ASSIGNMENT NO.

2
PHARMACOPHORE MAPPING &
VIRTUAL SCREENING
SUBMITTED BY:
SHAGUFI NAZAR
M.PHARM( 2nd sem )

P’ceutical chemistry

Jamia hamdard

SUBMITTED TO:

Dr.GITA CHAWLA

PROFESSOR

P’ceutical chemistry

Jamia hamdard

DATE:8TH MAY 2019

CONCEPT OF PHARMACOPHORE
WHAT IS A PHARMACOPHORE?

A pharmacophore is an abstract description of molecular features which are necessary for molecular
recognition of a ligand by a biological macromolecule.

HISTORICAL PERSPECTIVE :

The original concept of the pharmacophore was developed by Paul Ehrlich during the late 1800s as
certain “chemical groups” or functions in a molecule is responsible for a biological effect, and
molecules with similar effect had similar functions in common.

The word pharmacophore was coined much later, by Schueler in his 1960 book Chemobiodynamics
and Drug Design, and was defined as “a molecular framework that carries (phoros) the essential
features responsible for a drug’s (pharmacon) biological activity.” The definition of a pharmacophore
was therefore no longer concerned with “chemical groups” but “patterns of abstract features.”

Later on since 1997, a pharmacophore has been defined by the IUPAC as: A pharmacophore is the
ensemble of steric and electronic features that is necessary to ensure the optimal supramolecular
interactions with a specific biological target and to trigger (or block) its biological response

The pharmacophore describes the essential, steric and electronic, functiondetermining points
necessary for an optimal interaction with a relevant pharmacological target. The pharmacophore
dose not represent a real molecule or a real association of functional groups, but a purely abstract
concept of compound towards their target structure.

Pharmacophores are not specific functional groups (e.g. sulphonamides) or “pieces of molecules”
(e.g. dihydropyridines, aryl piperazines). In general, a pharmacophore is a representation of common
molecular features and properties that are considered to be responsible for a desired biological
activity including;

3D (hydrophobic groups, charged/ionisable groups, hydrogen bond donors/acceptors)

2D (substructures)

1D (physical or biological)

E.g.., antihistamines

A commonly used 3D pharmacophore contains 2 aromatic rings and a tertiary nitrogen

PHARMACOPHORE MAPPING:
The process of deriving pharmacophore is known as pharmacophore mapping. Pharmacophore
Mapping is the placement of pharmacophoric features in a 3D spatial arrangement such that it aligns
perfectly with the biological molecule.

Unlike the QSAR which is focused on a set of descriptors ,pharmacophore mapping is rather a
geometrical approach. It can be thought of a 3D model of characteristic features of binding site of
investigated protein.

TWO REQUIREMENTS :

That substructure of a molecule that is responsible for its pharmacological activity (Pharmacophore).

A set of geometrical constraints between specific functional groups that enable the molecule to have
biological activity.

IT CONSIST OF THREE MAIN ASPECTS :

1. identifying common binding element that are responsible for the biological activity.

2. Determining the molecular conformation required (i.e., the bioactive conformation ).

3. Developing a superposition or alignment rule for the series of compounds.

SOFTWARES USED FOR PHARMACOPHORE MAPPING :

Discovery studio :Window ® and Linux® based protein modelling software.Produced by Accelrys
software company.Easy to use interface.

Examples of the programs that perform pharmacophore based searches are:

3D search UNITY, MACCS-3D and ROCS.

ROCS is using as shape based super position for identifying compound that have similar shaped.
CONFORMATIONAL SEARCH USED IN PHARMACOPHORE MAPPING
Conformations are different three-dimensional structures of molecules that arise from

− Rotation about single bonds (torsion angles)

− Different rings conformations

The biological activity of molecules is strongly dependent on their conformation. Conformational

analysis done by exploring the energy surface of a molecule and determining the conformation with
minimum energy.

GENERAL WORKFLOW OF A CONFORMATIONAL SEARCH:

Identification of the rotatable bonds in the molecule

Usually, only single bonds between heavy atoms are considered as rotatable and often single bonds
to terminal groups(e.g. a methyl group) are excluded.

