Archive for the ‘Compound Screening’ Category
New compounds may help develop drugs for degenerative nerve diseases
Last Updated on Wednesday, 12 May 2010 12:52 Written by Editor Wednesday, 12 May 2010 12:52
Scientists at Duke University Medical Centre have discovered certain compounds that could lead to promising new drugs for degenerative nerve diseases, such as Huntington’s disease, Alzheimer’s disease and Parkinson’s disease.
Misfolded proteins in nerve cells (neurons) are a common factor in all of these diseases.
These new compounds improve a cell’s ability to properly “fold†proteins.
It activates a master regulator to increase the supply of “protein chaperone†molecules that help fold proteins properly.
The scientists further explored one of the candidate molecules to activate the master regulator of chaperone gene expression, Heat Shock Factor 1 (HSF1), to learn whether it would work in model systems of Huntington’s disease, a devastating neurodegenerative disease of protein misfolding.
They were able to show that the molecule stimulated protein chaperones in cells and in an animal system.
The damage to early-state rat neurons was much lower in cells pre-treated with the HSF1 activator, and damage to the neurons of fruit flies that had a Huntington’s-like disorder was also greatly reduced.
The study provides a new approach to address the root cause of these diseases – protein misfolding.
“The advantage of our screen is that it identifies molecules that can elevate the levels of chaperones without inducing cellular stress and that don’t inhibit a key protein chaperone called Hsp90 that is needed for cells to function normally,†said senior author Dennis J. Thiele, Ph.D., Professor of Pharmacology and Cancer Biology.
“We found a creative way to identify new molecules that can activate the body’s natural protein folding machinery,†he added.
Lead author Daniel Neef, Ph.D., says they used genetically altered yeast to find compounds that might aid chaperone development.
The study appears online in PLoS Biology. (ANI)
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New class of brain-protecting drugs emerging
Last Updated on Wednesday, 12 May 2010 12:51 Written by Editor Wednesday, 12 May 2010 12:51
Researchers have identified a compound that mimics one of the brain’s own growth factors and can protect brain cells against damage in several animal models of neurological disease.
7,8-dihydroxyflavone is a member of the flavonoid family of chemicals, which are abundant in fruits and vegetables. The compound’s selective effects suggest that it could be the founder of a new class of brain-protecting drugs.
The results were published online this week in the Proceedings of the National Academy of Sciences.
Investigators at Emory University School of Medicine, led by Keqiang Ye, PhD, associate professor of pathology and laboratory medicine, were searching for a way to mimic a protein found in the brain called BDNF (brain-derived neurotrophic factor).
“BDNF has been studied extensively for its ability to protect neurons vulnerable to degeneration in several diseases, such as ALS, Parkinson’s and Alzheimer’s disease,” Ye says. “The trouble with BDNF is one of delivery. It’s a protein, so it can’t cross the blood-brain barrier and degrades quickly.”
Working with Ye, postdoctoral fellow Sung-Wuk Jang sifted through a library of chemicals to find those that could stimulate one of the proteins on the surfaces of neurons that BDNF binds to. They could show that 7,8-dihydroxyflavone sends survival signals to brain cells by pulling together two TrkB receiver-dish molecules, just like BDNF does.
Moreover, it is active in the brain when injected into the body cavity, meaning that it can cross the blood-brain barrier. Ye says many experimental “neuroprotectant” drugs have been unsuccessful in clinical trials for diseases such as stroke and Parkinson’s over the last decade.
“What’s different is this is a new pathway, offering us new opportunities,” he says. “This is the first molecule we’ve found that specifically triggers TrkB.”
7,8-dihydroxyflavone could partially prevent the death of neurons in experimental models of three neurological diseases:
- Seizure: Mice treated with the stimulant kainic acid
- Stroke: Loss of blood flow induced in mice by blocking a cerebral artery
- Parkinson’s disease: Mice treated with a toxin that kills the same neurons affected by Parkinson’s
To show that the effects of 7,8-dihydroxyflavone depended on TrkB, the authors used mice with a modified TrkB gene, which makes their neurons vulnerable to a chemical that is not otherwise toxic. That chemical could inhibit the effects of 7,8-dihydroxyflavone.
7,8-dihydroxyflavone is a member of a family of antioxidant compounds naturally found in foods ranging from cherries to soybeans. Tests in animals indicate that the compound has low chronic toxicity, Ye says. In clinical trials, BDNF itself can have side effects such as sensory alterations, weight loss or nausea.
“It is likely that many people take in small amounts of 7,8-dihydroxyflavone in their diets,” Ye says. “But drinking green tea or eating apples doesn’t give you enough for a sustained effect.”
In the initial screening process, several flavonoid compounds had similar properties to 7,8-dihydroxyflavone. Ye says his laboratory has already identified compounds that are several times more active. The next step is more animal studies to choose compounds likely to have the best drug profiles: stable and non-toxic.
Manuel Yepes, MD, assistant professor of neurology at Emory University School of Medicine, and his colleagues performed the stroke model experiments. Gary Miller, PhD, associate professor in Emory’s Rollins School of Public Health, and his colleagues performed the Parkinson’s-simulating toxin experiments.
Investigators from Georgia State University, UCLA, and the Centers for Disease Control and Prevention contributed to the research, which was supported by the National Institutes of Health.
Ye is an inventor of novel technology related to this research. Under Emory policies, he is eligible to receive a portion of any royalties or fees received by Emory from this technology. These relationships have been reviewed and approved by Emory University in compliance with its conflict of interest policies.
S.W. Jang, X. Liu, M. Yepes, K.R. Shepherd, G.W. Miller, Y. Liu, W.D. Wilson, G. Xiao, B. Blanchi, Y.E. Sun, and K. Ye.
A selective TrkB agonist with potent neurotrophic activities by 7,8-dihydroxyflavone. PNAS ##,## (2010)
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Open access drug discovery database launches with half a million compounds
Last Updated on Thursday, 6 May 2010 01:48 Written by Editor Thursday, 6 May 2010 01:48
ChEMBLdb, a vast online database of information on the properties and activities of drugs and drug-like small molecules and their targets, launches today with information on over half a million compounds. The data lie at the heart of translating information from the human genome into successful new drugs in the clinic.
The database is hosted by the European Molecular Biology Laboratory’s European Bioinformatics Institute (EMBL-EBI). It was transferred from biotech firm Galapagos NV in July 2008 through a £4.7 million Strategic Award from the Wellcome Trust.
ChEMBLdb is a unique resource because of its focus on drug discovery and its size: information on an additional 100 000 compounds has been added to the database for its launch, taking the number of small molecules to over 520 000, and it now contains over 2.4 million records of their effects on biological systems. The data include information about how small molecules bind to their targets, how these compounds affect cells and whole organisms, and information on the molecules’ absorption, distribution, metabolism, excretion and toxicity.
Dr John Overington, leader of the ChEMBL team at EMBL-EBI, said: “We hope ChEMBLdb will assist the translation of genomic-based insights into innovative drug therapies. We are pleased that there has already been big demand for ChEMBLdb data – not only from large pharmaceutical companies but also from academic institutions and small companies who will particularly benefit from free access to the data.”
The human genome sequence provided a molecular ‘parts list’ for a human being, comprising all the genes and proteins that are encoded by our genetic blueprint. In order to develop new medicines, it is important to catalogue how each of these ‘parts’ interacts with drugs and drug-like molecules. ChEMBLdb brings together information from the interface of the genome with chemistry into a set of ‘chemogenomic’ databases that can be used to help determine whether a particular molecule has the right properties to make an effective drug.
Professor Janet Thornton, Director of EMBL-EBI, said: “We are delighted to augment the biological data archived and served from EMBL-EBI with the ChEMBLdb resource. The database adds an important new capability to address the needs of the pharmaceutical and biotechnology industries, and provide the academic chemical biology communities with previously inaccessible data.”
