1. Does classical/receptor pharmacology still play an important
role in 21st
Century drug development?
Dimitrios Brachos
2155944
Instructor:
Dr. William Miller
Principles of Pharmacology
M.Sc. Clinical Pharmacology
Institute of Cardiovascular and Medical Sciences
University of Glasgow

2. GUID: 2155944
1
Pharmacology is one of the pivotal parts of the drug discovery and
development process. It studies the effects of drugs and how they exert their effects.
The discovery of medications can be based on substances which are found in plants,
animals, humans or that they are designed and tested in laboratories (Vallance et al,
2009).Even though pharmacology is a discipline that has a big heritage on its back,
nowadays, pharmacologists are mainly focused on molecular biology and genetics
especially on human genome (Rubin, 2007). The main outcome of this constant
pharmacological progress was that older perspectives on drug discovery and
development had become less popular and pharmacologists concentrated on what
was new and seemed promising.
The scientific field of classical/receptor pharmacology also known as forward
pharmacology is the technique which was used since the early days of drug discovery
and development of new pharmaceutical agents. This approach of drug discovery
has as a principle that we are based on the phenotypic screening of molecules of
natural or synthetic origin which could potentially serve as therapeutic agents. These
compounds/molecules are screened in cellular or animal models which have a
certain disease, aiming that the effect of the molecule will cause a desirable change
in the phenotype. During the 1980’s, the progress in genomics and molecular biology
resulted in the replacement of phenotypic screening by a technique called reverse
pharmacology. This method is a target-based screening, relying on the assumption
that a specific target is disease modifying and we screen for agents that can affect
the activity of the target (Takenaka, 2001). Then, these hits are tested in animals for
efficacy. However, some drug developers have raised serious doubts on if over-
reliance on geneticsfor target validation has contributed to reduced success in drug
discovery (Zheng et al, 2013).
Figure 1: schematic representation of classical and reverse methodology in drug
discovery and development (Hacker et al, 2009 p. 577).
Classical Pharmacology
Addition of compound
Change of biological function
Determine mechanisms
(usually in vitro methods)
Reverse pharmacology
Isolation of therapeutic agent
Identification of the compound affecting the
target
Modify drug to maximize in vivo effects
Demonstration of the desired biological
function in vivo
Note: we check the in vitro activity before the in vivo.

3. GUID: 2155944
2
The contribution of reverse pharmacology and target-based screening played
a very significant role in drug development of many drugs in the last decades and
therefore it should not be underestimated. A study using reverse pharmacology was
made in Mali in order to find a phytomedicine that can act as an anti-malarial drug
and resulted in a new standardized anti-malarian drug. They first selected a remedy
to develop, via a retrospective study focusing on the outcome of treatment. After
that, they conducted a clinical trial where they checked different dosage of the
candidate drug that presented a dose-response effect and helped that the most
adequate and safest dosage was chosen. Thirdly they compared the first line
standard treatment with the phytomedicine after a randomized control trial was
carried out. Finally, they identified active compounds which can be exploited as
biomarkers for quality control and standardization. In this way the research team
claimed that through reverse pharmacology a standardized phytomedicine can be
developed in amore timely efficient manner than common medicines (Willcox et al,
2011).Another study conducted by Aggarwal B. et al, 2011 in an effort to identify
new Anti-inflammatory compoundscombating chronic conditions. Using the reverse
pharmacology approach it was found that Ayurveda can be a source of potential
Anti-inflammatory agents. Therefore, they reviewed plants in an attempt to describe
various other plants of the same species that are currently used, their various active
components, and the pathways inhibited.
A relatively recent effort to find new genes and pathways using gene
expression profiling influenced by methotrexate and to search for genotype
association of these genes which respond to methotrexate treatment. The changes
in the gene expression profile of 11 children with juvenile idiopathic arthritis was
investigated using methotrexate, in whom response to treatment after six months
was defined. The genes exhibiting the most significant changes after treatment were
used for single nucleotide polymorphism genotyping. They then compared the
prevalence of SNPs between responders and non-responders. A Validation of the
results was available from an independent cohort. The evaluation of gene expression
in children withjuvenile idiopathic arthritisprior and post treatment and the analysis
of genetic variation in differential gene expression was conducted for the first time.
The changes in gene expression showed 1222 probe sets were differentially
expressed.From those 6 were selected due to high differential expression and were
further analyzed for genetic association in response to methotrexate. Three SNPs in
the SLC16A7 gene indicated notable association with response to methotrexate.
Finally, one SNP showed confirmed association in an independent cohort which
means that this specific gene may influence the genetic variability in methotrexate
response in juvenile idiopathic arthritis, setting as proof the statement that genes
that are expressed differentially at the level of mRNA after drug is administered may
bereliable for gene analysis (Moncrieffe et al, 2010).

