Abstract— Increasing the odds of hit identification in screening
is of significance for drug discovery. The odds for finding a hit
are closely related either to the diversity of libraries or to the
availability of focused libraries. There are no truly diverse
libraries and it is difficult to design focused libraries without
sufficient information. Hence it is helpful to consider alternative
approaches that can enhance the odds using existing libraries.
Multiple members of a protein family have been considered
collectively in inhibitor design, on the basis of the correlation
between protein families and ligands derived from specific
compound classes. Such a correlation has been exploited in
various drug discovery studies and a general receptor-homologbased
screening scheme may be devised. The feasibility of such a
scheme in enhancing the odds of hit identification is discussed.
Index Terms—Homolgy, inhibitors, screening.
I. INTRODUCTION
High-throughput screening and virtual screening has
been extensively used in drug discovery [1, 2]. The odds
for finding a hit depends on the diversity of compound
libraries used [3]. There is no truly universal set of
representative compounds and the screening has practically
been conducted against subsets of molecules [4, 5x], which
may result in useful hits being missed. This problem was
illustrated recently by Oldenburg [3] in an example of two
highly similar compounds of the steroid family, testosterone
and estrogen, which differ only by a methyl group and a few
double bonds. If only the first is included in a library for
screening against estrogen receptor, estrogen would not be
discovered. The same is true if one starts with estrogen and
tests it against testosterone receptor.
Such a problem is likely of particular concern to
screening for agonist/activator drugs that generally require
more specific structural binding configuration than that of
antagonist/inhibitor drugs. A search of Medline shows that
less than 12% of the publications in drug screening are related
to agonist/activator drugs, which may be partly due to the
difficulty in finding an agonist or activator hit.
Agonist/activator drugs constitute an important drug class. A
search of the therapeutic target database [5] finds 44 targets of
agonist/activator drugs, many of which are important
receptors. Hence, methods for improving the odds of
screening of agonist/activator drugs as well as
antagonist/inhibitor drugs are potentially useful in new drug
discovery.
Increasing the size of libraries in a random fashion may not
always be effective or practical for solving this problem [4].
Drug-like compounds have been found sparsely distributed
through chemistry space [6]. As a result, the design of focused
libraries can be a difficult task without sufficient information.
Therefore it is helpful to consider alternative approaches that
can potentially enhance the odds of hit identification without
relying solely on the simple expansion of existing libraries.
Recent developments in exploiting the correlation between
protein families and ligands from specific compound classes
point to a receptor-homolog-based screening scheme for
improving the odds of hit identification.
II. EXPLOITATION OF THE CORRELATION BETWEEN
PROTEIN FAMILIES AND LIGANDS DERIVED FROM
SPECIFIC COMPUND CLASSES
In the search of inhibitors of specific cyclin-dependent
kinases, multiple members of the kinase family were
considered in a collective manner on the basis of their
common feature of ligand-binding mode [7]. Based on the
known binding mode of purine olomoucine at the ATPbinding
site, compounds were designed from combinatorial
libraries of 2,6,9-trisubstituted purines. Selective inhibitors for
subsets of cyclin-dependent kinases were developed from
these libraries.
The correlation between members of kinase family and
inhibitors derived from specific compound classes has been
shown and exploited in various studies [8]. For instance,
compounds based on quinazoline scaffold were found to
exhibit good structure-activity relationship against EGFR
tyrosine kinase and other related kinases (30-34). Potent ATPbinding
site inhibitors were derived from this scaffold for
EGFR tyrosine kinase (31 or 32), c-erbB2/c-erbB4/EGFR (42,
45, 62), RAF kinase (61), CSF-1R (47, 51), and VEGFR (63-
72), some of which are undergoing clinical trials. Inhibitors
were derived from the phenylamino-pyrimidine class for
PDGFR (80?), PKC-a (75) and EGFR tyrosine kinase (82). A
number of indolocarbazole derivatives were found to be
