65 nM
Fragment
Lead for protein kinase inhibitor
Congreve, M. et al. Drug Discov.Today 2003,8, 876-877
Mr =456
Molecular Weight Mr ~300 Da
H-bond donors (HBD) <3
H-bond acceptors (HBA) <3
What Qualifies Compounds to be Fragments?
Clog P=1.92PSA=48.14
Clog P=3.07PSA= 77.6
Congreve, M. et al. Drug Discov.Today 2003,8, 876-877 Ertl, P.et. al. J.Med.Chem. 2000, 43,3714-3717
Clog P <3 A measure of Lipophilicity of a compound
Polar Surface Area (PSA) <60 A measure of permeability through the cell membrane.
N
NH2
O
N
NH2
O
Cl
NH
N
O
FO
Fragment
Lead for protein kinase inhibitor
Some Common Drug-Based Fragments
Hartshorn, M.J., Murray, C.W.et.al. J. Med. Chem. 2005, 48, 403-413
HN
N
N
N
N
N
HN
N
S
O
O
NH2
NH
NH2
O
NH
OH
OHN
HN N
NH
N
ON
N NH
O
Ring system from drug Heterocyclic system Side chains
Conventional HTS vs. Fragonomics Based on Central Scaffold
A library with 1 million compounds
100 X R1
100 X R2, and 100 X R3 yields
100 100 100
Variations yield a library of only 300 compounds
Carr, R, and Hann, M. Modern Drug Discov. 2002, 45-48
N N
NH
R1
HN R3
O
R2
N N
NH
R1
HN CH3
O
CH3
N N
NH
H3CHN CH3
O
R2
N N
NH
H3CHN R3
O
CH3
Conventional (HTS) Drug Design
Screen
100 Hits
~100,000,0008
104 Hypotheticalfragments
Erlanson, D.A, Hansen, K.S. Curr Opin Chem Biol. 2004, 8,399-406.
Conventional (HTS) and Fragment Based Drug Design
SiteA
10 Ligands
10 Ligands
Site B100 Hits
Synthesis of 10 4
compounds
10,100 Molecules
Screen
100 Hits
~100,000,0008
104 Hypotheticalfragments
Erlanson, D.A, Hansen, K.S. Curr Opin Chem Biol. 2004, 8,399-406.
1. Prepare set of potential binding elements with a common chemical linkage group
Maly, D.J., Choong, D.J., and Ellman, J.A. Proc.Natl.Acad.Sci.USA. 2000,97,2419-2424
Fragment Based Drug Design
1. Prepare set of potential binding elements with a common chemical linkage group
2.Screen Potential binding elements
Maly, D.J., Choong, D.J., and Ellman, J.A. Proc.Natl.Acad.Sci.USA. 2000,97,2419-2424
Fragment Based Drug Design
Fragment Based Drug Design1. Prepare set of potential binding elements with a common chemical linkage group
2.Screen Potential binding elements
3. Prepare library of all possible combinations of linked binding elements.
Maly, D.J., Choong, D.J., and Ellman, J.A. Proc.Natl.Acad.Sci.USA. 2000,97,2419-2424
Fragment Based Drug Design1. Prepare set of potential binding elements with a common chemical linkage group
2.Screen Potential binding elements
3. Prepare library of all possible combinations of linked binding elements.
4.Screen library of linked binding elements
Maly, D.J., Choong, D.J., and Ellman, J.A. Proc.Natl.Acad.Sci.USA. 2000,97,2419-2424
Application of Fragment based drug design
Maly, D.J., Choong, D.J., and Ellman, J.A. Proc.Natl.Acad.Sci.USA. 2000,97,2419-2424
1. Protein kinase inhibitors
Tyrosine kinase (Src) activate numerous signaling pathways within cells, leading to cell proliferation, differentiation , migration and metabolic changes
Src kinases have been implicated in the pathology of tumors, osteoclast-mediated Bone resorption and disorders associated with T-cell proliferation
Scapin,G. Drug Discovery today. 2002, 7,2002
Library for the Protein kinase inhibitor
NH
NOCH3
O
N OCH3
O
N OCH3
OH
N OCH3
S
N OCH3
N OCH3
OH
OH
N
NOCH3
Cl
O
N OCH3
ClCl
N OCH3
N OCH3
OH
N
S N
OO
N