Dr Alan Schafer, Director of Science Funding at the Wellcome Trust, said: “This unprecedented transfer of pharmaceutical data resources from the private sector to the public domain should have the greatest impact on researchers in academia and in small companies on limited budgets. ChEMBLdb will be a major resource of information for driving forward medicinal chemistry and drug development in the UK and internationally.”
The launch of ChEMBLdb is accompanied by the release of Kinase SARfari, an integrated resource of sequence, compound and screening data from a variety of sources for the protein kinases, a key family for drug discovery.
Image: Drug delivery capsules. Credit: Anna Tanczos, Wellcome Images
Contact
Craig Brierley
Senior Media Officer
Wellcome Trust
T +44 (0)20 7611 7329
E c.brierley@wellcome.ac.uk
Louisa Wright
Outreach Programme Project Leader
EMBL-EBI
T +44 (0)1223 494665
E louisa@ebi.ac.uk
Katrina Pavelin
Scientific Outreach Officer
EMBL-EBI
T +44 (0)1223 494452
E katrina@ebi.ac.uk
source; wellcome.ac.uk
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Product Focus: Automated Liquid Handling
Last Updated on Thursday, 6 May 2010 10:13 Written by Editor Thursday, 6 May 2010 10:13
Liquid handlers are sold in a variety of fluid-dispensing configurations, from single-channel through eight (one row of a 96-well microtiter plate), 96, and 384 channels. As the successors to manual pipettes, automated liquid handlers are the principal enablers of rapid experiments and assays conducted in tubes, vials, or microtiter plates. Liquid handlers are often just one component of systems consisting of microplate handlers, washers, readers, stackers, shakers, and incubators. Automation became necessary as assays were miniaturized from vials to tubes, and finally to microplates, and as researchers switched from radionuclide-based assays to tests that used non-radioactive detection.
Biology, medical testing, and screening of development-stage drugs are the primary markets for automated liquid handling. The energy, environmental, and heavy industries also use liquid handlers when accuracy and reproducibility, but not necessarily high throughput, are desired. “Any time you work with many samples and small quantities of fluids, automating liquid handling with a workstation will provide good return on investment,†says Scott Eaton, director of robotics marketing at Hamilton (Reno, NV).
Assessing workflow requirements is essential when selecting an automation system. Liquid transfers take time, which adds up rapidly as dispensing and other operations increase. Users who work with labile or highly toxic samples or reagents may prefer to process a smaller number of plates per run in order to move them rapidly through the protocol.
Another factor to consider, Eaton says, is the effect of physical forces on very small liquid-dispensing volumes used in higherdensity plates. “While 96-well plates remain the most common, 384- and even 1,586- well systems that employ sub-microliter volumes are gaining in popularity. At these volumes, evaporation and absorption onto the plastic plate surface become issues.â€
Automated liquid handlers have evolved from automated pipetting systems to workstations that employ liquid handling as one component, according to Nance Hall, vice president for automation and detection systems at PerkinElmer (Waltham, MA). Today’s systems perform washing, incubation, and plate manipulation in addition to dispensing. “In the past, liquid handlers performed just one function; today, they are ‘application solutions’ in which liquid handling is part of a larger picture,†Hall says.
Differentiators
Eaton believes a combination of ease-ofuse and flexibility in software is an important differentiator when selecting an automated liquid handler. “Some software is very easy to use, but it’s locked into specific applications.†The best of both worlds, he says, is a software package that presents operations graphically, provides “wizards†or templates for routine tasks, and that adapts to different assays. 
Hall suggests that potential buyers analyze their liquid-handling needs the way a cook examines a recipe. “What are the ‘ingredients’? What labware are we dispensing from and into? What do I expect from the automation component? What volumes are involved, and what sample-tip options are available?†Hall says. “Users who fail to optimize the liquid handler’s fluidics design to desired volumes will be forced to compromise either on performance or throughput.â€
Users should weigh throughput considerations when considering a liquid-handler purchase, says Jason Greene, liquid-handling product manager at BioTek (Winooski, VT). “The cutoff point for automation versus a multi-channel handheld pipette is several strips [rows or columns on a microplate] per day,†Greene says.
This seems like a small number of assays to justify the investment in automation, but as Greene notes, liquid handling is just one component of what may be a complex workflow. “Operating manually, users must work through the various reagent additions, incubations, washing, and reading steps,†he says. “Nobody likes to wash microplates. It’s pretty easy to get users to buy into the idea of automation on that function alone.†Moreover, he says, even low-throughput labs come to value the reproducibility of automated systems.
For Nadine Gassner, associate director of the Chemical Screening Center at the University of California-Santa Cruz, experience with a particular vendor is a major factor in selecting a liquid-handling system. The center, which performs highthroughput screening on natural-product and newly synthesized drug candidates, has the capability of testing hundreds of thousands of compounds in one experiment using 96- and 384-well plates.
Gassner had already been using a PerkinElmer plate reader. During the startup phase of the screening center, she visited the company and was impressed with the ability of its liquid handlers to service a variety of assays. “We were also looking for a strong industry track record and considered our experiences with PerkinElmer’s excellent service.â€
Angelo DePalma holds a Ph.D. in organic chemistry and has worked in the pharmaceutical industry. You can reach him at angelo@adepalma.com.
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C. elegans Assisted Screening of New Drugs
Last Updated on Thursday, 6 May 2010 10:10 Written by Editor Thursday, 6 May 2010 10:10
Researchers at McMaster University have developed a way to propel and direct microscopic-sized worms (C. elegans nematodes) along a narrow channel using a mild electric field. The discovery opens up significant possibilities for developing high-throughput micro-screening devices for drug discovery and other applications.
The research is described in the January 21, 2010 issue of Lab on a Chip, a leading international journal in the field of nanotechnology and bioengineering. The researchers demonstrate movement of the worms forward and in reverse inside a microchannel, guided by the direction of the electric field (electrotaxis).
“This is the first time that worms have been stimulated to move in a micro-channel device in a very precise and directed way,†said Bhagwati Gupta, assistant professor of biology. “It will allow researchers to study in real time how a proposed drug affects neurons and muscles that control motion of a live specimen.â€
“The electrotaxis of the worms has the potential to automate what is currently a slow, manual process for drug screening on worms,†said Ravi Selvaganapathy, assistant professor of mechanical engineering. “The system is fairly easy and inexpensive to scale up to conduct rapid screening of tens of thousands of chemicals in worms to identify drug candidates in a cost-effective manner. Such discovery could accelerate clinical trials in people by allowing scientists to focus only on relevant drugs and would use limited resources more efficiently.â€
C. elegans is a proven animal model for the study of human diseases because it utilizes many of the same proteins and molecules as humans. It also has a generation time of approximately only four days and a lifespan of about two to three weeks. This accelerates the understanding of the function of disease-related proteins. The use of C. elegans as a genetic model organism was first undertaken by Sydney Brenner in 1974. He was presented with the Nobel Prize in Physiology or Medicine in 2002 for his work in this area. Researchers working with C. elegans were also awarded Nobel prizes in 2006 and 2008.
Currently, researchers observe worms individually under a microscope as they move in a random manner or in a direction forced by pressure. The new development retains a worm’s natural motion and causes no harm to the worm. Researchers also found that the response of the worms was dependent on its age and neuronal development. This allows for large numbers of worms to be sorted and handled in an automated manner. This discovery allow the researchers to study how neurons respond to electricity. It can also be used to fabricate new kinds of devices to handle and manipulate large numbers of worms.