inhibitors of NGF receptor (210), protein kinase C (215), and
PDGR (213). Pyrazolo[d]pyrimidine derivatives were
designed as inhibitors of LcK (120), v-Src (220), CSF-1R
(47), and EGFR (47).
Increasing The Odds Of Hit Iidentification By
Screening Against Receptor Homologs
Yuzong Chen, Congzhong Cai , Zerong Li , Lianyi Han and Jifeng Wang
H
This correlation also appears in other protein families and
applies to agonists/activators. A few examples are serine
protease and peptide-like inhibitors (x1), nuclear hormone
receptors and steroid agonists (x2), and members of G-protein
coupled receptors and catecholamine agonists (x2).
There have been suggestions of exploiting this relationship
to various drug discovery problems [10]. It was proposed that,
by screening a common and diverse set of small molecule
inhibitors against a set of proteins from a family, specific
structure-activity relationship homology can be derived from
which potential drug discovery targets can be grouped (z1).
Based on the common molecular theme for ligands with a
certain classes of drug targets it was suggested that, in stead of
putting barriers of high-risk targets through expensive screens
of large compound collections, focused libraries of specific
compound classes should be tested first [9]. In order to direct
discovery processes to tractable chemical libraries, potential
targets can be screened from all members of a gene family that
have proven records in drug development and chemistry effort
can then be focused on the most intriguing targets (z2).
A. Screening against of receptor-homologs as a general
scheme?
It is of interest to explore the possibility of further
extending the protein-family-based approach into a more
general receptor-homolog-based scheme for drug screening.
In this scheme, as illustrated in Figure 1, screening is
conducted against a receptor and its homologs which are
defined as proteins of similar sequence in the ligand-binding
domain. These homologs likely share common structural
features at ligand-binding sites and structurally similar ligands
may exist for some of these homologs. If one or more of these
ligands is identified as a hit for the entire homolog group, the
rest may be generated by focused library design based on the
identified hits. The subsequent screening of these focused
libraries against the receptor may lead to the identification of
the specific ligands for that receptor.
In the testosterone-estrogen example, if the same set of
compounds is used for screening against estrogen receptor and
all of its sequence homologs (including testosterone receptor),
testosterone would be identified as a hit for the entire receptor
group. A focused library of steroid analogs may be
constructed based on the framework of testosterone, which
likely include estrogen. A subsequent screening of this
focused library against estrogen receptor would discover
estrogen. The odds of finding an agonist hit are thus
significantly increased.
The feasibility of this scheme depends on the existence of
structurally similar ligands, particularly agonists/activators,
for at least some of the homologs. Structurally similar agonists
are searched for two receptors, estrogen receptor and
adrenoceptor, and their sequence homologs. The identified
homologs of these receptors along with structurally similar
agonists are given in Table 1 and 2 respectively. Certain
homologs are excluded from these two groups which include
orphan receptors with unknown ligand and peptide-agonist
receptors or lipid-agonist receptors with binding sites covering
sections different from that of the respective common binding
sites defined by the estrogen receptor or adrenoceptor [11,12].
Structurally similar agonists are found for 7 out of 8 homologs
of estrogen receptor [13-15] and for 4 out of 8 homologs of
adrenoceptor [16-19] respectively. Moreover, agonist
superficially similar to the other agonists is found for a
homolog of estrogen receptor and that of adrenoceptor
respectively. Thus it appears that compounds of common
structural framework can be found for a substantial portion of
receptor homologs.
III. CONCLUSION
The correlation between protein families and ligands of
common structural theme has been exploited in various drug
discovery studies. A general receptor-homolog-based
screening scheme may be devised based on this correlation.
From the study of specific cases, it appears to be feasible to
use this scheme to enhance the odds of hit identification.
References
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Source: dspace.mit.edu