Generation of conformations with the implemented algorithm:

no.of conformations of a molecule is determined by following formula:

Where, N= total no. of conformations.,n= no.of rotatable bonds. ,k= increments in the angle.

Checking for duplicates and very similar geometries:

identical conformation or those that are very similar have to be identified and removed.

The usually applied metric for this comparison is the RMSD value (root mean square deviation)

where N is the number of heavy atoms (non-hydrogen atoms), d is the distance between the ith
corresponding atom pair of the conformations.

Selection of a set of representative conformations:

METHOD USED FOR CONFORMATIONAL SEARCH:

1. CLIQUE DETECTION METHOD FOR PHARMACOPHORE.

2. ENSEMBLE DISTANCE GEOMETRY.

3. COMPUTIONAL METHODS : alignment or bioactive conformation method.

1. CLIQUE DETECTION METHOD: When many pharmacophoric groups are present in the molecule
it may be very difficult to identify all possible combinations of the functional groups. Clique is
defined as a 'maximal completely connected sub graph'. Clique detection algorithms can be applied
to a set of precalculated conformations of the molecules. Cliques are based upon the graph-
theoretical approach to molecular structure.

Graph G: G is not a completely connected graph, because there is not an edge between all the
nodes.

sub graph S1 is not a completely connected sub graph, because there is no edge between nodes 1
and 8

S2 is a completely connected sub-graph

S2 is not a clique, because it is not a maximal completely connected sub graph

S2 can be converted Another clique C2 into a clique C1 by adding node 8

2.ENSEMBLE DISTANCE GEOMETRY: Used to simultaneously derive a set of conformations with a

previously defined set of pharmacophoric groups overlaid. Special Feature : conformational spaces
of all the molecules are considered simultaneously.

3. COMPUTATIONAL METHOD: It includes

GASP (Genetic Algorithm Superposition Program): Genetic Algorithm Superposition Program

(GASP). GASP is the determination of the pharmacophore features: rings, donors (protons) and
acceptors (lone pairs). The atoms defined as HBA carriers can be aliphatic and aromatic nitrogen's,
alcohols, ethers, carbonyl, carboxyl oxygen's and halogens ,HBD carriers include amines and
hydroxyls.
SEAL (Steric And Electrostatic Alignment method): Steric And Electrostatic Alignment method
(SEAL). SEAL method uses steric and electrostatics features along with exhaustive searching to
compare all possible orientations

AUTOFIT program: Pharmacophore mapping attempts to find features important for receptor
binding. Therefore researchers developing alignment technique that focus on deriving and
comparing potential receptor-binding sites from ligands, rather then just using the atom of ligands.

DISCO tech: It simultaneously determine bioactive conformations and superposition rules.(i.e.,

active ligands) for the biological target of interest. An important feature of DISCO is calculation of
hydrogen bond accepting and donating sites.

IDENTIFICATION OF PHARMACOPHORE FEATURES:

In drug design, the term 'pharmacophore‘ refers to a set of features that is common to a series of
active molecules.

PHARMACOPHORE FEATURE: Every type of atom or group in a molecule that exhibits certain
properties related to molecular recognition can be reduced to a pharmacophore feature. These
molecular patterns can be labelled as hydrogen bond donors or acceptors, cationic, anionic,
aromatic, or hydrophobic regions. We will refer to such features as 'pharmacophoric groups‘.

There are three main levels of resolution for defining the features:

1.Atom-Based: One of the simplest ways to define a feature is by the 3D position of an atom,
associated with the atom type.

2.Topology-Based: the atoms are grouped into topological features like phenyl ring and carbonyl
group.

3.Function-Based: the atoms are grouped into chemical functional features that describe the kind of
interactions important for ligand-receptor binding.

