EBI is an Outstation of the European Molecular Biology Laboratory. Small Molecules in Bioinformatics EBI Bioinformatics Roadshow 16th March 2011 Dusseldorf

EBI is an Outstation of the European Molecular Biology Laboratory.

Small Molecules in Bioinformatics

EBI Bioinformatics Roadshow16th March 2011

Dusseldorf

Small molecule resources at the EBI21.04.232

Agenda

• Introduction

• Small molecule resources • ChEMBL• ChEBI

• Searching and browsing

• Hands-on Exercises

Annotation of bioinformatics data

• Essential for capturing and understanding and knowledge associated with core data

• Often captured in free text, which is easier to read and better for conveying understanding to a human audience, but…

• Difficult for computers to parse• Quality varies from database to database• Terminology used varies from annotator to annotator

• Towards annotation using standard vocabularies: ontologies within bioinformatics


Small molecules participate in all processes of life

What are Small Molecules?

• A small molecule is defined as a low molecular weight organic compound.

• Most drugs are small molecules to allow passage over cell membranes and oral bioavailability.

• They are also able to bind to proteins and enzymes, thereby altering function, which can lead to a therapeutic effect.

• Small molecules are used in everyday life.

Some common

small molecules:

Amino Acids

Signalingγ-aminobutyric acid

GABA: chief inhibitory neurotransmitter in the mammalian central nervous system.

In humans, also regulates muscle tone.

• synthesized by neurons

• found mostly as a zwitterion, that is, with the carboxyl group deprotonated and the amino group protonated

• conformational flexibility of GABA is important for its biological function, as it has been found to bind to different receptors with different conformations

• GABA deficiency linked to

• anxiety disorder, depression, alcoholism

• multiple sclerosis, action tremors, tardive dyskinesia


Metabolism

Adenosine 5’-triphosphate (ATP): the

"molecular unit of currency" of intracellular

energy transfer.

• generated in the cell by energy-consuming processes, broken down by energy-releasing processes

• proteins that bind ATP do so in a characteristic protein fold known as the Rossmann fold, which is a general nucleotide-binding structural domain that can also bind the cofactor NAD

Adenosine 5'-triphosphate


Enzymes

• Enzyme inhibitors are molecules that bind to enzymes and decrease their activity.

• Many drugs are enzyme inhibitors. They are also used as herbicides and pesticides.

• Enzyme activators bind to enzymes and increase their enzymatic activity.

• Enzyme activators are often involved in the allosteric regulation of enzymes in the control of metabolism.

clavulanic acid acts as a suicide inhibitor of bacterial β-lactamase

enzymes


Pathways

http://www.genome.jp/kegg-bin/highlight_pathway?scale=1.0&map=map00231&keyword=tryptophan


Systems biology

BioModels: quantitative models of biochemical and cellular systems

tryptophan

D-enantiomer: sweet L-enantiomer: bitter


Drug types 2003 - 2009

'Small molecules' in various shades of blue (http://chembl.blogspot.com/)


Small Molecule Databases

• Small Molecule Databases can be used to:

• Investigate historical compounds and associated bioactivity data.• To give fresh insight into previously rejected drugs.

• Create Structure-Activity Relationships (SARs)

• Look at how changing a functional group can change the biological activity of a compound – before you start your own synthesis.

21.04.2313 Small molecule resources at the EBI

• Direct synthesis• Could reduce number of compounds made – if any similar

compounds have significant toxicity or unfavourable binding data, you can save time by not making analogues.

• Direct end product testing• Suggest what testing could be carried out – the database can

give you an idea of what testing has given ‘good’ (i.e. clear) results.

• Reduce number of compounds put through High Throughput Screening (HTS).


ChEBI and ChEMBL

Small molecule resources at the EBI

What is ChEBI?