Source: biomedme.com
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Product Focus: Ultra-High-Throughput Screening
Last Updated on Wednesday, 5 May 2010 05:14 Written by Editor Wednesday, 5 May 2010 05:14
Ultra-high-throughput screening (uHTS) is an automation-based methodology for conducting hundreds of thousands of biological or chemical screening tests per day. The cutoff between high-throughput screening (HTS) and ultra-high-throughput is somewhat arbitrary. “There is no fixed boundary,†says Simon Sheard, Ph.D., business development manager at RTS Life Science (Manchester, UK), which supplies automated sample management equipment used in uHTS. The generally accepted crossover point today is 100,000 tests per day.uHTS is conducted in microtiter plates. To provide numerical perspective, 100,000 tests per day require 1,450 96-well plates (by far the most commonly used type), 261 384-well plates, or 65 1536-well plates. uHTS programs that exceed 1 million screens per day use ten times as many plates.
Equipment for conducting uHTS is indistinguishable from a standard microplate handling system, consisting of a robotic microplate handler, a liquid dispenser, and a plate reader. Additional components for washing, agitation, bar code reading and incubation are also possible.
uHTS achieves its speed through a combination of higher-density microtiter plates and multichannel (384 and higher) liquid dispensing. Equally important in achieving high throughput, however, is assay simplicity. Most ultrafast screens involve simple binding and rapid reading of results. For this reason, uHTS lends itself most readily to drug screening where, classically, tens of thousands or hundreds of thousands of wells are plated with entries from a large compound library, and the assay reagents (protein, enzyme, cell, or receptor, plus reporting reagent) remain constant in every well. Depending on the nature of the detection event, the interaction between compound and target is read as fluorescence or luminescence.
It is possible to “cheat†in HTS/uHTS by utilizing unpurified compounds, mixtures of compounds, or even multiple targets, a technique known as high-content screening because a multiple of the information normally available is collected. Wells that “light up†are examined more closely, for example by purifying mixtures or plating components individually.
The pharmaceutical connection
Parallel screening methods have been used for decades in the pharmaceutical industry. The advent of automated plate-handling and reading instrumentation, and the replacement of radiolabeling assays with luminescence- and fluorescence-based screens, created the opportunity for the several-hundredfold improvement in throughput represented by uHTS. Original equipment was expensive, but over the past decade instrumentation prices have fallen in terms of cost per assay per day, to the point where uHTS is now accessible to small drug discovery firms and academic groups. Numerous service providers also conduct uHTS services for organizations that lack this capability or whose own systems are overcommitted.
Wei Zheng, Ph.D., a group leader at the NIH Chemical Genomics Center (Rockville, Md.) learned the HTS and uHTS trades while screening drug candidates at Merck and Amgen. One of the instruments in use at the NIH Center is a plate-handling robotic system, codeveloped by Zheng at Merck, that processes hundreds of thousands of wells per day and has 1,536-well capability. “It runs between half a million and a million screens per day, depending on the assay,†Zheng told Lab Manager Magazine. The system uses plate readers from PerkinElmer and GE, and core robotics from Kalypsys Systems.
Zheng’s group uses 1,536-well plates almost exclusively, as do most pharmaceutical labs. “Miniaturization saves time and enables higher throughput at reduced cost,†he notes. However, minuscule assay volumes sometimes create difficulties for cell-based assays. “It’s often difficult to deliver the number of cells you need for an assay at such low volumes. In these circumstances the screens cannot be run at 1,536-well density.â€
Recently, researchers from the Chemical Genomics Center, in collaboration with scientists at Trinity College (Dublin, Ireland) reported on a screen of 17,143 FDA-approved and experimental drugs. The biological target in this case was a panel of human liver enzymes that metabolize drugs, and hence are critical to a medicine’s effectiveness.
uHTS received a bad reputation around the beginning of the decade, based on a perceived low success rate in identifying new drugs. The fault, says Zheng, was not with uHTS methods but with the drug companies’ choice of screening targets.
Simon Sheard agrees. “We hear comments about the failure of the ‘law of big numbers’ regularly. That’s a generalization, and the approach of cranking the handle faster has not completely fallen out of use. Nevertheless, what we have seen during the last few years is a shift away from uHTS to automated screening of smaller compound sets through assays that provide more information per well, or higher-quality data.â€
HTS and uHTS systems don’t differ much in terms of instrumentation. What changes is the trend towards modularity. “Both systems employ a collection of instruments linked by software and robotics,†Sheard observes. As assay strategies become more sophisticated and screens more numerous, the number of components increases. uHTS is greatly facilitated, for example, by dedicated compound management systems that store compounds directly in readyto- test plates. At some point, Sheard notes, “It may not be sensible to have a single robot feeding plates to numerous instruments.†And all this added functionality necessitates software products that tie everything together seamlessly.
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Pomegranate ‘can combat MRSA and other superbugs’
Last Updated on Wednesday, 5 May 2010 01:27 Written by Editor Wednesday, 5 May 2010 01:27
In a series of tests conducted over three years, academics found that mixing the fruit’s rind with two other natural products – metal salts and vitamin C – greatly enhanced its infection-fighting properties. The discovery could pave the way for a lotion to be developed for patients or perhaps, in time, a new antibiotic.
Declan Naughton, professor of biomolecular sciences at Kingston, described the breakthrough as “significant”.
Naughton said scientists were searching for a way to create new antibiotics because of the rise in infections resistant to drugs on the market. One way to go about it was to screen natural products, he said.
“A great deal of medicines come from plants, but the normal approach taken by the pharmaceutical industry is to try to find one particular active molecule,” he said. “After a considerable number of screening experiments, we found that combining three ingredients – pomegranate rind, vitamin C and a metal salt – gave a much more potent effect: killing off or inhibiting drug-resistant microbes from growing.
“It was the mix that fantastically increased the activity – there was synergy, where the combined effects were much greater than those exhibited by individual components. It shows nature still has a few tricks up its sleeve.”
The tests were conducted using microbes taken from hospital patients. Scientists found that pomegranate rind mixed with metal salts were most effective against MRSA, while adding vitamin C helped tackle other common hospital infections.
Naughton said the idea of using foodstuffs was unusual but meant that the body should be more able to cope with its application. “Patients are less likely to experience any major side-effects,” he added.
It is not the first time pomegranate has been shown to have medical benefits. The fruit has already been hailed a super-food with claims that its juice can help protect against a range of ailments, from heart disease to male impotence.
Other scientists welcomed the findings but pointed out that they were limited to tests in the laboratory – and had yet to be developed for use on people.
Anthony Coates, professor of medical microbiology at St George’s in London, said: “What is the significance of all of this? Well, there is no doubt that these natural products like pomegranate are of interest. This observation – the fact it has acted against MRSA and other drug-resistant infections – is potentially significant. But we need to remember it is early research, of an observational nature, in vitro.” Coates said much more work needed to be done to answer questions such as which component was the most active and to look at toxicity when applying the treatment to humans.
However, he pointed to other studies that had also highlighted the benefits of the fruit. One trial on 60 patients found that it had an anti-dental plaque effect, for instance.
Any discovery that was a potential step towards a new antibiotic was a positive thing, he added.
“The need for new antibiotics is acute,” said Coates. “To put it in context, about 20 new classes of antibiotics were marketed between 1940 and 1962 yet only three have been marketed since. In all classes, resistance has arisen. Most antibiotics come from nature, so it is very valid to look at natural sources.”
■The leaves and bark of the willow tree have been talked about for centuries as a remedy for some illnesses. This is because they contain salicylic acid – the precursor to aspirin. The Royal Society published findings about the medicinal properties of the natural ingredient in the 18th century.
â– Taxol (paclitaxel) is a chemotherapy drug that has been in use for many years. It is extracted from yew bark and needles. The white powder is turned into a clear, colourless liquid and administered intravenously. It is used in the treatment of ovarian, testicular and lung cancer among others.
■The same scientists who discovered that pomegranate rind could counter MRSA also discovered that white tea could help prevent cancer and heart disease. It comes from the same plant as other teas, but the leaves are picked and harvested before fully open – when the buds are covered by fine, white hair.
â– Rhubarb root has naturally occurring anthraquinones (organic compounds) which have a laxative effect.
â– Animals can also provide medicines. For example, a series of antibiotic peptides were extracted from the skin of the African clawed frog.