In its search for new molecules against malaria, MMV has so far supported the screening of more than 5 million compounds for their potential activity against the malaria parasite. Three partners of the MMV-supported early discovery projects have gone a step further and released the data pertaining to the active molecules into the public domain. This bold move will enable scientists the world over to access these data ensuring their antimalarial potential be used to the full.

GlaxoSmithKline (GSK), have worked to screen more than 2 million compounds from their in-house library for activity against Plasmodium falciparum – the most prevalent strain of malaria parasite in Africa. The screen resulted in the identification of more than 13,500 compounds with activity against the parasite. The largest group of compounds with a known mode of action to be identified was the kinase inhibitors, which are currently being explored in two other MMV-supported projects: GSK miniportfolio and Monash University’s Kinase Platform.

The Genomics Institute of the Novartis Research Foundation has screened more than 800,000 compounds, from a range of sources, resulting in more than 5,600 molecules active against the parasite. Four of these chemical series have been selected for further development at Novartis Institute of Tropical Diseases (NITD). One series is now undergoing candidate selection for clinical trials, while the other three are in lead optimization.

The team from St Jude Children’s Research Hospital has screened more than 300,000 unique chemical structures also against P. falciparum, yielding over 1,100 potent and selective drug discovery starting points. In collaboration with Rutgers University in New Jersey the team at St Jude is working to take these starting points to the next level.

These projects bring the total number of promising compounds to over 20,000.

Data from GSK and St Jude’s screening work has been published in Nature, while all data, including those of Novartis, can be found online through the European Bioinformatics Institute (EMBL-EBI). Sharing these data will not only expedite the discovery and development of future antimalarials it has also set a precedent for other actors in neglected disease drug discovery to follow the open access model.

source: mmv.org

The humble nematode worm could prove of inestimable in screening new compounds on account of active drugs, chic enquiry published today suggests.

Soil-homestead nematodes have a programmed avoidance response to harmful chemicals, which they detect from one end to the other nerves exposed to their conditions. Scientists led by the Wellcome Sureness Sanger Start possess genetically modified the worm C. elegans to make human proteins called receptors in these nerves: the modified worms detect and steer clear of human signalling molecules and soporific candidates.

The exciting results, reported today, 20 July 2006, in the untaken receptive-access journal BMC Biology, promise a simple assay that can be acquainted with to interview thousands of compounds for vim against human proteins – a foundation of drug event.

“The worm is a great tool to understand biology,” said Dr Michelle Teng of the Wellcome Assurance Sanger Alliance, a lead author on the discharge. “Because we understand it so well – it has a undesigning doubtlessly studied in a tizzy set-up – the role on account of each nerve has been mapped in detail. We also would rather a good truce of the signalling mechanisms in nerves that drive the responses.

“We showed that the biochemical response of the receptors emulated that seen in humans. It is very recently that, in the worm, the effects of that response are to make them toady away from the chemical stimulus. This forthright response could be adapted to to test many unknown medicine candidates.”

Medicines continually interact with receptors, which are “sensors” at the show up of cells. The crew introduced the somatostatin receptor (Sstr2) and the chemokine receptor 5 (CCR5) in the nerves that touched by to environmental cues. Somatostatin is a hormone that mediates a wide sphere of activities in humans and chemokines play an important situation in the immune system. The CCR5 receptor used is also the gateway that HIV/AIDS virus uses to set cells. Both receptors belong to a receptor family called GPCRs, which pretend to be up to 50% of current opiate targets.

The response was specific. In tests, worms responded by avoiding somatostatin or chemokine placed in their paths alone when the pertinent receptor was made in the appropriate nerves.

“We have shown that we can hijack the cellular machinery of the worm so that the man receptor proteins drive the avoidance response,” explained Dr John McCafferty, Principal Investigator at the Wellcome Trust Sanger Institute and senior author. “We chose two receptors with to a large differing functions in humans. The responses were personal to to the compounds we added and could be inhibited in the same way a effect in humans could be restrained.”

The worms could also be desensitized by pre-exposure to somatostatin or chemokine: desensitization is an important have a share of normal philanthropist response, because it ensures that our receptors can recover repayment for a fresh volley of stimulus. This is the cardinal over and over again that activation has been programmed in these nerves and the team have shown that the human receptors integrate into the worm signalling machinery.

“Systems exist already to study the response of cells in evaluation-tubes to added compounds,” continued Dr McCafferty. “However, because these are ground-dwelling worms which supply on bacteria, we could evaluate crude samples for the purpose antidepressant candidates. Together, these results make us very optimistic that these models longing be widely apt and that development of a boisterous-throughput system is practical.”

The span used a instantaneous sorting system to isolate the genetically modified worms. Although, for this swat, worm responses were scored below the microscope, automation could be integrated to achieve a higher rate of testing.

