Modern Method of Drug

Discovery

In Silico Drug Designing
Guided by:
Mr. Manjunath N.S.
Lecturer , Dept. of Biotechnology
Presented by:
Priya Ranjan Kumar
8
th
Sem, Dept. of Biotechnology
Background
Research based pharmaceutical companies, on average, spend about 20% of their
sales on research and development (R&D).
Despite these enormous expenditures, there has been a steady decline in the number
of drugs introduced each year into human therapy.
70-100 in the 60s
60-70 in the 70s
~50 in the 80s
~40 in the 90s

(Innovation Deficit - coined in 1996 by Jurgen Drews, president
of research at Hoffmann-LaRoche.)


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Reasons for the innovation deficit:

Increased demand on safety for drugs.
- the average number of clinical trials per new drug application (NDA) increased from
30 in the 70s to 40 in the 80s, to 70 in the 90s.
- the increased demand on safety is also reflected in a prolonged duration of the drug
development process.

In the 60s, total development time was 8.1 yrs
In the 70s, total development time was 11.8 yrs
In the 80s, total development time was 14.2 yrs
In the 90s, total development time was 14.9 yrs
Currently, total development time is ~16 yrs



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Drug Discovery & Development
Identify disease
Isolate protein
involved in
disease (2-5 years)
Find a drug effective
against disease protein
(2-5 years)
Preclinical testing
(1-3 years)
Formulation &
Scale-up
Human clinical trials
(2-10 years)
FDA approval
(2-3 years)


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Allocation of R&D time

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A new drug today costs ~$880 million and takes ~15-16 yrs to
develop.

Allocation of R&D funds

About 75% of this cost ($660 million) is attributable to failure
during development.

90% of all drug development candidates fail to make it to
market.

Methods that enhance the drug discovery process and reduce
failure rates are highly desirable!


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Impact of new technology on drug
discovery
The last few years have seen a number of revolutionary new technologies:
Gene chips, genomics and HGP
Bioinformatics & Molecular biology
More protein structures
High-throughput screening & assays
Virtual screening and library design
Docking
Combinatorial chemistry
In-vitro ADME testing
Other computational methods
How do we make it all work for us?

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Identify disease
Isolate protein
Find drug
Preclinical testing
GENOMICS, PROTEOMICS & BIOPHARM.
HIGH THROUGHPUT SCREENING
MOLECULAR MODELING
VIRTUAL SCREENING
COMBINATORIAL CHEMISTRY
IN VITRO & IN SILICO ADME MODELS
Potentially producing many more targets
and personalized targets
Screening up to 100,000 compounds a
day for activity against a target protein
Using a computer to
predict activity
Rapidly producing vast numbers
of compounds
Computer graphics & models help improve activity
Tissue and computer models begin to replace animal testing


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1) Drug Target Identification

The identification of new, clinically relevant, molecular targets is of most
importance to the discovery of innovative drugs.
It has been estimated that up to 10 genes contribute to multifactoral diseases.
Science 287:1960-1964 (2000)
Typically these disease genes are linked to another 5 to 10 gene products in
physiological circuits which are also suitable for pharmaceutical intervention.
If these numbers are multiplied with the number of diseases that pose a major
medical problem in the industrial world, then there are ~5000 to 10000 potential
drug targets.

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Contd.
Current therapy is based upon less than 500 molecular targets
45% of which are G-protein coupled receptors
28% are enzymes
11% are hormones and factors
5% ion channels
2% nuclear receptors
Therefore, many more drug targets exist! How to identify them?
Besides classical methods of cellular and molecular biology, new techniques of
target identification are becoming increasingly important. These include:
a) Genomics
b) Bioinformatics
c) Proteomics
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a) Genomics

Term was coined in the mid 80s.
Evolved from 2 independent advances:
1) Automation resulting in a significant increase in the number of experiments that
could be constructed in a given time. (eg. DNA sequencing)
2) Informatics- the ability to transform raw data into meaningful information by
applying computerized techniques for managing, analyzing, and interpreting data.
The identification of new biological targets has benefited from the genomics approach:
eg. The sequencing of the human genome
Blueprint of all proteins
Bioinformatics methods are used to transform the raw sequence into meaningful information
(eg. genes and their encoded proteins) and to compare whole genomes (disease vs. not).
Drug Discovery Today 5:135-143 2000