• Chemical Entities of Biological Interest• Freely available• Focused on ‘small’ chemical entities (no proteins or

nucleic acids)• Illustrated dictionary of chemical nomenclature• High quality, manually annotated• Provides chemical ontology

Access ChEBI at http://www.ebi.ac.uk/chebi/


ChEBI home page


ChEBI data overview

Visualisation

caffeine1,3,7-trimethylxanthine methyltheobromine

Nomenclature

Formula: C8H10N4O2Charge: 0 Mass: 194.19

Chemical data

metaboliteCNS stimulanttrimethylxanthines

Ontology

MSDchem: CFFKEGG DRUG: D00528

Database Xrefs

Chemical Informatics

InChI=1/C8H10N4O2/c1-10-4-9-6-5(10)7(13)12(3)8(14)11(6)2/h4H,1-3H3

SMILES: CN1C(=O)N(C)c2ncn(C)c2C1=O

ChEBI – Chemical Entities of Biological Interest21.04.2319

ChEBI entry view

Chemical Structures

• Chemical structure may be interactively exploredusing MarvinView applet

• Available in formats• Image• Molfile• InChI and InChIKey• SMILES



Automatic Cross-references

What is ChEMBL?

• Database of bioactive, drug-like small molecules.• Contains 2D structures, calculated properties (logP, mol

weight, Lipinski etc)• Contains abstracted bioactivity data, e.g. binding data

and IC50, from multiple primary scientific journals• Covers about 30 years of compound synthesis and

testing• Annotated FDA-approved drugs

Access ChEMBL at https://www.ebi.ac.uk/chembldb/


ChEMBL Main Search Page


Master headline21.04.2324

Calc. properties

Drug Information

Clickable structure


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Structural Representations



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Parent and Salt Forms

Database links


ChEBI Link:


This will take you back to ChEMBL

ChemSpider Links:

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The link works both ways. They

link TO ChemSpider and

FROM ChemSpider.

They link on Standard_Inchi


Wikipedia Links:

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We also have links with Wikipedia. These also use the Standard_Inchi as the common identifier. These

links will link to the Compound Report Card in

ChEMBL.

The links are added by a ChemoBot and can be

updated with each release, if required.


STRUCTURAL REPRESENTATION


Stereoisomers

• Compounds that have same molecular formula and configuration, but differ in the 3-dimensional orientations.

• The central tetrahedral carbon has 4 different molecular groups/atoms attached. This is known as the chiral centre.


Stereoisomerism Example - Thalidomide

• Caused thousands of deformities in babies across 46 countries between 1957 and 1961.

• The R isomer is to control morning sickness but the S isomer was teratogenic.

• Sparked more tightly controlled laboratory practices across the world.


Stereoisomers

• Where known, the stereochemistry of the compound is noted in the structure and in the name.

• If a stereoisomer of an existing compound is submitted, it is given a separate id number.

• If a mixture of two stereoisomers had data submitted, we will also give this a separate id number if the activity of the compounds can not be isolated.

• If you draw a planar compound into the structure search, you will receive data on all stereoisomers.


Ofloxacin, Levofloxacin and Dextrofloxacin

• Fluoroquinolone antibiotics• Ofloxacin is a racemic (equal) mixture of Levo and Dextro

isomers.• Levofloxacin is the more active stereoisomer• Dextrofloxacin is the less active stereoisomer

• ChEMBL has data on each with separate bioactivities.


Tautomers (keto-enol form)

• Two forms readily interconvert via the migration of a hydrogen to the adjacent oxygen and the swapping of a single to a double bond, and vice versa.

• ChEMBL does not differentiate between different tautomers.• The preferred tautomeric structure is retained.

• ChEBI does differentiate and will store the separate tautomers.


Salts• About 50% of marketed drugs are combined with salts to

aid in their activity.• Some salts prevent the drug from being absorbed in the mouth.• Some salts help the drug be activated in the intestines, rather than

the stomach.

• There are approx 40,200 ChEMBL compounds with salts.

• Bioactivity data is recorded against the parent drug and against the salt.

• Therefore, it’s important to give these compounds different ChEMBL ids.