The worm model can also alleviate to define which regions of a novel compound are foremost due to the fact that its biological effect, which can be crucial in compensation producing noticeable drugs. The conspire were able to usefulness the worm assay to identify four superior building blocks within the somatostatin molecule which are known to be necessary in the interest of its make.

“These results show the power of oafish organisms such as the worm to help us not only in our understanding of biology but also in the search for untrained ways to improve healthcare,” said Professor Ronald Plasterk, Professor of Developmental Genetics at the University of Utrecht and Director of the Hubrecht Laboratory, in the Netherlands. “It is a comfortably irony of history that the worm was chosen for biomedical examination by Sydney Brenner forty years ago in Cambridge, sole a not many miles from the Sanger Institute. Then twenty years ago John Sulston started to make a gene map of the animal, and long run read its sequence as the elementary of all animal genomes.

“And infrequently a new generation of researchers again in the Cambridge area uses it to test office-seeker drugs that are immediately relevant to sympathetic vigorousness.”

Publication details
Teng MS et al. (2006) Face of mammalian GPCRs in C. elegans generates novel behavioural responses to human ligands. BMC Biology 4:22 doi:10.1186/1741-7007-4-22

The publication, which is available able of charge, according to BioMed Central’s into operation-access policy, is at: http://www.biomedcentral.com/1741-7007/4/22.

The BMC Biology website is http://www.biomedcentral.com/bmcbiol.

Participating Centres and websites

Wellcome Confidence in Sanger Institute – C. elegans lab:

http://www.sanger.ac.uk/Teams/Team37

Wellcome Trust Sanger Institute – ATLAS Engagement:

http://www.sanger.ac.uk/Teams/Team86

Erasmus Medical Concentrate – Jansen lab:

http://www2.eur.nl/fgg/ch1/cellbiology/jansen

The Wellcome Trust Sanger Initiate, which receives the majority of its funding from the Wellcome Trust, was founded in 1992 as the centre for UK sequencing efforts. The Institute is responsible suited for the completion of the sequence of approximately one-third of the human genome as equably as genomes of version organisms such as mouse and zebrafish, and more than 90 pathogen genomes. In October 2006, further funding was awarded by the Wellcome Empower to enable the Institute to build on its on cloud nine-class systematic achievements and turn to account the bounty of genome data now available to answer important questions connected with health and disease. These programmes are built surrounding a Faculty of more than 30 higher- ranking researchers. The Wellcome Sureness Sanger Association is based in Hinxton, Cambridge, UK.

http://www.sanger.ac.uk

The Wellcome Trust is the most diverse biomedical fact-finding generosity in the world, spending adjacent to £450 million every year both in the UK and internationally to support and plug delving that will improve the health of humans and animals. The Credit was established impaired the will of Sir Henry Wellcome, and is funded from a unsociable allowance, which is managed with sustained-length of time stability and crop in rail at.

Wellcome Trust Sanger Institute
Hinxton, Cambs, CB10 1SA, UK

http://www.sanger.ac.uk

source: abundanttanzania.qanka.biz

U.S. scientists say they are using a novel screening technique to identify new effects of drugs in shifting cellular energy metabolism.Researchers at Massachusetts General Hospital, which led the study, said drugs that target the way cells convert nutrients into energy could offer new approaches to treating a range of conditions, including heart attack and stroke.

The team said it identified several FDA-approved agents, including an over-the-counter anti-nausea drug, that can shift cellular energy metabolism processes in animals.

“Shifts in cells’ energy production pathways take place naturally during development and in response to demanding activities — like sprinting versus long-distance running,” said Dr. Vamsi Mootha, the lead investigator. “They are also known to be involved in several disease states. We wanted to identify compounds that can safely induce this shift … and investigate their therapeutic potential in animal models.”

The researchers said their findings, which may lead to new therapeutic strategies to treat several serious health problems, appear in the early online edition of the journal Nature Biotechnology.

source: upi.com

Drugs that target the way cells convert nutrients into energy could offer new approaches to treating a range of conditions including heart attack and stroke. Using a new way to screen for potential drugs, a team led by Massachusetts General Hospital (MGH) researchers has identified several FDA-approved agents, including an over-the-counter anti-nausea drug, that can shift cellular energy metabolism processes in animals. Their findings, being published online in Nature Biotechnology, may open the door to new therapeutic strategies for several serious health problems.