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b) Bioinformatics

The in silico identification of novel drug targets is now feasible by systematically
searching for paralogs (related proteins within an organism) of known drug targets
(eg. may be able to modify an existing drug to bind to the paralog).
Can compare the entire genome of pathogenic and nonpathogenic strains of a
microbe and identify genes/proteins associated with pathogenism.
Current Opin. Microbiol 1:572-579 1998
Using gene expression microarrays and gene chip technologies, a single device can
be used to evaluate and compare the expression of up to 20000 genes of healthy and
diseased individuals at once.
Trends Biotechnol 19:412-415 2001

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c) Proteomics

It is becoming increasingly evident that the complexity of biological
systems lies at the level of the proteins, and that genomics alone will
not suffice to understand these systems.
It is also at the protein level that disease processes become manifest,
and at which most (91%) drugs act.
Therefore, the analysis of proteins (including protein-protein,
protein-nucleic acid, and protein-ligand interactions) will be most
importance to target discovery.


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2) Target Validation

Involves demonstrating the relevance of the target protein in a disease process/
pathogenicity and ideally requires both gain and loss of function studies.
This is accomplished primarily with knock-out or knock-in animal models, small
molecule inhibitors/agonists/antagonists, antisense nucleic acid constructs,
ribozymes, and neutralizing antibodies.
Since strong interactions between a protein and its ligand are characterized by a
high degree of complementarity, knowledge of the protein three dimensional
structure will enable the prediction of druggability of the protein.


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3) Lead Compound Identification

Compounds are identified which interact with the target protein and
modulate its activity.
Hundreds- thousands of chemical compounds are made and tested in
an effort to find one that can achieve desirable results .
only one in about 10,000 compounds can ever reach market .
Compounds are mainly identified using random (screening) or rational
(design) approaches.



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A) High-throughput Screening

Used to test large numbers of compounds for their ability to affect the activity of
target proteins.
Natural product and synthetic compound libraries with millions of compounds are
screened using a test assay.
There are concerns with the numbers approach to screening for a lead molecule.
In theory generating the entire chemical space for drug molecules and testing
them would be an elegant approach to drug discovery.
In practice, this isnt feasible.
Therefore, concepts are needed to synthesize and select biologically relevant
compounds.



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Contd.
One solution may be to accumulate as much knowledge as possible on
biological targets (eg. structure, function, interactions, ligands) and choose
targeted approaches to chemical synthesis.
Another crucial point for reliable high-throughput screening results is the
robustness and quality of the biological test assays.

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B) Structure Based Drug Design

Three dimensional structures of compounds from virtual or physically
existing libraries are docked into binding sites of target proteins with
known or predicted structure.
Scoring functions evaluate the steric and electrostatic complementarity
between compounds and the target protein.
The highest ranked compounds are then suggested for biological testing.
Drug Discovery Today 7:64-70 2002

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Contd.
Once hits (compounds that elicit a positive response in an assay) have been identified via
the screening approach, these are validated by re-testing them and checking the purity
and structure of the compounds.
Only if the hits fulfill certain criteria are then regarded as leads. The criteria can
originate from:
1) Pharmacodynamic properties - efficacy, potency, selectivity
2) Physiochemical properties - water solubility, chemical stability,
Lipinskis rule-of-five.
3) Pharmacokinetic properties - metabolic stability and toxological aspects.
4) Chemical optimization potential - ease of chemical synthesis.
5) Patentability

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4) Lead Optimization

Molecules are chemically modified and subsequently characterized in order
to obtain compounds with suitable properties to become a drug.
Leads are characterized with respect to pharmacodynamic properties such
as efficacy and potency in vitro and in vivo, physiochemical properties,
pharmacokinetic properties, and toxicological aspects.

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Efficacy vs Potency

Potency refers to the amount of drug required for its specific effect to occur; it
is measured simply as the inverse of the EC50 for that drug.
Efficacy measures the maximum strength of the effect itself, at saturating drug
concentrations.





Drug Red exceeds drug Black in potency, while the opposite is true of the
efficacy.