4. GUID: 2155944
3
On the other hand, current drug development and discovery turnover again
to classical pharmacology approach was seriously questioning reverse pharmacology
pathway by the results from an analysis of the strategies for discovery and the
mechanism of action in molecular level for new molecules authorized by the US Food
and Drug Administration(1999 to 2008). The analysis of the data shows that from 75
which were first-in-class drugs with new molecular mechanisms, 50 (67%) were small
molecules and 25 (33%) were biologics. Furthermore, the results indicated that
phenotypic screening contribution on that small molecules which were first in class
was beyond those with target-based approach. 28 of them were found target-based
and 17 via phenotypic screening in a period when the target-based approach was
dominant in drug discovery. The inference of the previous study is that for first-in-
class drugs no cogitation of a better molecular mechanism of action may lead to
poor or unfavorable results in drug discovery and development (Swinney et al,
2011).We should also take also into account that since the 1990’s most research
projects were based on target-based drug discovery and that is why these evidences
are remarkable (Lee et al, 2013).
Other studies that aimed to solve one of the most serious diseases
concerning the central nervous system Alzheimer’s disease found to give very
disappointing or mixed results. Eli Lilly and Company tried to develop a new drug for
AD called Semagacestat, a γ-secretase inhibitor aiming to reduce levels of amyloid-β
(Aβ) because this peptide aggregates and by that it is responsible for the formation
of plaques in the brain which leads to Alzheimer’s disease. The β- and γ-
secretasescut amyloid precursor protein leading to Aβproduction. Apparently,
Semagacestat failed on in Phase III trials even though that phase II trial results
proved out that it cuts the Aβ showing inhibition of generation of 47% with dose of
100mg, 52% with 140mg, and 84% with 280mg over a 12-hour period (Bateman et al,
2009).But, unfortunately that was not the only unfavorable results in the hunt for a
new AD agent drug. A study by a company namedMyriad Genetics on another γ-
secretase inhibitor called tarenflurbilfailed in Phase III trialsbecause it did not show
any benefit on Alzheimer’s patients at that stage. At the same year, a very similar
approach on AD with Tramiprosate designed to bind in Aβ amyloid failed at late-
stage clinical trials in the United States(Opar, 2008). These disappointing results
came in addition to other previous failures and rose concerns about the approach of
research on the pathway since the reverse method used consumed a big amount of
time and expanses without any prosperous or encouraging results.
At the same time and while classical pharmacology is yet the method that led
to the discovery of most of the drugs we use until nowadays, researchers using
phenotypic drug development have achieved to exemplify many molecular details
that are involved in cell cycle, metastasis differentiation and tissue regeneration. A
study on small molecular inhibitors based on a connection of two phenotypic screens
on a posttransitional change and on visualization of chromatin and microtubules
investigated how they affect cell mitosis. The one molecule named monastrol found
to inhibit the motility of a mitotic motor protein kinasin Eg5 that is needed for