Source: guardian.co.uk
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Screening for Antiviral Activities of Isolated Compounds from Essential Oils
Last Updated on Wednesday, 5 May 2010 12:40 Written by Editor Wednesday, 5 May 2010 12:40
Akram Astani1,2, Jürgen Reichling3 and Paul Schnitzler1 1Department of Infectious Diseases, Virology, University of Heidelberg, Heidelberg, Germany, 2Yazd Shahid Sadoghi University of Medical Science, Safaieh, Yazd, Iran and 3Department of Biology, Institute of Pharmacy and Molecular Biotechnology, University of Heidelberg, Germany
Essential oil of star anise as well as phenylpropanoids and sesquiterpenes, e.g. trans-anethole, eugenol, β-eudesmol, farnesol, β-caryophyllene and β-caryophyllene oxide, which are present in many essential oils, were examined for their antiviral activity against herpes simplex virus type 1 (HSV-1) in vitro. Antiviral activity was analyzed by plaque reduction assays and mode of antiviral action was determined by addition of the drugs to uninfected cells, to the virus prior to infection or to herpesvirus-infected cells. Star anise oil reduced viral infectivity by >99%, phenylpropanoids inhibited HSV infectivity by about 60–80% and sesquiterpenes suppressed herpes virus infection by 40–98%. Both, star anise essential oil and all isolated compounds exhibited anti-HSV-1 activity by direct inactivation of free virus particles in viral suspension assays. All tested drugs interacted in a dose-dependent manner with herpesvirus particles, thereby inactivating viral infectivity. Star anise oil, rich in trans-anethole, revealed a high selectivity index of 160 against HSV, whereas among the isolated compounds only β-caryophyllene displayed a high selectivity index of 140. The presence of β-caryophyllene in many essential oils might contribute strongly to their antiviral ability. These results indicate that phenylpropanoids and sesquiterpenes present in essential oils contribute to their antiviral activity against HSV.
Keywords: antiviral activity – herpes simplex virus – mode of action – phenylpropanoid – selectivity index – sesquiterpenes – star anise essential oil
For reprints and all correspondence: Paul Schnitzler, Department of Infectious Diseases, Virology, University of Heidelberg, Im Neuenheimer Feld 324, 69120 Heidelberg, Germany. Tel: +49-6221-56 50 16; Fax: +49-6221-56 50 03; E-mail: paul_schnitzler@med.uni-heidelberg.deReceived June 22, 2009; accepted October 15, 2009
Source: oxfordjournals.org
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James Thomson’s Cellular Dynamics Launches iCell Cardiomyocytes Commercially for Drug Candidate Toxicity Screening
Last Updated on Wednesday, 5 May 2010 11:52 Written by Editor Wednesday, 5 May 2010 11:52
The University of Wisconsin’s James Thomson, whose vision was behind the founding of Cellular Dynamics, has from the initial creation of Induced Pluripotent Stem Cells felt that their use in the testing of new drugs would mark their greatest contribution, at least initially.Thursday the company announced the commercial launch of iCell Cardiomyocytes for use in testing of new drug candidates by the pharmaceutical industry. These human heart cells are designed to aid drug discovery and improve the predictability of drug compound efficacy and toxicity screens, weeding out ineffective and potentially toxic compounds early in the pharmaceutical pipeline process before significant time and resources have been invested.
iCell Cardiomyocytes are the first product developed by anyone from iPS cells and were discovered by CDI senior research fellow Junying Yu, Ph.D., then a postdoctoral research associate in the University of Wisconsin-Madison laboratory of James Thomson. Yu’s discovery followed a similar and almost simultaneous discovery by Shinya Yamanaka at Kyoto University.
Derived from induced pluripotent stem (iPS) cells, iCell Cardiomyocytes spontaneously beat in vitro and exhibit the electrophysiological and biochemical properties of normal human heart cells. Thus, both logically and according to Cellular Dynamics, iCell Cardiomyocytes provide significant advances over non-human cell models, which may exhibit a different response than human tissue. The same advantage is suggested to hold over tumor-derived cell models, which are genetically different than normal cells; and cadaveric cells, which exhibit batch-to-batch variability, de-differentiate under in vitro conditions, and exhibit non-cardiomyocyte behavior.
iCell Cardiomyocytes are produced in-house by Cellular Dynamics from a master cell bank of iPS cells expanded from a single clonal population reprogrammed from fully mature human cells using Dr. Thomson’s patented technology. Cellular Dynamics has reportedly developed a proprietary process to industrialize iCell Cardiomyocytes production so that the cardiomyocytes are manufactured at the high quantity, quality and purity required by pharmaceutical companies. The company has successfully engaged in pre-launch validation testing with several pharmaceutical customers.
James Thomson, chief scientific officer, had the following to say about the iCell launch: “Rapid application of stem cell technology has been a goal both of my laboratory at the University of Wisconsin and Cellular Dynamics. Utilizing human iPS cells for new drug toxicity testing should improve the drug discovery process in a timeframe that has an effect on human healthcare now, not 10 years from now. Ultimately applications of stem cell technology in drug discovery will provide great utility and enable movement toward a long-term goal of cellular-based therapeutics and personalized medicine.”
Adapted from the Cellular Dynamics announcement.
Source: stemcelldigest.net
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Hi-tech microscopes make androgen therapy ‘personal’
Last Updated on Thursday, 28 January 2010 06:03 Written by Editor Thursday, 28 January 2010 06:03
HOUSTON — (December 9, 2009) — On rare occasions, an infant is born with outward appearance of a female but the XY chromosomes of a male. If the child has a normal Y chromosome — the chromosome responsible for testicular development — the condition is known as androgen insensitivity syndrome.
Experts estimate such births occur in about one in 20,000 infants. Other children are born with a partial form of the condition that can affect their genitalia and/or fertility, but how many is not known.
The cause is a wide range of androgen receptor (AR) mutations that fail to perceive the presence of the male hormones testosterone and dihydrotestosterone to differing degrees. How to overcome the problem remained a mystery until Baylor College of Medicine and Michael E. DeBakey Veterans Affairs Medical Center experts used a high throughput, automated microscopy technique called high content analysis to solve the puzzle. A report of their findings appears in the current issue of PLoS One, an open access journal.
Reverse effect of mutation
They not only identified the functional abnormality of the AR, but also used high content analysis to “personalize” a treatment that reverses the effects of that mutation.
“With this microscopy technique, we have been able to quantify how the receptor moves and functions inside cells taken from children with normal receptors and in those with the mutation,” said Dr. Marco Marcelli, professor of medicine-endocrinology at BCM and a physician at the Michael E. DeBakey Veterans Affairs Medical Center. He and Dr. Michael Mancini, associate professor of molecular and cellular biology at BCM, and director of its Integrated Microscopy Core, are senior authors of the report.
Androgen insensitivity syndrome
They used banked cells taken from patients – both those with the mutation and those without – to study the action of the receptor in cell cultures grown in the laboratory. Dr. Michael J. McPhaul, a collaborator on the study and a professor of internal medicine—endocrinology at The University of Texas Southwestern School of Medicine Dallas, provided the samples from patients with androgen insensitivity syndrome.
“We did this on a cell-by-cell basis, using high content analysis,” said Mancini. “It is a proof-of-principle study carried out as though we had a patient and a library of hormones. We tried to find the perfect hormone for the mutation through high-speed collection of dozens of measurements from thousands of cells.”
“In two of the three specimens we tested, we found we were able to reverse the activity of the mutated receptor to almost normal,” said Marcelli.
In one patient, large doses of the male hormone dihydrotestosterone were sufficient. In another, they used a synthetic androgen that also activated the receptor.
These approaches overcame the central problem – the mutation changes the shape of the receptor and prevents it from maintaining normal contact with the hormone. It is as though a key is bent and can no longer turn the tumblers in a lock. In these cases, the hormone is designed to go into a pocket created by the receptor. When the pocket is changed by the mutation, the hormone is unable to establish good contact.