The various stages involved in identification of common features of pharmacophore are :

1.Constructive stage

2.Subtractive stage

3.Optimization stage

1.The constructive stage: identifies pharmacophore candidates that are common among the most
active set of ligands. Set of maximum eight most active compounds is determined. pharmacophore
candidates consisting of up to five features between the two most active ligands are identified.
pharmacophore candidates which fit a minimum subset of features of the remaining most active
compounds are retained.
2.The subtractive stage: removes those pharmacophore candidates constructed in the previous
stage that are also present in more than half of the least active ligands.

3.The optimization stage: attempts to improve the score of the pharmacophore candidates that
pass the subtractive stage by simulated annealing.

PHARMACOPHORE MODELLING
A pharmacophore model is a hypothesis accounting for the observed biological activities of a set of
molecules that bind to a common biological target

The process for developing a pharmacophore model generally involves the following steps:

Select a training set of ligands – Structurally diverse set of molecules are chosen, that includes
both active and inactive compounds.

Conformational analysis – Generate a set of low energy conformations that is likely to contain the
bioactive conformation for each of the selected molecules.

Molecular superimposition – Superimpose ("fit") all combinations of the low-energy

conformations of the molecules. Similar (bioisosteric) functional groups common to all molecules in
the set might be fitted (e.g., phenyl rings or carboxylic acid groups). The set of conformations (one
conformation from each active molecule) that results in the best fit is presumed to be the active
conformation.

Abstraction – Transform the superimposed molecules into an abstract representation. For

example, superimposed phenyl rings might be referred to more conceptually as an 'aromatic ring'
pharmacophore element. Likewise, hydroxyl groups could be designated as a 'hydrogen-bond
donor/acceptor' pharmacophore element.

Validation – The model is only valid in so far as it is able to account for differences in biological
activity of a range of molecules.

Computational approach:

All molecular modelling studies were performed using specific catalyst.

All the structures were built and geometry optimized using CHARMm force field implemented in the
program.

The conformations were generated using the maximum limit of 255 conformations within a 20 Kcal
cut-off for the common feature pharmacophore generation using the Hip-hop module. [Smellie, A.,
et al., 1995a; Brooks, B. R., et al., 1983]
Common feature-based (Hip-hop) pharmacophore model for the synthesized quinolone molecules(
ant tubercular drugs). The Model (Hypo 1) contains eight features: three hydrophobic (cyan), four
hydrogen bond acceptor-lipid (green), and one aromatic ring (orange) & Distance between chemical
features in Å unit.

IN SILICO DRUG DESIGN & VIRTUAL SCREENING

TECHNIQUES
In Silico is an expression used to mean “performed on computer or via computer simulation.”In
Silico drug designing is defined as the identification of the drug target molecule by employing
bioinformatics tools.

Virtual screening (VS):

Computational technique.

Producing large libraries of compound that docked in to the binding site using computer
programme. The goal is finding interesting new scaffolds rather than many hits.

The aim is to score, rank or filter a set of structures using one or more computational procedures

It can be used:

to help decide which compounds to screen (experimentally)

which libraries to synthesise

lyse the results of an experiment,

such as HTS run.

TYPES:

It is of two types: ligand based( pharmacophore based )

structure based.

SIMILARITY BASED SCREENING

Chemical, pharmacological or biological properties of two compounds match. The more the common
features, the higher the similarity between two molecules.

Chemical:The two structures on top are chemically similar to each other.

Pharmacophore:The two structures above are less similar chemically (topologically) yet have the
same pharmacological activity, namely they both are Angiotensin-Converting Enzyme(ACE)
inhibitors.

Requirement for a similarity search:

A Database SQL or NoSQL ( Postgres, MySQL,MongoDB) or flat file of descriptors e.g. ChemFP

Chemical Cartridge to generate fingerprints(descriptors) for molecules ( RDKit, openbabel)

Similarity function to calculate similarity( Jaccard, Dice,Tversky) this can be written in c,c++ or
python as a function Inside SQL databases.

2D based similarity:
Based on 2D fingerprint of molecule.

Each bit in the bit string represents one molecular fragment. Typical length is ~1000 bits

Similarity is based on determining the number of bits that are common to two structures.