OCH3
O
N OCH3
OH
1
23 4
56
9 10
78
11 12
D.J.Maly, I.C.Choong and J.A, Ellman,Proc.Natl.Acad.Sci.USA,2000,97,2419-2424
Library for the Protein kinase inhibitor
D.J.Maly, I.C.Choong and J.A, Ellman,Proc.Natl.Acad.Sci.USA,2000,97,2419-2424
Ki =41µM
NH
NOCH3
O
N OCH3
O
N OCH3
OH
N OCH3
S
N OCH3
N OCH3
OH
OH
N
NOCH3
Cl
O
N OCH3
ClCl
N OCH3
N OCH3
OH
N
S N
OO
N
OCH3
O
N OCH3
OH
1
23 4
56
9 10
78
11 12
Library of protein kinase inhibitor
D.J.Maly, I.C.Choong and J.A, Ellman,Proc.Natl.Acad.Sci.USA,2000,97,2419-2424
F
F
F
F
F
N OCH3
O
BrBr
O
N OCH3
N
OH
N OCH3
N
N OCH3
O
N OCH3
HO
HO
N OCH3
O
OO
N OCH3
S
N OCH3
N OCH3S
N
OCH3
H3CO
C6H5H2CO
N
OCH3N
OCH3
16
17
18 1920
1314 15
21 22 23 24
Ki= 40M
Fragment-Based Design : Protein Kinase Inhibition
D.J.Maly, I.C.Choong and J.A, Ellman,Proc.Natl.Acad.Sci.USA,2000,97,2419-2424
Compound IC50µM
C-Src Fyn Lyn Lck
[7] 41± 5 >1000 >1000 >1000
[16] 40± 16 64± 50 400± 170 >500
[7,16] 0.064± 0.038 5.0± 2.4 13 ± 2.4 >250
N
N OCH3
N
NO
N
O
HO
OHOH
OH
NH3CO
7 16
Entry
1
2
3
4
6
7
8
Compound
7,16, n=2
Linker c-Src IC50, M
(CH2)n
7,16, n=3
7,16, n=4
7,16, n=5
7,16, n=6
7,16, cis
7,16, trans (1R,2R)
(CH2)n
(CH2)n
(CH2)n
(CH2)n
0.0640.038
1.10.2
6.53.0
6.50.8
5.32.1
1.20.6
0.620.02
N
N N
O
HO
OH
Olinker
Correlation of linker structure with IC50 values for c-Src Inhibition
Application of Fragment Based Drug Design
2. Matrix Metalloproteinase inhibitors
Matrix Metalloproteinases is a family of zinc-dependent endopeptidases.
Implicated in a variety of diseases including arthritis and tumor metastasis.
Conventional high-throughput screening failed to get non-peptide inhibitor.
Haiduk, P.J. et al. JACS. 1997,119, 5818-5827
Application of Fragment Based Drug Design
2. Matrix Metalloproteinase Inhibitors
HO
N
HN
O
HO
N
O
HN
O
HO
Kd=17 mMKd=0.2 mM
IC50=57 nM
Haiduk, P.J. et al. JACS. 1997,119, 5818-5827 Puerta, D.T, Lewis J.A. JACS. 2004, 126, 8389
Application of Fragment Based Drug Design
2 Matrix Metalloproteinase Inhibitors
O
O
FF
F
S
O
O
O
HO
O
O
IC 50 = 0.5 nM
Wada, C.K, et al. J.Med.Chem. 2002, 45, 219-232
ABT-518 , a drug candidate in clinical trial by Abbot Pharmaceutical Company
Application of Fragment Based Drug Design
3. Thymidylate synthase (TS)
Is the sole source for production of thymidine monophosphate (dTMP). dTMP plays a central role in DNA synthesis . It has been a target for dividing cancer cells.
Banerjee D, Mayer-Kuckuk P, Capiaux G, et al. Biochim. Biophys. Acta, 2002, 1587,:164-73.
Application of Fragment based Drug Design
SHS S S SH S S S
S
NH2
S
H2N
S
NH2
S
NH2
Screen against library of Disulfide-containing small Molecules
R NH
SS
NH2
O
Erlanson, D.A, Braisted, A.C .Proc.Natl.Acad.Sci.USA. 2000, 97 ,9367–9372
TS
Site Directed Ligand Discovery for TS
Preparation of Disulfide-Containing Library Members
Parlow, J.J. & Normansell, J.E.Mol.Diversity 1995,1, 266-269
O
OH
N
O
O COOH 1,3-disopropylcarbodiimide (DIC)
O
O
N
O
O
NH
SS
H2N
O
O
THF
NH
S
S
HNO
O
OTFA
DCMNH2
S
SHN
O
ODMF
Synthesis of Sulfonyl Libraries
HNS
SNH2
OO
DCM
R S
O
O
Cl
NPoly vinyl pyridine
HNS
SNH
OO
S
O
O
R
HN
S
SNH
O
O
S
O
O
RTFA
H2N
S
SNH S
O
O
R
filteration
Erlanson, D.A, Braisted, A.C .Proc.Natl.Acad.Sci.USA. 2000, 97 ,9367–9372
Thymidylate Synthase Inhibitor
N
SO
O
N
S
O
ON
linker
linker
HN
SO
O
linker
NH
SO
O
linker