“Large amounts of testosterone may create more stable contact,” said Marcelli. “The synthetic androgen may have a conformation that establishes better contact.”
Superandrogen
In the future, scientists may be able to screen large banks of such compounds to find a “superandrogen” that may be even more efficient.
“We might be able to use this technique to create a personalized medicine test,” said Mancini.
Similar techniques might be used to screen drugs for treatment of different cancers, particularly those in which the androgen receptor is responsible for cancer progression. This study proves that the concept is valid providing quantitative information collected quickly on numerous measurements normally requiring separate biochemical tests and huge numbers of cells.
Marcelli said they have yet to use this kind of technique in patients, and such studies will require careful preparation, and go through a variety of approvals before it can be used in clinics. He also said it would be used only in individuals with the partial form of the syndrome. Finding well-matched hormones to defective androgen receptors through screening of thousands of compounds from available libraries could be one of the future developments of this technique.
The paper’s first author, Dr. Adam T. Szafran, an M.D./Ph.D. student who worked in Mancini’s laboratory, championed these studies, said Mancini. Szafran is now finishing the clinical part of his studies at BCM.
Others who took part in this study include Drs. Sean Hartig and Ivan P. Uray, Maria Szwarc, Jennifer Bell, Huiying Sun, Yuqing Shen and Sanjay N. Mediwala, all of BCM. Sun, Shen and Mediwala are also of the MEDVAMC.
Funding for this work came from the National Institute of Diabetes and Digestive and Kidney Diseases, the John S. Dunn Foundation and the Veterans Administration.
Source: bcm.edu
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Alzheimer’s Research Provides Potential Treatment for UTI
Last Updated on Monday, 11 January 2010 06:05 Written by Editor Monday, 11 January 2010 06:05
One element links the disparate areas of research: amyloids, which are fibrous, sticky protein aggregates. Some infectious bacteria use amyloids to attach to host cells and to build biofilms, which are bacterial communities bound together in a film that helps resist antibiotics and immune attacks.
Amyloids also form in the nervous system in Alzheimer’s disease, Parkinson’s disease and many other neurodegenerative disorders.
To probe amyloids’ contributions to neurodegenerative diseases, scientists altered potential UTI-fighting compounds originally selected for their ability to block bacteria’s ability to make amyloids and form biofilms. But when they brought the compounds back to UTI research after the neurology studies, they found the changes had also unexpectedly made them more effective UTI treatments.
“Thanks to this research, we have evidence for the first time that we may be able to use a single compound to impair both the bacteria’s ability to start infections and their ability to defend themselves in biofilms,” said senior author Scott J. Hultgren, Ph.D., the Helen L. Stoever Professor of Molecular Microbiology at Washington University.
The findings were reported online in Nature Chemical Biology.
The National Institutes of Health has estimated that over 80 percent of microbial infections are caused by bacteria growing in a biofilm, according to Hultgren. Scientists in Hultgren’s laboratory have worked for decades to understand the links between biofilms and UTIs.
“UTIs occur mainly in women and cause around $1.6 billion in medical expenses every year in the United States,” said co-lead author Jerome S. Pinkner, laboratory manager for Hultgren.
“We think it’s likely that women who are troubled by recurrent bouts of UTIs are actually being plagued by a single persistent infection that hides in biofilms to elude treatment,” Pinkner added.
Co-lead author Matthew R. Chapman, Ph.D., now associate professor of molecular, cellular and developmental biology at the University of Michigan, was a postdoctoral fellow in Hultgren’s lab in 2002 when he discovered that the same bacterium that causes most UTIs, Escherichia coli, deliberately makes amyloids. The amyloids go into fibers known as curli that are extruded by the bacteria to strengthen the structures of biofilms.
To treat UTIs, Hultgren’s lab has been working with Fredrik Almqvist, Ph.D., a chemist at the University of Umea in Sweden, to develop compounds that block bacteria’s ability to make curli, disrupting their ability to make biofilms and leaving them more vulnerable to antibiotics or immune system attacks.
Almqvist recently suggested altering a group of the most promising curli-blockers to see if they could also block the processes that form amyloids in Alzheimer’s disease.
The alterations worked: In laboratory tests, the new compounds prevented the protein fragment known as amyloid beta from aggregating into amyloid plaques like those found in the brain in Alzheimer’s disease.
When scientists took the new compounds back to a mouse model of UTIs, though, they received a surprise. The altered compounds were better at reducing the virulence of infections, inhibiting not only curli formation but also the formation of a second type of bacterial fibers, the pili.
“Pili aren’t made of amyloids, but they are essential to both biofilms and to the bacteria’s ability to initiate an infection,” Hultgren said.
Hultgren and colleagues are already developing even more potent infection and amyloid fighters, screening a library of thousands of chemicals similar to the most promising compounds from the study.
Chapman cautions that it’s too early to tell which, if any, of the compounds will be helpful in treating neurodegenerative diseases.
“Much neurodegenerative drug development has focused on ways to break up amyloids or prevent them from forming, but because amyloids may also be an important part of normal cellular physiology, we need to identify molecules that will target only the toxic amyloid state,” he said.
Source: farsnews.com
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iThemba Pharmaceuticals (Pty) Ltd and Pyxis Discovery B.V. Announce Collaborative Agreement to Offer
Last Updated on Monday, 11 January 2010 06:01 Written by Editor Monday, 11 January 2010 06:01
iThemba Pharmaceuticals’ service division and Pyxis Discovery announced today that they have signed a collaborative agreement to jointly market both companies’ services.
Pyxis’s world class computational chemistry and lead discovery expertise will be coupled with iThemba’s service division to provide medicinal and synthetic chemistry support to projects identified through Pyxis’ international client network. iThemba Pharmaceuticals and Pyxis Discovery also announced today that they are entering into a co-marketing agreement to offer virtual libraries which will be exclusively synthesized for clients. Pyxis’s smart approach of designing and selecting compounds facilitates a rapid and efficient lead discovery and library design process and this coupled to iThemba’s expertise in synthetic and medicinal chemistry will provide our customers with a unique service offering opportunity.
“The intellectual and technological support from Pyxis will enhance both of our service offerings,” said Chris Edlin, CSO of iThemba. “Our customers will gain the advantage of our coupled expertise in design, synthesis and medicinal chemistry prowess.”
“Combining the outstanding medicinal chemistry expertise of iThemba with our design approach helps us to provide our clients with a more complete set of services, resulting in swiftly progressing lead discovery and optimization projects.” said Ron van der Valk, Managing Director of Pyxis Discovery. “In addition to this, we hope that our collaboration with iThemba will support their ambition to bring affordable medicines to the less fortunate people in this world.”
About iThemba Pharmaceutical (Pty) Ltd. (http://www.ithembapharma.com)
iThemba Pharmaceuticals (Pty) Ltd., based in Modderfontein, Gauteng, South Africa is founded to discover and develop new and affordable medicines for the diseases of poverty in Africa. The company is funded by the Biotechnology Regional Innovation Centers, LIFElab and BioPAD of the Department of Science and Technology, Government of South Africa. Utilizing leading edge proprietary technology and its expertise in synthetic organic chemistry, iThemba Pharmaceuticals will become the premier research focal point in Africa for infectious diseases including HIV, tuberculosis, malaria and their associated co-infections. The company will create shareholder value by coupling the company’s own drug discovery efforts with collaborative research initiatives and cash-generating contracts to reduce the risks and costs of developing medicines for neglected diseases and low profit-margin markets.