“Shifts in cells’ energy production pathways take place naturally during development and in response to demanding activities — like sprinting versus long-distance running. They are also known to be involved in several disease states,” explains Vamsi Mootha, MD, of the MGH Center for Human Genetic Research, who led the study. “We wanted to identify compounds that can safely induce this shift — those that have previously been discovered are too toxic — and investigate their therapeutic potential in animal models.”

Normally cells convert nutrients into energy by relying on two cellular processes. One involves the uptake of sugars that are broken down in the cytoplasm into a molecule called lactate via a process called glycolysis, which quickly yields a small amount of ATP, the enzyme that provides cellular energy. Alternatively, sugars and proteins can be processed in cellular structures called mitochondria to release greater amounts of ATP through a more efficient process called cellular respiration.

In cancer cells and other rapidly proliferating cells, energy is produced predominantly by glycolysis, suggesting that a shift away from that mechanism might suppress tumor growth. Previous animal studies suggested that a reduction in mitochondrial respiration could mimic a process called ischemic preconditioning, in which brief episodes of ischemia ? a reduction in blood flow ? actually protect tissue against being damaged if its blood supply is later cut off completely.

To search for compounds that shift cells from respiration to glycolysis, Mootha’s team devised a novel screening strategy. They cultured skin cells in two different nutrient environments ? glucose, which provides energy through both glycolysis and respiration, or galactose, which forces cells to rely on mitochondrial respiration alone. A drug that redirects energy metabolism from respiration to glycolysis would stop growth in the galactose-cultured cells while having little effect on cells grown in glucose. Their initial screen of almost 3,700 compounds, including nearly half of all FDA-approved drugs, identified several drugs known to inhibit cellular respiration on one end of the scale and several anti-cancer drugs that halt the growth of rapidly proliferating cells at the other, which verified the approach.

Because most agents known to mimic ischemic preconditioning in animal models are too toxic to use in human patients, the researchers were most interested in finding drugs that cause subtle metabolic shifts. The screen identified eight approved drugs that produced a less pronounced but still significant shift away from cellular respiration. One of those agents was meclizine, an over-the-counter drug used to treat nausea and vertigo ? suggesting that it passes the blood-brain barrier ? with few negative side effects.

To investigate meclizine’s potential to prevent tissue damage in heart attack or stroke, Mootha’s team collaborated with University of Rochester researchers who had developed rat models of heart attack damage and an MGH Pathology group with a mouse model of stroke damage. Blinded experiments using both animal models showed that pretreatment with meclizine dramatically reduced ischemic damage to cardiac cells in the heart attack model and to brain cells in the stroke model. They also found that meclizine’s ischemia protective effects do not appear to involve its known mechanisms.

While the study results suggest that treatment with drugs like meclizine may someday be useful for reducing the damage associated with heart attack or stroke, Mootha stresses that much additional study is needed. “Before we can think about human studies, we need to do rigorous animal testing to determine optimal, safe dosing regimens and learn more about how this drug works,” he says. He also notes that the drug-screening strategy developed by his team could help to identify previously unsuspected beneficial or detrimental effects of other approved drugs.

Mootha is an associate professor of Systems Biology at Harvard Medical School and an associate member of the Broad Institute of MIT and Harvard. Co-lead authors of the Nature Biotechnology article are Vishal Gohil, PhD, and Sunil Sheth, MD, MGH Center for Human Genetic Research (CHGR). Additional co-authors are Roland Nilsson, PhD, Fabiana Perocchi, PhD and William Chen, MGH-CHGR; Jeong Hyun Lee, PhD, and Cenk Ayata, MD, MGH Pathology; Andrew Wojtovich and Paul Brookes, PhD, University of Rochester Medical Center; and Clary Clish, PhD, Broad Institute. The study was supported by grants from the American Diabetes Association and the Smith Family Foundation.

