Download - in silico small molecule discovery
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in silico small molecule discovery
SalesTargetgene
Discoverhit
Hit tolead
Optimiselead
Clinical
Target gene identified with a viable assay
High throughput screen
in silico
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Case 1 – receptor structure known
Novel in silico hits~ 100
ComputerDatabase ofMolecules100,000 +
dock moleculesinto receptor
Secondary assay? hits
IC50 < 10 µM
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How successful is this method?
• From Shoichet’s group on target – protein tyrosine phosphate 1B
• None of the in silico hits found by HTS• But unpredictable - other systems yielding < 1%
Method Compounds tested
Hits with IC50 < 10µ M Hit rate
High throughput screening (HTS)
400,000 6 0.001%
In silico docking 365 from docking
18 5%
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How does one get the receptor structure?
• X-ray structure available already at RCSB databank
• Set up a structure determination
• Predict structure
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Cloning Recombinant protein Expression
Protein purification –
mg quantities
Protein crystalsElectron density map
X-ray diffraction patternProtein structure
Crystallization
X-ray crystallography pipeline
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Prediction protein structure by homology
Query sequence
Matchedfold
Match sequence against library of known folds
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Phyre- www.sbg.bio.ic.ac.uk
Phyre and predecessor 3DPSSM > 1,000 citations
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Case 2: Ligand activity data available
Novel in silico hits
database
Observed activity Structure-activity rules
Screen
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INDDExTM –A logic-based method
• Muggleton & Sternberg developed a logic-based strategy
• Method now incorporated into INDDEx within an Imperial spin-out Equinox Pharma
• INDDEx designed to exploit availability of active and inactive data on a at least c. 5 but ideally more ligands
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Logic-rules lead to new chemotypes
C D7Å
AB B C
Fragment B is bonded to fragment C
Fragment C is bonded tofragment D
AB C D
7Å
INDDEx can learn complex rule from simpler facts Fragment A is 7Å from fragment B which is bondedto fragment C which is bonded to fragment D
Fragment A is 7Å from fragment B
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Rules can be understood by chemists
Standard programs:
Activity = 0.45 LogP + 0.56667 Lumo +1.65 V
AB C D
7Å
ILP rule:In an active molecule:Fragment A is 7Å from fragment B which is bondedto fragment C which is bonded to fragment D
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Chemistrt a
Blind trial of hit discovery on GPCR-1 Data from literature
250 novel in silico hits
Order
Observed activity- From Literature
157 Compounds30 Verified in vitro hitsNEW CHEMOTYPES Test
Cerep
in silico at Equinox
Equinox outsourced wet chemistry and biology
INDDEx
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GPCR-1: training set
Distribution of 686 training molecules collected from public domain
ActivesInactives
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GPCR Target 1 hits for optimatisation
4.7M molecules in Zinc database
400,000 drug like molecules
500 in silico hits
250 hits & new chemotypes
157 tested for inhibition
76 actives
39 for IC50
30 confirmed
30 chemotypes
30
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GPCR-1: results of primary screening
Number of in silico hits: 157 (10µM concentration)Number of actives: 76 Number of inactives: 81Primary screen success rate = 48%
10 919 22
16
81
010
2030
4050
6070
8090
Num
ber o
f hits
>70% 60%-70% 50%-60% 40%-50% 30%-40% <30%
Percent of specific binding
CB1 results - primary screening
True hitsFalse hits
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GPCR-1: new chemotypes
Distribution of hits based on their diversity (Tanimoto coefficients)
8
14
8
0
2
4
6
8
10
12
14
Num
ber o
f hits
<0.60 0.60-0.70 0.70-0.75
Tanimoto coefficient
CB1 results - new chemotype
New chemotype
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Chemistrt a
INDDEx
Equinox hit discovery on GPCR-2 - Data from BioPrint (Cerep)
250 novel in silico hits
Order
Observed activity- From BioPrint
94 Compounds28 Verified in vitro hitsTest
Cerep
in silico at Equinox
Equinox outsources wet chemistry and biology
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Confirmed hit rate of in silico predictions on secondary screen c. 35%
Target 1 Target 2In silco hits 157 94Primary screen hits(>30% binding at 10µM)
76 42
No. compounds tested for IC50 39 28IC50 results (<12µM) 30 28Estimated secondary hits if all primary hits tested
40 42
Estimated hit rate = estimated secondary hitsIn silico hits
38/157 = 24 %
42 /94= 45 %
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Comparative hit rates
Company / approach Target Hit Rate Technology
INDDEx GPCR 1 & 2 + unknown target
35 % Ligand-based
Structure-based Multiple targets Average< 2% Docking into 3D
structure
High throughput Multiple targets Average 0.001% Experimental
screening
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Concluding remarks
• If protein structure available can initiative an in silico screening approach to find hits.– Success rate generally <.2%– X-ray structure determination requires mgs of material– Prediction of structure if sequence identity > 50%
• If structure- activity data available then in silico methods can yield far better hit rates c. 35%
• in silco methods complement high throughput and can find different hits
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In silico small molecule discovery
• Michael Sternberg, Ata Amini, Paul Freemont & Michael Sternberg
• Imperial Collge Lond– www.sbg.bio.ic.ac.uk & www.doc.ic.ac.uk/~shm– www.equinoxpharma.com