Salt Example: Morphine• Morphine can be adminstered with many different salts:

• Hydrochloride (HCl)• Sulphate (SO4)• Tartrate• Acetate• Citrate• Methobromide (MeBr)• Hyrobromide (HBr)• Hydroiodide (HI)• Lactate• Chloride (Cl)• Bitartrate


Dealing with Salts in ChEMBL

• Each compound, if in a salt form, is analysed and matched to a ‘parent’ – i.e. the base form of the compound. (Not inorganic compounds)

• For example, morphine hydrochloride (CHEMBL556578), morphine sulfate (CHEMBL422878) and morphine sulfate hydrate (CHEMBL1200603) are matched to their parent morphine (CHEMBL70)

• This relationship is shown on the interface of the compound page.

• Additionally, when you run a search for a compound, you will only be brought back the parent form in the results grid.


Parents and Salts on the Compound Page

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PARENT (compound report

page)

SALTS (with hyperlinks beneath)


• Clicking on the Bioactivity Summary pie chart will give you the bioactivity data for ALL forms of the compound

• To get salt specific bioactivity data, click on the hyperlink beneath the salt form of interest to be taken to its compound page.

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Morphine - All Data Morphine HCl specific data


Naming and Classification


Chemical names

Common or trivial names are those that are highly used.

Advantages of common names include simplicity, pronounceability and universally recognised

The main disadvantage is ambiguity – the same common name may refer to more than one type of chemical.


Systematic names

A systematic name is one which corresponds to the chemical structure such that the structure can be determined from the name, e.g. 1,2-dimethyl-naphthalene

Software packages exist which can generate structures from the systematic names (e.g. ACD/Name, ChemOffice, MarvinSketch).

More than one correct systematic name can be assigned to the same molecular structure, depending on the manner in which naming rules are applied.


Examples of common and systematic names

Common names Systematic names

caffeine

guaranine

theine

1,3,7-trimethyl-3,7-dihydro-1H-purine-2,6-

dione

7-methyltheophylline

1,3,7-trimethyl-2,6-dioxopurine


SEARCHING IN CHEBI

Why?

• Ontological data• Structure classification

• Chemical entity, e.g. hydrocarbon• Role, e.g. ligand• Subatomic particle, e.g. electron

• Links to other databases• Kegg• DrugBank• PDBEChem

• Citations

How?Text-based

Drawing

The ChEBI ontology

Organised into three sub-ontologies, namely• Molecular structure ontology

• Subatomic particle ontology

• Role ontology

(R)-adrenaline


Molecular structure ontology


Role ontology



ChEBI ontology relationships

• Generic ontology relationships

• Chemistry-specific relationships


Viewing ChEBI ontology

Simple and advanced text search

Narrow by category

Narrow by categoryAND, OR

and BUT NOT

AND, OR and BUT

NOT


Structure searchSearch optionsSearch options

Structure drawing tools

Structure drawing tools


Search Results

Click to go to compound page

Click to go to compound page

Hover-over for search menu

Hover-over for search menu


Types of structure search

• Identity – based on InChI

• Substructure – uses fingerprints to narrow search range, then performs full substructure search algorithm

• Similarity – based on Tanimoto coefficient calculated between the fingerprints

InChI=1/H2O/h1H2

10101101110010110010

1010110111

0010110010Tanimoto(a,b)

= c / (a+b-c)

= 4 / (4+7-4) = 0.57

a

b


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Browse via Periodic Table

Molecular entities / Elements

Molecular entities / Elements


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Navigate via links in ontology

Click to follow linksClick to follow links


CHEBI SEARCH EXAMPLE

ChEBI example

• Search for ‘Glycine’• What is the ChEBI ID for this?• Is it available as a Kegg compound?• What are the IUPAC names?• What is ‘glycine zwitterion’?• • 15428• Yes• Glycine, aminoacetic acid• It is a tautomer of glycine

SEARCHING IN CHEMBL


How to search in ChEMBL:

• Keywords• Compound name – dopamine, haloperidol• Assay name – cytotoxicity, liver hepatotoxicity• Target – RAF-1, IRAK-4

• Structure

• BLAST search – FASTA sequence from UniProt

• Protein or taxonomy hierarchy


Where to search:


Using the search field (found on main page):

• Best for single words• E.g. ‘dopamine’, ‘Muscarinic’

• Looks for matching text in compound name, key or synonym• 3-o-methyl-alpha-methyldopamine• Muscarinic receptor 4