Calculation of similarity: Sequences/vectors of bits, or numeric values that can be compared by

distance functions, similarity metrics .

E= Euclidean distance T = Tanimoto index /Jaccard

3D based similarity:
Shape-based

- ROCS (Rapid Overlay of Chemical Structures)

flexibility

− Rigid search - based on a single conformer

− Flexible search - Conformation explored at search time

- Ensemble of conformers generated prior to search time with each conformer of each molecule
considered in turn.
OVERVIEW OF PHARMACOPHORE BASED DESIGN

PHARMACOPHORE BASED SCREENING

Usually pharmacophore based search are done in two steps.

First the software checks whether the compound has the atom types or functional groups required
by the pharmacophore,

Than its checks whether the spatial arrangement of this element matches the query.

-Flexible 3D searches identified a higher number of hits than rigid searches do.

-However flexible searches are more time consuming than rigid ones.

-There are two main approaches for including conformational flexibility in to the search

-one is top generate a user defined number of representative conformation for each molecules
when the database is to created, the other is to generate conformation during the search.

Pharmacophore model provide powerful filter tools for virtual screening even in case where the
protein structure is not available. pharmacophore filter are much faster than docking approaches,
and there for greatly reduce the number of compound subjected to the more expensive docking
application.

2-D Pharmacophore searching

Important for accelerating the drug discovery different strategies are pursued to search a 2D.
Database to identified the compound of the interest. Substructure search identified larger molecules
that contain user define query. Biochemical data obtainable from these compounds can be used for
generating structure-activity-relationship (SAR) even before synthetic plans are made for lead
optimization.
In contrast, superstructure search are used to find smaller molecules that are embedded in the
query. One problem that can arise from substructure search is that the number of the compound
identified can reach into the thousands. One solution o this problem is raking of the compound
based on similarity between compound in the database and in the query.

3-D Pharmacophore searching

1. Ligand based pharmacophore generation:

Ligand based pharmacophores are generally used when crystallographic solution structure or
moulded structure of protein cannot be obtained.

When a set of active compound is known and it is hypothesized that all the compounds bind in the
similar way to the protein, then common group should interact with the same protein residue.

2.Manual pharmacophore generation:

Manual pharmacophore generation is used when there is an easy way to identify the common
feature in a set of active compounds and/or there is experimental evidence that same functional
groups should be present in the ligand for good activity. An example is the development of a
pharmacophore model for dopamine-transporter (DAT) inhibitor. Pharmacophores should also have
some flexibility built in, thus justifying the use of distance ranges.

3.Automatic pharmacophore generation:

There are several programs Hip Hop, Hypogen, Disco, Gaps, flo,APEX, and ROCS, that can
automatically generate potential pharmacophore from a list of known inhibitors. The performance
of these programs in automated pharmacophore generation varies depending on the training set.
These all program use algorithms that identified the common pharmacophore features in the
training set molecules; they scoring function to rank the identified pharmacophores.

4.Receptor based pharmacophore generation:

If the 3D structure of receptor is known, a pharmacophore model can be derived based on the
receptor active site. Biochemical data used to identify the key residue that is important for substrate
and/or inhibiting binding. This can greatly improve the chance of finding small molecules that inhibit
the protein because the search is focused on a region of the binding side that is crucial for binding
substrate and inhibitors.
 REFERENCES
BOOK:

Foye’s principle of medicinal chemistry, 9th edition.

WEBSITES:

https://www.dovepress.com/pharmacophore-modeling-advances-limitations-andcurrent-utility-in-
dru-peer-reviewed-fulltext-article-JRLCR

http://www-oc.chemie.uniregensburg.de/OCP/ch/chv/oc22/script/glossar/Pharmacophore.pdf

http://shodhganga.inflibnet.ac.in/bitstream/10603/123674/10/chapter%209%20pharm
acophore%20modelling.pdf

SLIDES:

https://www.slideshare.net/VishakhaGiradkar1/conformational-search-used-inpharmacophore-

http://csmres.co.uk/cs.public.upd/article-downloads/2010-2011_2.pdf