Massachusetts General Hospital, established in 1811, is the original and largest teaching hospital of Harvard Medical School. The MGH conducts the largest hospital-based research program in the United States, with an annual research budget of more than $600 million and major research centers in AIDS, cardiovascular research, cancer, computational and integrative biology, cutaneous biology, human genetics, medical imaging, neurodegenerative disorders, regenerative medicine, systems biology, transplantation biology and photomedicine.
source: scienceblog.com

The search for new drug compounds is probably worse than looking for a needle in a haystack because scientists are limited in the size of the haystacks they can rummage through — time and money make it virtually impossible to screen or search through super-large libraries of potential compounds. This is a serious problem, because there is enormous interest in identifying synthetic molecules that bind to proteins for applications in drug discovery, biology, and proteomics, and larger libraries should mean higher odds of success.But large libraries come with large problems. Because even compounds with only modest affinity (binding to the target protein receptor with less force than those with high affinity) are usually marked as hits, researchers often end up with several hundred of them and, because of practical constraints involving time and money, no easy way to determine which might be the highest affinity or best compound to serve as a starting point to design a drug. These limitations and others have drastically blunted the use of very large libraries — monster libraries — in binding assays.

Now, in research published in the most recent issue of the journal Chemistry & Biology, Tom Kodadek, a professor at The Scripps Research Institute’s Florida campus, and his colleagues at Scripps Florida and the University of Texas Southwestern Medical Center have devised an innovative new way to solve this longstanding problem.

“Current methods severely limit the size of the libraries you can screen,” said Kodadek. “If you get 20 hits out of a 100,000 compound library, it’s feasible to re-synthesize each of those hits to test which are the most effective. But what if you want to screen 10 million compounds? It takes an impossible amount of time to re-synthesize promising compounds for further study. To find the most potent ligands, our new method stands head and shoulders over what is available to researchers today.”

Ligands are compounds that attach to proteins and alter their expression, potentially affecting a particular biomolecular activity, say, a protein pathway involved in a disease.

The new method displays millions of compounds on the surface of resin-based beads, each type of compound on a different bead. The hits are culled from the beads using a unique magnetic signature and then transferred to a microarray format — glass slides or silicon chips that can hold large numbers of compounds on their surface. The microarray format allows quantitative comparison of binding affinity that can be carried out without the need for tedious re-synthesis of many different compounds.

In the study, the team used mixed peptide/peptoid libraries — peptides make up proteins; peptoids are molecules closely related to, but more stable than peptides, making them more convenient for testing — but the method could be applied to any class of compound, according to Kodadek.

Changing the Paradigm

The Kodadek group’s method combines several different technical advances to enable this convenient and efficient screening.

These days, most active molecules are discovered through screening of two basic types. There are functional screens, in which small molecules are introduced into the wells of microtiter plates — flat plates with multiple wells that can reach as high as 9,600 — and tested individually for their ability to alter the activity of an enzyme. Alternatively, there are binding assays, an approach first developed for bead-displayed peptide libraries, where each bead displays many copies of a single molecule.

“Our new method for screening synthetic libraries and characterizing the resultant hits combines many of the features of bead library screening and microarray-based analysis in a seamless fashion,” Kodadek said. “The new technique uses several million beads, each of which displays a unique ligand — theoretically as many as 64 million compounds. The target protein has an antibody attached to it that is covered with iron oxide particles — magnetic dust. If the peptoid ligand is a legitimate ligand, and attaches to the protein, we can pull it from the mass by using a magnetized centrifuge.”

The selected compounds are then removed from the beads through a unique cleaving process and attached to glass microarray slides. These arrays are mixed with different concentrations of the target protein, allowing the affinity strength of each compound on the array to be determined quickly and efficiently.

“This technology is relevant to custom libraries that are produced on beads,” Kodadek said. “Right now, that probably constitutes five percent of screening going on. My guess, however, is that ratio will change once researchers begin to adopt this new method.”

Adoption of this new technique will take time and something of a paradigm shift, Kodadek notes. The new screening technology monitors binding of the bead-immobilized molecule to the target protein; currently, the most widely used high-throughput screens monitor function of the compound. In addition, not all laboratories currently have the equipment and expertise necessary to make microarrays of small molecules.

“I think our method can revolutionize medicinal chemistry,” said Kodadek, “but this is only the first step.”

source: sciencedaily.com