5. GUID: 2155944
4
spindle bipolarity. This finding is very important because all small molecules we
knew were that affected the mitosis were targeting tubulin and therefore monastrol
will help us understand the mitotic mechanisms in mammalian cells (Mayer et al,
1999). Another study was performed to quantitate invadopodia which is a
membrane salient in cancer cells and which contributes in the remodeling of the
cellular matrix and tissue invasion. After screening by an image-based assay 1280
pharmacological active compounds included in the Library of pharmacologically
active compounds (LOPAC) they discovered that two compounds, one of which is
called placitaxel, could contribute in the increase of both invadopodia number and
had invasive behavior. But they also found out compounds that inhibited the
formation of invadopodia without causing any toxicity and they were characterized
as cyclin-dependent kinase inhibitors and cdk5 showed to promote the formation of
invadopodia and cancer invasion through phosphorylation process and the down
regulation of caldesmon a protein which regulates actin (Quintavalle et al, 2011). A
promising approach on how small molecules could contribute on adult stem
therapies was recently published saying that the whole tissue regeneration research
has focused on transplantation therapies using embryonic pluripotent stem cells
which in many cases cause mutations, rejection due to immune responses and that
face ethical objections. Therefore, an alternative method is proposed with in vivo
modulation or an ex vivo approach of expansion of adult stem cell populations of
endogenous origin with small molecules which are drug-like. By this method it is
supported that it may lead to the discovery of new medicines for a range of human
diseases such as cardiovascular or neurodegenerative diseases (Lairson et al, 2013).
In a drug targets analysis starting at 2000 until 2012 it was found that
theactual mechanism of the 8% of those small-molecule drugs approved by FDA were
unknown (Munos, 2013). A manuscript on the mechanism of action of biguanides for
example metformin which is a first-line medicine for diabetes used over six decades
indicates that even though our knowledge is limited on the identity of molecular
drug targets, the drug can be safe and effective ( Miller et al, 2013). Researchers and
biotechnological organizations mostly use classical pharmacological approach
whereas the pharmaceutical industry prefers more certain pathways of drug
development and they use reverse pharmacology(Chasman, 2003, p. 537).
Of course both methods have advantages and disadvantages and these need
to be weighted up. In reverse pharmacology you must know the target while in
phenotypic drug discovery or development we do not need to know the target and
the fact that we do not know that can be a disadvantage. Target-based
pharmacology has also the disadvantage that the assay can be less biologically
relevant compared to in vitrophenotypic-based screening. Furthermore, reverse
pharmacology has the advantage that by knowing the mechanism of action of the
lead compound it can accelerate the preclinical drug development. On the other
hand, classical pharmacology can have multiple signaling pathways targets. A very
important disadvantage of classical pharmacology is that target identification may be
required and that can be very complicated and time consuming. In order to optimize

6. GUID: 2155944
5
structure-activity relationship in case of classical pharmacology, it may need to
develop a more targeted assay. Not to forget, in target-based or reversed
pharmacology drug target may not be relevant to the disease, as lack of human
efficacy found in late-stage clinical trials while in classical pharmacology drug target
is usually disease relevant and it may target more complex diseases. Lack of efficacy
which is found in late stage clinical trial in target-based screening is basically the
reason for phase III and submission failures as 66% from 2007 until 2010 in which
period 83 submissions for Phase III trials were made while the success of the
submission have fallen 50% (Arrowsmith, 2011).The same in a bigger extend has
happened at Phase II trials where the rates of failure are greater than any other
phase of drug development and this is again by 51% due to lack of efficacy.
(Arrowsmith, 2011).
It is obvious that the method of reverse pharmacologyto discover and
develop new drugs is a viable approach which has contributed to the scientific
research. But even though that the genomics revealed many new molecular targets,
reverse pharmacological approach did not succeed to increase the number of new
drugs discovered (Munos et al, 2009). Classical pharmacology is still serving its most
in drug discovery and it seems that scientists still rely on phenotypic drug
development because it seems more cost-effective and with higher
productivity(Reaume et al, 2011). On the other hand, over the last decades
pharmaceutical companies have chosen the pathway of drastic restriction of this
approach and the monopoly of interest was absorbed by the reverse method. The
dependence of drug discovery and development on a target-driven strategy may
increase the reduction of medicine productivity in future and therefore
pharmaceutical companies need to minimize their interventions in the scientific field
and let the scientists decide on which path the science will follow. Therefore,
classical pharmacology plays an important role on drug development on the 21st
century and can be used in future but, a way of detachment of the interests of
pharmaceutical companies from the drug discovery and developmentmust be found
to let science improve its techniques and methods and guide the research without
any ‘’extra weights’’.