About Pyxis Discovery B.V. (http://www.pyxis-discovery.com)
The ambition of Pyxis Discovery is to be the preferred chemistry service provider for companies that are active in small molecule drug discovery. Pyxis Discovery’s Smart approach of designing and selecting compound libraries facilitates a rapid and efficient lead discovery process, yielding lead compounds with excellent pharmacological profiles. Pyxis Discovery uses proprietary software algorithms for compound design and selection and a Global Supplier Database of nearly all commercially available screening compounds to provide its clients with screening libraries that are tailored to their specific needs. Furthermore, Pyxis Discovery offers high quality compound libraries off the shelf. Pyxis Discovery is headquartered in the Netherlands and has a worldwide presence with also an office in Boston, Massachusetts and representation in Japan.
Source: Pyxis Discovery B.V.
Source: melodika.net
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Drug giant will allow screening of its library of compounds to seek potential treatments for neglected diseases
Last Updated on Monday, 11 January 2010 05:48 Written by Editor Monday, 11 January 2010 05:48
The Drugs for Neglected Diseases initiative (DNDi) has announced an agreement with drug giant Pfizer that will allow it access to the Pfizer library of novel chemical entities, in order to screen it for compounds that could be developed into new treatments for three of the most neglected infectious diseases of poverty: human African trypanosomiasis (HAT), visceral leishmaniasis (VL) and Chagas disease.
Pfizer vice president Dr Manos Perros said, “We are expanding our commitment to the fight against tropical diseases by joining forces with DNDi by sharing our collection of chemical compounds and the knowledge and expertise associated with these chemical entitiesâ€. His colleague Dr Sam Azoulay said, “We are confident that the significant resources and expertise that public-private partnerships such as this one bring together, will accelerate and significantly increase the chances of success in the search for effective new drugs against serious infections that disproportionately affect the poorâ€.
Under the agreement, scientists in institutes affiliated with DNDi will test at least 150,000 compounds in the Pfizer library against Trypanosoma brucei, Leishmania donovani and Trypanosoma cruzi, the kinetoplastid parasites that cause HAT, VL and Chagas disease, respectively. The researchers will seek compounds that show initial activity against the parasites, and thus might form the basis for novel drug discovery programmes to treat the diseases. The screening will be undertaken at the Eskitis Institute for Cell and Molecular Therapies, Griffith University in Brisbane, Australia (for HAT) and the Institut Pasteur Korea (VL and Chagas disease).
“This agreement provides us access to a compound library of novel chemical entities that has never been explored for kinetoplastid diseases. This marks an important step towards DNDi’s objective of building a robust portfolio and to feed the research and development pipeline with new promising compounds,†said Dr Shing Chang, R&D director at DNDi. In July this year, DNDi announced a similar agreement with Merck – see press release
.
Within the same week as the finalising of its agreement with Pfizer, DNDi also announced it is to receive $15 million of Gates Foundation funding over the next five years, which it will use for the development of fexinidazole, currently the only new drug candidate in clinical development for sleeping sickness – see press release. Further information about DNDi is available here.
Source: tropika.net
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University of Minnesota invention will help speed development of drug treatments for heart failure
Last Updated on Monday, 11 January 2010 05:33 Written by Editor Monday, 11 January 2010 05:33
Contacts: Patty Mattern, University News Service, (612) 624-2801, mattern@umn.edu
John Merritt, Office of the Vice President for Research, (612) 624-2609
Stacie Byars, Celladon, (206) 660-2588
MINNEAPOLIS / ST. PAUL (11/23/2009) —Research conducted by University of Minnesota scientists, in collaboration with Celladon Corporation, has led to the invention of technology to more rapidly identify compounds for the treatment of heart failure.
Chronic heart failure is an increasingly important health problem. It is the leading medical cause of hospitalization and is expected to result in an estimated direct and indirect cost to the health care system of $37.2 billion in 2009 alone. About 5.7 million people in the United States have heart failure, and it contributes to or causes some 290,000 deaths annually. However, developing new treatments is an extremely costly and time-consuming process, taking nearly a decade to gain regulatory approval and requiring hundreds of millions of dollars.
The technology, developed by the universitys David Thomas and Razvan Cornea and Celladon Corporations Krisztina Zsebo, allows for increased screening efficiency of compounds capable of disrupting the interactions of proteins implicated in the development of heart failure. Fluorescence resonance energy transfer (FRET) is used to measure disruption of the calcium regulatory system, which has long been implicated in cardiovascular disease. This will provide key information on a particular drugs likelihood of success early in the screening process, since compounds that decrease FRET are good candidates for further development.
“Dr. Cornea and I, along with our students, have worked for more than a decade developing methods for preparing membranes from purified components, and using FRET to detect changes in protein interactions,” Thomas said. “Scientists from Celladon saw the potential for drug discovery, and this resulted in a breakthrough that has added an exciting new dimension to our research program.”
The high-throughput assay, developed by the university team, is based on a reconstituted membrane system composed of purified lipid and protein components. This technique is especially important because the interactions of integral membrane proteins are more complex than soluble proteins, making it very difficult to produce a synthetic system that recapitulates the cellular interactions in a large-scale and reproducible manner.
Celladon, based in La Jolla, Calif., has acquired an exclusive license for the technology from the University of Minnesota for the development of molecular therapies for cardiovascular diseases. Celladon also provided funding for the research that allowed Thomas to further refine the assay.
“This technology is very important to the efficient selection and advancement of compounds with the potential to increase cardiac contractility and potentially accelerates product opportunities that will ultimately benefit patients and development partners alike,” said Krisztina M. Zsebo, Ph.D., president and chief executive officer of Celladon Corporation. “Celladon’s investigation and development of first-in-class CDN small molecules as intravenous and oral drugs for the treatment of acute and chronic heart failure sets us apart in the cardiovascular field and presents multiple partnering opportunities.”
Source: umn.edu
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Developments in Focused Kinase Libraries
Last Updated on Monday, 11 January 2010 03:14 Written by Editor Monday, 11 January 2010 03:14
Chemical libraries have long been a mainstay in the search for new pharmaceutical compounds, and they have been created using many different paradigms. Vast diverse collections of unique compounds have been screened at high throughput to find appropriate effects on target proteins. Such large libraries continue to be used for drug discovery, but screening smaller, more focused libraries, can provide more efficient solutions with better overall hit rates.
Ten years ago, BioFocus®, a Galapagos company, launched a nonexclusive compound library specifically designed to target serine-threonine and tyrosine kinases. This library, SoftFocus® Kinase library one (SFK01), was designed to mimic the binding of ATP to the catalytic (hinge) region and had a rather simplistic design based around an aminopyrimidine core (Figure 1).
Targeting Kinases
| Protein kinases are enzymes that phosphorylate substrate proteins at specified residues such as serine, threonine, and tyrosine. The phosphorylation of the substrate protein initiates a cascade, that in turn, modulates the transcription of a gene or set of genes. Kinases play pivotal roles in modulating diverse cellular activities, including growth, differentiation, metabolism, adhesion, motility, and death, and have been implicated as important mediators of certain forms of cancer.
Kinases, therefore, represent key druggable target proteins. The initial realization that most kinases possess highly conserved catalytic domains initially made kinase targets ideally suited for compound screening via focused chemical libraries in which the library compounds were specifically created to bind to the catalytic (hinge) regions. Despite the slightly higher costs generally associated with the design and synthesis of such focused libraries compared to large and diverse compound collections, true savings can be gained as a result of shortened project cycles coupled with the reduced costs of screening, storage, and quality control of a smaller screening library. The increasing wealth of structural data available along with a number of new techniques such as in silico design has enabled the continual development of kinase-focused collections, providing increasingly more sophisticated chemical structures. One of the key benefits of this is that the current range of SoftFocus kinase libraries has been designed to target additional binding modes to those involving the hinge region; most notably the DFG-out binding mode and the novel binding mode first observed in the kinase PIM-1. |
In Silico Design
Current in silico design processes enable automated docking and scoring of scaffold ideas into a variety of known x-ray structures that have been selected, not only for broad coverage of the kinome, but also different conformational states of individual kinase enzymes. The design premise is that, if the core of the molecule or scaffold contains the key recognition groups for binding into one of the known conformational states, it has the potential to target any kinase.