linker N
S
O
O
F
linker
N
S
O
O
linker
N
S
O
O
linker
Cl
Erlanson, D.A, Braisted, A.C .Proc.Natl.Acad.Sci.USA. 2000, 97 ,9367–9372
Selected Non selected
linker = NH
SS
O
NH2
N
O
HO
S
O
O
R
R'
N S
O
O
O
NHCO2H
CO2H
CH3
Compund R
NH
CO2H
O CO2H
NH
CO2NH2
O CO2H
NH
CO2H
O CO2NH2
NH
CO2H
O
Ki
1100
35
61
373
246
R' Ki
NH2
O
H
NH
CO2H
O
NH
N
O
NH
O
37
noncompetitive
0.33
12
> 100 only
2
3
4
5
Compound
1 6
7
8
9
10
Erlanson, D.A, Braisted, A.C .Proc.Natl.Acad.Sci.USA. 2000, 97, 9367–9372
Thymidylate Synthase Inhibitor
NS
O
O
CO2H
HOOC
COOHHN O
N
O
HN
H2N N NH
N
H
HOOC
COOHHN
O
H
NS
O
O
CO2H
HOOC
COOHHN
O
H
NS
O
O O
NHCOOH
MethylenetetrahydrofolateKm= 14 M
N-tosyl-D-prolineki= 1.10.25 mM
ki= 24 7 M ki= 33040 nM
Erlanson, D.A, Braisted, A.C .Proc.Natl.Acad.Sci.USA. 2000, 97 ,9367–9372
Application of Fragment Based Drug Design
4. Cysteine Aspartyl Protease-3 ( Caspase-3)
Mediator of apoptosis ( programmed cell death).
They are responsible for the cleavage of the key cellular proteins such as cytoskeleton proteins.
Reducing the apoptotic response in diseases with dysregulated apoptosis such as myocardial infarction, stroke, traumatic brain, Alzheimer’s disease, and Parkinson diseases could benefit .
Hotchkiss, R.S. et al. Nat. Immunol. 2000, 1 , 496-501
Tethering with Extenders-dynamically Assembling Fragments
SH SRLG S SHS S S
S
H2N
S
NH2
S
NH2
S S S
xA
Caspase-3 using extender A
SH SRLG S SHS S S
S
H2N
S
NH2
S
NH2
S S S
NH2
Caspase-3 using extender B
S S SSH
Erlanson, D.A, Hansen, K.S. Curr Opin Chem Biol. 2004, 8,399-406.
SH
SRLGxA
S SH
OHN
S
Cl
Cl O
O
HOOCO
O
OHN
S
Cl
Cl O
O
HOOC
O
Extender A Extender B
Casp SHN
SH
O
HOOCO
Caspase-3
S
OO
Caspase-3
Casp SHN
S
O
HOOC
O
SH
O
Assembly of the Extender with Enzyme and with Fragment Library
Casp SHN
SH
O
HOOCO
Casp SHN
S
O
HOOC
O
SH
O
S SH
H2NS
SNH
SO O
COOH
OH
H2NS
SNH
SO O
COOH
H2NS
SNH
SO O
s SO2
FRAGMENTS
S S S
Assembly of the Extender with Enzyme and with Fragment Library
Erlanson, D.A, Lam, J.W, Wesmann ,C. Nat. Biotechnol. 2003, 21, 308-314
A
B
c
O HN
HOOC
SS
ONH
S COOH
OH
O O
H
OHN
HOOCO
NHS
O O
H
OHN
HOOCO
NHS COOH
OH
O O
H
OHN
HOOCO
NHS
O OCOOH
H
OHN
HOOCO
NHS
O OCOOH
OH
SCaspase
Extender A + Fragment A
Compound Ki( M)
2.8
15.3
>100
0.2
1
2
3
4
Erlanson, D.A, Lam, J.W, Wesmann ,C. Nat. Biotechnol. 2003, 21, 308-314
Assembling the inhibitor
Erlanson, D.A, Lam, J.W, Wesmann ,C. Nat. Biotechnol. 2003, 21, 308-314
Assembling the inhibitor
Caspase-3
Superimposition of Inhibitor 1(Gray) and compound 4 (salmon) with Capsase 3
Erlanson, D.A, Lam, J.W, Wesmann ,C. Nat.Biotechnol.2003 , 21,308-314
Summary
The use of fragment based drug design accompanied by different means of detection could increase the chance of finding new medical entities.
Site directed ligand discovery and fragment based lead discovery are still in their infancy, but the success of these emerging approaches could success.
No single technology will suffice, and the combination of HTS, site directed , and fragment-based lead discovery will likely become increasingly important.
Acknowledgements Prof. Kevin D. Walker Prof. Babak Borhan Prof. Bill Wulff Prof. Bob Hausinger Dr. Philip J. Hajduk , Abbott Laboratories
Lab members:,Mark, Irosha, Washington, Danielle, Behnaz
Friends: Khassay, Mercy, Rahman, Anil, Munmun, Luis