• Needs an exact match• Can’t use wildcards, e.g. ‘%’, ‘?’…


Using the Protein Sequence Search

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• Useful for searching for a specific protein or a protein from the same family

• The results brought back will show a percentage similarity to the inputted sequence. • An exact match will give 100%.• Same targets but different organisms will give ~90%


Compound Drawing

• Can draw the full structure of interest or a partial structure

• Using the Substructure Search you can find compounds containing your partial structure

• Using the Similarity Search, you can find similar compounds – based on a percentage score (70-100%)


DOWNLOAD AND ANALYSIS OF CHEMBL RESULTS


• The compounds can be downloaded as an *.SDFile.


• The bioactivity data can be downloaded as *.XLS



CHEMBL WORKED EXAMPLE

STRUCTURE ACTIVITY RELATIONSHIPS


Drug design

• Ligand-based: relies on knowledge of other molecules that bind to the biological target of interest.

• Structure-based: relies on knowledge of the 3D structure of the biological target.

• A lead has• evidence that modulation of the target will have therapeutic value: e.g. disease

linkage studies showing associations between mutations in the biological target and certain disease states.

• evidence that the target is druggable, i.e. capable of binding to a small molecule and that its activity can be modulated by the small molecule.

• Target is cloned and expressed, then libraries of potential drug compounds are screened using screening assays


Drug Discovery Process

> 2,900,000 bioactivities

> 600,000 compounds

~30,000 distinct lead series

~12,000 candidates ~2000

drugs

Target Discovery

Lead Discovery

Lead Optimisatio

n

Preclinical Development

Phase 1

Phase 2

Phase 3

Launch

•Target identification•Microarray

profiling•Target

validation•Assay

development•Biochemistry

•Clinical/Animaldisease models

•High-throughputScreening (HTS)•Fragment-based

screening•Focused libraries

•Screening collection

•Medicinal Chemistry

•Structure-baseddrug design•Selectivity

screens•ADMET screens•Cellular/Animaldisease models•Pharmacokineti

cs

•Toxicology•In vivo safety pharmacology•Formulation

•Dose prediction

PK

tolerability

Efficacy

Safety

&

Efficacy

Indication

Discovery & expansion

Med. Chem. SAR Clinical Candidates

Drugs

Discovery Development Use

Clinical Trials

ChEMBL database


SAR DataSAR Data

CompoundCompoundA

ssay

Ass

ay

Ki=4.5 nM

>Thrombin MAHVRGLQLPGCLALAALCSLVHSQHVFLAPQQARSLLQRVRRANTFLEEVRKGNLERECVEETCSYEEAFEALESSTATDVFWAKYTACETARTPRDKLAACLEGNCAEGLGTNYRGHVNITRSGIECQLWRSRYPHKPEINSTTHPGADLQENFCRNPDSSTTGPWCYTTDPTVRRQECSIPVCGQDQVTVAMTPRSEGSSVNLSPPLEQCVPDRGQQYQGRLAVTTHGLPCLAWASAQAKALSKHQDFNSAVQLVENFCRNPDGDEEGVWCYVAGKPGDFGYCDLNYCEEAVEEETGDGLDEDSDRAIEGRTATSEYQTFFNPRTFGSGEADCGLRPLFEKKSLEDKTERELLESYIDGRIVEGSDAEIGMSPWQVMLFRKSPQELLCGASLISDRWVLTAAHCLLYPPWDKNFTENDLLVRIGKHSRTRYERNIEKISMLEKIYIHPRYNWRENLDRDIALMKLKKPVAFSDYIHPVCLPDRETAASLLQAGYKGRVTGWGNLKETWTANVGKGQPSVLQVVNLPIVERPVCKDSTRIRITDNMFCAGYKPDEGKRGDACEGDSGGPFVMKSPFNNRWYQMGIVSWGEGCDRDGKYGFYTHVFRLKKWIQKVIDQFGE

APTT 11 min

Target

Compound

Bioactivity


Current Data Content (ChEMBL_09)

• Abstracted from 39,094 papers from 16 journals• Ongoing curation and clean-up of all data

• 759,220 compound records• 623,012 distinct compound structures

• 8,091 targets• 4,912 protein molecular targets

• 3,030,317 experimental bioactivities • binding measurements, functional assays and ADMET


ChEMBL Assay Data

• ChEMBL contains >3 million data points relating compounds to targets or effects.