However, as different side chains or monomers are added, the potential to gain specificity for one target over another becomes reality. The various docking methods used in the design of the BioFocus libraries include hinge binding, the DFG-out model, and novel binding modes.
Hinge Binding
| Hinge-binding library designs are validated by docking a minimally substituted scaffold into various different high-resolution kinase x-ray structures. These structures have been selected from across the phylogenetic tree to ensure broad coverage of tyrosine and serine/threonine kinases.
The BioFocus Kinase Toolkitâ„¢ provides a two-dimensional map (2D Roadmap) of the key ligand-binding features within a customized ATP-site model, allowing predictions of affinity, selectivity, and likely off-target issues based on the compositions of the individual sub-sites. This knowledge can then be used to select the appropriate side chains or monomers with which to decorate the scaffolds. |
DFG-Out Model
An approach to the design of kinase libraries with higher selectivity potential is to target the DFG-out allosteric pocket adjacent to the ATP site. BioFocus has developed a generalized binding model of the DFG-out pocket that enables the targeting of a range of inactive kinase conformations.
Binding Modes
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Focusing on Success
By constantly refining the compounds using in silico methods, including those described above and with the development of innovative in silico models and applications such as Cresset BioMolecular’s molecular Field technology, the latest advances in kinase research can be incorporated into novel libraries. Consequently, this may also provide a strong intellectual property position to those who screen them. Indeed, based on the screening of SoftFocus kinase libraries alone, over 70 known patents have been applied for or granted.
A recent example from Galapagos highlights this. Following a screen of some 16,000 BioFocus focused kinase compounds, three hit series were identified that showed structure-activity relationships against a novel rheumatoid arthritis target. Two of these compound series were progressed to the hit-to-lead phase and subsequently one series was optimized, and a compound is currently undergoing clinical trials. The project time from screening to preclinical candidate nomination was three years.
There is no doubt that the drug discovery process should be shortened whenever possible. It is essential to improve the lead discovery process. Focused compound libraries such as the SoftFocus collection have the inherent capabilities to provide a robust hit discovery process, with good hit-to-lead conversion rates and shorter development times.
Furthermore, such libraries can maximize the benefit of new techniques such as in silico modeling, enabling them to be consistently developed over time. BioFocus has also maintained flexibility in its library-generation processes using these techniques, which has enabled the development of novel libraries with proven results.
Source: genengnews.com
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DG-AMMOS: A New tool to generate 3D conformation of small molecules using Distance Geometry and Automated Molecular Mechanics Optimization for in silico Screening.
Last Updated on Monday, 11 January 2010 12:27 Written by Editor Monday, 11 January 2010 12:27
Discovery of new bioactive molecules that could enter drug discovery programs or that could serve as chemical probes is a very complex and costly endeavor. Structure-based and ligand-based in silico screening approaches are nowadays extensively used to complement experimental screening approaches in order to increase the effectiveness of the process and facilitating the screening of thousands or millions of small molecules against a biomolecular target.
Both in silico screening methods require as input a suitable chemical compound collection and most often the 3D structure of the small molecules has to be generated since compounds are usually delivered in 1D SMILES, CANSMILES or in 2D SDF formats.
Results: Here, we describe the new open source program DG-AMMOS which allows the generation of the 3D conformation of small molecules using Distance Geometry and their optimization via Automated Molecular Mechanics Optimization. The program is validated on the Astex dataset, the ChemBridge Diversity database and on a number of small molecules with known crystal structures extracted from the Cambridge Structural Database.
A comparison with the free program Balloon and the well-known commercial program Omega generating the 3D of small molecules is carried out. The results show that the new free program DG-AMMOS is a very efficient 3D structure generator engine.
Conclusions: DG-AMMOS provides fast, automated and reliable access to the generation of 3D conformation of small molecules and facilitates the preparation of a compound collection prior to high-throughput virtual screening computations.
The validation of DG-AMMOS on several different datasets proves that generated structures are generally of equal quality or sometimes better than structures obtained by other tested methods.
Author: David LagorceTania PenchevaBruno VilloutreixMaria Miteva
Credits/Source: BMC Chemical Biology 2009, 9:6
Posted under Cell Analysis, Cell-based Assays, Compound Screening, Discoveries, Innovations and Patents, Industry News, Press Releases | Comments Off
Lumpy Assay Results
Last Updated on Monday, 11 January 2010 12:23 Written by Editor Monday, 11 January 2010 12:23
When we screen zillions of compounds from our files against a new drug target, what can we expect? How many hits will we get, and what percentage of those are actually worth looking at in more detail?
These are long-running questions, but over the last twenty years some lessons have been learned. A new paper in J. Med. Chem. emphasizes one of the biggest ones: if at all possible, run your assays with some sort of detergent in them.
Why would you do a thing like that? Compound aggregation. The last few years have seen a rapidly growing appreciation of this problem. Many small molecules will, under some conditions, clump together in solution and make a new species that has little or nothing to do with their individual members. These new aggregates can bind to protein surfaces, mess up fluorescent readouts, cause the target protein to stick to their surfaces instead, and cause all kinds of trouble. Adding detergent to the assay system cuts this down a great deal, and any compound that’s a hit without detergent but loses activity with it should be viewed with strong suspicion.
The authors of this paper (from the NIH’s Chemical Genomics Center and Brian Shoichet’s lab at UCSF) were screening against the cysteine protease cruzain, a target for Chagas disease. They ran their whole library of compounds through under both detergent-free and detergent conditions and compared the results. In an earlier screening effort of this sort against beta-lactamase, nearly 95% of the hits (many of them rather weak) turned out to be aggregator compounds. This campaign showed similar numbers.
There were 15 times as many apparent hits in the detergent-free assay, for one thing. Some of these were apparently activating the enzyme, which is always a bit of an odd thing to explain, since inhibiting enzyme activity is a lot more likely. These activators almost completely disappeared under the detergent conditions, though. And even looking just at the inhibitors, 90% of the hit set in the detergent-free assay went away when detergent was added. (I should note that control cruzain inhibitors performed fine under both sets of assays, so it’s not like the detergent itself was messing with the enzyme to any significant degree).
They point out another benefit to the detergent assay – it seems to improve the data by keeping the enzyme from sticking to the walls of the plastic tubes. That’s a real problem which can kick your data around all over the place – I’ve encountered it myself, and heard a few horror stories over the years. But it’s not something that’s well appreciated outside of the people who set up assays for a living (and not always even among some of them).
So, let’s get rid of those nasty aggegators, right? Not so fast. It turns out that some of the compounds that showed this problem during the earlier beta-lactamase work didn’t cause a problem here, and vice versa. Even using different assays designed to detect aggregation alone gave varying results among sets of compounds. It appears that aggregation is quite sensitive to the specific assay conditions you’re using, so trying to assemble a blacklist of aggregators is probably not going to work. You have to check things every time.
One other interesting point from this paper (and the previous one): curators of large screening collections spend a lot of time weeding out reactive compounds. They don’t want things that will come in and react nonspecifically with labile groups on the target proteins, and that seems like a reasonable thing to do. But in these screens, the compounds with “hot” functional groups didn’t have a particularly high hit rate. You’d expect a cysteine protease to be especially sensitive to this sort of thing, with that reactive thiol right in the active site, but not so. This ties in with the work from Benjamin Cravatt’s group at Scripps, suggesting that even fairly reactive groups have a lot of constraints on them – they have to line up just right to form a covalent bond, and that just doesn’t happen that often.
So perhaps we’ve all been worrying too much about reactive compounds, and not enough about the innocent-looking ones that clump up while we’re not looking. Detergent is your friend!
Source: corante.com
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Families of Spinal Muscular Atrophy Funded Program Shows Quinazoline Compounds Give Survival Benefit in a Severe Mouse Model of SMA.