7. GUID: 2155944
6
References
Aggarwal B.B, Prasad S., Reuter S., Kannappan R., Yodav V. R., Park B., Hye kim J.,
Gupta S. C., Phromnoi K., Sundaram C., Prasad S., Chaturvedi M. M., Sung B.. (2011).
Identification of Novel Anti-inflammatory Agents from Ayurvedic Medicine for
prevention of chronic diseases: ‘’Reverse Pharmacology’’ and ‘’Bedside to Bench’’
Approach. Curr Drug Targets. 2011.12(11): 1595–1653.
Arrowsmith, J. (2011). Trial watch: Phase II failures: 2008–2010. Nat Rev Drug Discov,
10(5), pp.328-329.
Arrowsmith, J. (2011). Trial watch: Phase III and submission failures: 2007–2010. Nat
Rev Drug Discov, 10(2), pp.87-87.
Bateman, R., Siemers, E., Mawuenyega, K., Wen, G., Browning, K., Sigurdson, W.,
Yarasheski, K., Friedrich, S., DeMattos, R., May, P., Paul, S. and Holtzman, D. (2009).
A γ-secretase inhibitor decreases amyloid-β production in the central nervous
system. Annals of Neurology, 66(1), pp.48-54.
Chasman, D. (2003). Protein structure. New York: Marcel Dekker.
Hacker, M., Messer, W. and Bachmann, K. (2009). Pharmacology. London: Academic.
Lairson, L., Lyssiotis, C., Zhu, S. and Schultz, P. (2013). Small Molecule–Based
Approaches to Adult Stem Cell Therapies. Annu. Rev. Pharmacol. Toxicol., 53(1),
pp.107-125.
Lee, J. and Berg, E. (2013). Neoclassic Drug Discovery: The Case for Lead Generation
Using Phenotypic and Functional Approaches. Journal of Biomolecular Screening,
18(10), pp.1143-1155.
Mayer, T. (1999). Small Molecule Inhibitor of Mitotic Spindle Bipolarity Identified in a
Phenotype-Based Screen. Science, 286(5441), pp.971-974.
Miller, R., Chu, Q., Xie, J., Foretz, M., Viollet, B. and Birnbaum, M. (2013). Biguanides
suppress hepatic glucagon signalling by decreasing production of cyclic AMP. Nature,
494(7436), pp.256-260.
Moncrieffe, H., Hinks, A., Ursu, S., Kassoumeri, L., Etheridge, A., Hubank, M., Martin,
P., Weiler, T., Glass, D., Thompson, S., Thomson, W. and Wedderburn, L. (2010).
Generation of novel pharmacogenomic candidates in response to methotrexate in
juvenile idiopathic arthritis: correlation between gene expression and
genotype. Pharmacogenetics and Genomics, 20(11), pp.665-676.
Munos, B. (2009). Lessons from 60 years of pharmaceutical innovation. Nat Rev Drug
Discov, 8(12), pp.959-968.

8. GUID: 2155944
7
Munos, B. (2013). A Forensic Analysis of Drug Targets From 2000 Through 2012. Clin
Pharmacol Ther, 94(3), pp.407-411.
Opar, A. (2008). Mixed results for disease-modification strategies for Alzheimer's
disease. Nat Rev Drug Discov, 7(9), pp.717-718.
Quintavalle, M., Elia, L., Price, J., Heynen-Genel, S. and Courtneidge, S. (2011). A Cell-
Based High-Content Screening Assay Reveals Activators and Inhibitors of Cancer Cell
Invasion. Science Signaling, 4(183), pp.ra49-ra49.
Reaume, A. (2011). Drug repurposing through nonhypothesis driven phenotypic
screening. Drug Discovery Today: Therapeutic Strategies, 8(3-4), pp.85-88.
Rubin, R. (2007). A Brief History of Great Discoveries in Pharmacology: In Celebration
of the Centennial Anniversary of the Founding of the American Society of
Pharmacology and Experimental Therapeutics. Pharmacological Reviews, 59(4),
pp.289-359.
Swinney, D. (2013). Phenotypic vs. Target-Based Drug Discovery for First-in-Class
Medicines. Clin Pharmacol Ther, 93(4), pp.299-301.
Swinney, D. and Anthony, J. (2011). How were new medicines discovered?. Nat Rev
Drug Discov, 10(7), pp.507-519.
Vallance, P. and Smart, T. (2006). The future of pharmacology. British Journal of
Pharmacology, 147(S1), pp.S304-S307.
Willcox, M., Graz, B., Falquet, J., Diakite, C., Giani, S. and Diallo, D. (2011). A “reverse
pharmacology” approach for developing an anti-malarial phytomedicine. Malar J,
10(Suppl 1), p.S8.