• These activities come from ~490K assays reported in medicinal chemistry literature.

• Assays can be classified as:• binding measurements

e.g., IC50

• functional assay endpointse.g., Vasodilation

• ADME/toxicity datae.g., LD50


Compound Properties and Selectivity

• Stores a wide range of calculated compound properties (e.g., mol wt, logP, RO5 violations) • Can be used to identify compounds most likely to have good in vivo

properties (Absorption, Distribution, Metabolism, Excretion)

• Contains activity information against liability targets (e.g., cytochrome P450s, HERG K+ channel)• If compounds have been tested in these assays, can avoid those

with potential toxicity issues

• Contains data on a wide range of targets• If compounds have been tested against multiple targets, can get an

idea of their selectivity (important for validation studies)


Identifying Chemical Tools

• Search ChEMBL for protein of interest• Simple text search against protein names/synonyms• Browse protein family tree• Sequence search using BLAST (can find related proteins)

• Identify compounds active against this protein• Sort/filter by relevant activity types and potency• E.g., retrieve compounds with IC50/Ki < 100nM

• Retrieve other data for these compounds• Structures, chemical properties, other activities


Example SAR

1. Run a search on RAF-1

2. Filter on all IC50 values less than 100nM

3. Run the structures through an external source, such as Pipeline Pilot, to show the most common substructures.

• This will give you an idea of what type of compounds have a good IC50 for the target RAF-1.

• You can then design a similar compound(s) based on these substructures.

Assessing selectivity

• So far we have only identified compounds that may be active against a target of interest

• Often the aim is to find compounds that are selective for that target (i.e., not active against other targets)

• Need to consider all of the available activity data for each compound to see if it is known to be active against any other targets


• Extract the list of SMILES from the XLS spreadsheet

• Run this through ChEMBL SMILES list search tool

• Filter the bioactivity for IC50 > 100nM

• Download the filtered bioactivity as another XLS spreadsheet

• Run a filter on the spreadsheet• Not RAF-1• Collect the subset of compounds that showed the specificity for

RAF-1

• Selective for RAF-1

• Selective for RAF-1 and inactive for other targets

• You can use external programs like Pipeline Pilot™ and Spotfire Decision Site™ to analyse the results.


Pipeline pilot protocol to extract all data with an IC50 of 0-100nM


IC50 vs Molecular Weight - Spotfire™


Downloads and programmatic access

Downloading ChEBI flavours

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• All downloads come in two flavours• 3 star only entries (manually annotated ChEBI

entries)• 2 and 3 star entries (manually annotated ChEBI,

ChEMBL and user submissions)


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Downloading ChEBI

• OBO file• Use on OBO-edit

• SDF File• Chemistry software compliant such as Bioclipse

• Flat file, tab delimited• Import all the data into Excel• Parse it into your own database structure

• Oracle binary dumps• Import into an oracle database

• Generic SQL insert statements• Import into MySQL or postgresql database


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The ChEBI web service

• Programmatic access to a ChEBI entry

• SOAP based Java implementation• Clients currently available in Java and perl

• Methods• getLiteEntity• getCompleteEntity and getCompleteEntityByList• getOntologyParents• getOntologyChildren and getAllOntologyChildrenInPath• getStructureSearch

• Documented at http://www.ebi.ac.uk/chebi/webServices.do.


Downloading ChEMBL

• Frequent releases (approx monthly)• SDFile • Text• MySQL• Oracle


Downloading ChEMBL


Help and Feedback

• Email addresses for support queries and feedback

• General questions and feedback on ChEMBL interface:

[email protected]• Reporting of data errors:

[email protected]

• General questions, support and feedback on ChEBI

[email protected]


Thank you

Documents

EBI is an Outstation of the European Molecular Biology Laboratory. Small Molecules in Bioinformatics EBI Bioinformatics Roadshow 16th March 2011 Dusseldorf