Last Updated on Monday, 28 December 2009 12:06 Written by Editor Monday, 28 December 2009 12:06
This publication, showing data from the testing of Quinazoline derivatives in a Spinal Muscular Atrophy mouse model, has been published in Human Molecular Genetics by lead author Dr. Matthew Butchbach from the laboratory of Dr. Arthur Burghes at the Ohio State University.
The generation of the Quinazoline compounds as a therapeutic drug candidate for Spinal Muscular Atrophy was fully funded by Families of SMA.
The paper explores whether the Quinazoline compounds, which increase the expression of SMN2, are useful as potential therapeutics for SMA. Ultrahigh-throughput screening identified substituted Quinazolines as potent SMN2 inducers. The drug-like properties of the initial screening hits were optimized through directed medicinal chemistry. This resulted in series of C5-Quinazoline derivatives.
Oral administration of three of these compounds (D152344, D153249 and D156844) to neonatal mice resulted in a dose-dependent increase in Smn promoter activity in the central nervous system.  The authors then examined the effect of these compounds on the progression of disease in SMNDelta7 SMA mice. Oral administration of D156844 significantly increased the mean lifespan of SMNDelta7 SMA mice by approximately 21-30% when given prior to motor neuron loss. Overall the authors summarize that the quinazoline derivative D156844 increases SMN expression in neonatal mouse neural tissues, delays motor neuron loss at PND11, and ameliorates the motor phenotype of SMNDelta7 SMA mice.
“This is the first compound series to go from hit-to-preclinical candidate that shows favorable pharmacology in the nervous system and shows benefit to severe SMA mice. This study shows that promising therapies for SMA can be developedâ€, said Matthew Butchbach, Ph.D., who is lead author on this publication.
“Families of SMA is pleased that the first test of this class of compounds in SMA mice shows potential therapeutic benefit. The clinical lead in this series called Quinazoline495, which is a more optimized compound than those tested here, has also been assessed in this animal model with similar results, as well as tested in a slightly less severe mouse model of SMA, in which it showed marked enhancement of survival”, says Jill Jarecki, Ph.D., FSMA research director.
The lead compound Quinazoline495 recently received orphan drug designation from the FDA for the treatment of spinal muscular atrophy. Please click here to read more.
Families of SMA recently licensed this series of compounds to Repligen Corporation for development as a drug treatment for Spinal Muscular Atrophy.
The full reference:
Butchbach ME, Singh J, Thornorsteinsdóttir M, Saieva L, Slominski E, Thurmond J, Andrésson T, Zhang J, Edwards JD, Simard LR, Pellizzoni L, Jarecki J, Burghes AH, Gurney ME. Effects of 2,4-diaminoquinazoline derivatives on SMN expression and phenotype in a mouse model for spinal muscular atrophy. (2009). Human Molecular Genetics, Epub ahead of print.
Source: fsma.org
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Enzyme binds both sides of the mirror
Last Updated on Monday, 28 December 2009 11:30 Written by Editor Monday, 28 December 2009 11:30
European chemists have discovered that both mirror-image forms of a particular compound can bind at the same time in the same site of an enzyme, a phenomenon that has never been seen before. The finding has significance for drug discovery screening and studies of how small molecules interact with proteins.
Rolf Breinbauer from Graz University of Technology, Austria, and Wulf Blankenfeldt from the Max Planck Institute of Molecular Physiology in Dortmund, Germany, were studying a metabolic enzyme from a species of the bacterium Burkholderia cepacia, using racemic mixtures of chiral probe molecules to find ones that bound in the enzyme’s active site. In most cases only one form of a chiral (or ‘handed’) molecule would bind at once, but they found that in one instance both enantiomeric forms occupied the binding site at the same time.
‘If you read the textbooks about enantiomers,’ says Breinbauer, ‘there’s a simplified notion that one enantiomer is good and the other is either bad or just idle.’ He explains that for most proteins (apart from certain enzymes that have evolved to cope with wide ranges of substrate molecules) either only one enantiomer will bind, or both can bind individually - with the assumption that one form will be significantly more active than the other. ‘Our findings show that the world is more complicated,’ he adds.
While each individual enantiomer can bind to the enzyme seperately, Breinbauer notes that the arrangement of the molecules within the binding site is quite different when both bind together. This could lead to cooperative effects, producing either an enhanced or diminished response relative to the individual enantiomers.
|
The three ways enantiomers can bind in enzymes: only one enantiomer binds (top); each binds individually (middle); both bind together (bottom)
© Angewandte Chemie
|
He adds that this could have relevance in drug discovery screening, where mixtures of both enantiomers of chiral compounds are routinely screened together to find initial hits. ‘People need to consider more options when interpreting binding data from racemic mixtures.’
Dafydd Owen of Pfizer Research Chemistry in Sandwich, UK, agrees that the finding is an important reminder that chemists need to be open-minded about interpreting screening data. It also highlights the inherent trade-offs made when screening mixtures - particularly in high-throughput screens when mixtures of several compounds are tested at once.
Owen sees most interest in the discovery in the area of fragment-based drug discovery, where small ‘fragment’ molecules found to bind to a drug target are linked together to make potential drug molecules. ‘As a medicinal chemist,’ he adds, ‘my immediate thought was to join the two structures together to incorporate the best of each and make a hybrid.’ He points out, however, that from a fragment point of view it is almost irrelevant to the enzyme that the two molecules happen to be mirror images of each other, ‘despite their apparent similarity, nature views enantiomers as very different molecules’.
Phillip Broadwith
Source: rsc.org
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UT professor receives grant for new process
Last Updated on Wednesday, 16 December 2009 10:59 Written by Editor Wednesday, 16 December 2009 10:59
Using a pair of tweezers, a UT graduate student carefully lifted a nylon mesh square about the size of a thumbnail out of a small flask in his team’s lab.
The nylon had been soaking in a clear, watery solution containing a chemical compound — the “capture agent†— that it would bind with during a process tweaked by the student and his team.
Jennifer Brodbelt, a chemistry and biochemistry professor, received a $734,068 grant from the National Institutes of Health Oct. 1 to further develop the process — Desorption Electrospray Ionization (TM-DESI) — and perfect the nylon squares which isolate desired compounds from solutions.
Brodbelt, UT graduate students and two professors from Southwestern University in Georgetown were given a two-year deadline to gather blood from people of varying ages and levels of health, and to develop a more efficient method of analyzing the samples.
Results will be used to spot trends in the frequency of certain biological compounds, including amino acids.
A mass spectrometer, the machine Brodbelt’s team uses, can identify specific compounds in a mixture like blood. The tricky part was getting the sample to spray into the machine.
Joe Chipuk, a graduate student currently working on the project, was struck by the idea of having samples sprayed directly through a sifting material into the spectrometer.
Chipuk ran home and began collecting mesh materials to spray water through. He cut up his screen door, his wife’s tea strainer and the aerator from his kitchen faucet.
He went outside, used a hose to spray water through the mesh materials and observed the water’s exit path.
He then drafted a plan to create a mesh material soaked in a chemical that allows certain compounds to travel through but traps enough as to not let every metabolite escape.
After the unwanted materials are sorted out, the desired compounds attached to the mesh are released and analyzed by the spectrometer.
Before Chipuk’s square, the desorption process played out very much like a complicated billiards shot. The spray came down at an angle, hit the slide holding the blood sample and ricocheted off carrying the compounds through the spectrometer.
The new technique allows the compounds to be sorted and analyzed at a much faster pace than before. Chipuk said they can now analyze 50 samples in approximately eight minutes, whereas before, analyzing 50 samples would have taken more than 24 hours.
The team is focusing on improving the reliability and consistency of the mesh squares, Brodbelt said.
“The hope is that this could be a way to diagnose patterns of disease or determine a prognosis based on the pattern of metabolites,†Brodbelt said. “The sooner you have an idea that you might have cancer, or that you are on track to develop cancer, you could have screening done earlier and more frequently.â€
Source: dailytexanonline.com
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