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Pharmacokinetics
determining the fate of xenobiotics.
(what the body does to the drug.)
Often divided into areas examining the extent and rate of
adsorption, distribution, metabolism, and excretion (ADME).
Pharmacodynamics
determining the biochemical and physiological effects of
drugs, the mechanism of drug action, and the relationship
between drug concentration and effect.
(what the drug does to the body)
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Contd.
This process ideally requires the simultaneous optimization of multiple parameters
and is thus a time consuming and costly step.
This is often the tightest bottleneck in drug discovery.
Hints on how to modify a lead compound can originate from molecular modeling,
quantitative structure-activity relationships, and from structural biology (structure-
based drug design).
In parallel to compound characterization with respect to potency and selectivity, in
vitro assays for the prediction of pharmacokinetic properties should be performed.
Once compounds with desirable in vitro profiles have been identified, these are
characterized using in vivo models.





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5) Preclinical and Clinical Development
Preclinical studies involve in vitro studies and trials on animal populations.
Wide ranging dosages of the compounds are introduced to the cell line or animal in order
to obtain preliminary efficacy and pharmacokinetic information.
After successful completion of Preclinical studies, the drug goes through the 4 phages of
Clinical trials during which the safety and efficacy of the drug on human being is to be
examine.
Once a drug has proven acceptable, the trial results are combined into a large document
which includes a comprehensive description of manufacturing procedures, formulation
details, shelf life, etc. This document is submitted to the FDA for review.



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Softwares for Drug designing
1. Sanjeevini
A complete drug design software.
2. Drug-DNA Interaction Energy server
Calculates the Drug-DNA interaction energy.
3. Binding Affinity Prediction of Protein-Ligand Server(BAPPL)
Computes the binding free energy of a protein-ligand complex.
4. ParDOCK - Automated Server for Rigid Docking
Predicts the binding mode of the ligand in receptor target site.
5. Active Site Prediction

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Contd
6. Lipinski Filters
Checks whether a drug satisfies the 5 Lipinski rules.
7. Molecular Volume Calculator
Calculates the volume of a molecule
8. DNA Sequence to Structure
Generates double helical secondary structure of DNA using conformational parameters
taken from experimental fiber-diffraction studies.
9. RASPD for Preliminary Screening of Drugs
Preliminary screening of ligand molecules based on physico-chemical properties of the
ligand and the active site of the protein. This will predict binding energy of drug/target at a
preliminary stage.

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Discovery Studio

Life Science Modeling and
Simulations Tool
Summary
R&D in the pharmaceutical industry is undergoing a lot of technological changes,
and there is pressure to make the investment pay off
There is a big need to sensibly use the large amounts of chemical and biological-
related information produced in the process
Thoughtful use of new technologies, methods and softwares are becoming crucial
to the success of drug discovery
An in silico revolution is emerging in this field that can alter the conduct of early
drug development.
The use of computers and computational methods permeates all aspects of drug
discovery today and forms the core of structure-based drug design.

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References
Aurther M. Lesk. Introduction to Bioinformatics. Oxford University Press, USA (May 9, 2002) | ISBN-10:
0199251967
Madsen, Ulf Krogsgaard-Larsen, Povl Liljefors, Tommy (2002). Textbook of Drug Design and Discovery.
Third Edition, Washington, DC: Taylor & Francis.
Alexander Zien, Robert Kffner, Theo Mevissen, Ralf Zimmer and Thomas Lengauer. Identification of
Drug Target Proteins. ERCIM News No.43 - October 2000.
Lata N., Jayaram B. A binding affinity based computational pathway for active site directed lead molecule
design: Some promises and prespective. 2005, Drug Design Reviews-Online, 2(2), 145.
The Drug Development Company Database and the Alzheimer's Drug Discovery Foundation (ADDF) and
the Alzheimer Research Forum. Retrieved April 4, 2011, from http://www.alzforum.org/drg/tut/tutorial.asp/.
Supercomputing Facility for Bioinformatics & Computational Biology, IIT Delhi. Retrieved April 5, 2011,
from http://www.scfbio-iitd.res.in/
Ludwig Institute for Cancer Research (2010, February 4). New computational tool for cancer treatment.
Science Daily. Retrieved April 5, 2011, from
http://www.sciencedaily.com/releases/2010/01/100129151756.htm
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