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Chemical Biology
Intervention in Cells Based on Chemical Principles
Chemical Biology
Intervention in Cells Based on Chemical Principles
Keynote Speaker: Stuart L. Schreiber (Harvard University) Presented by the Hunter College Center for Study of Gene Structure and Function, Research Centers in Minority Institutions Program of the National Center for Research Resources, NIH, and the Chemical Biology Discussion Group Posted March 17, 2005 Overview
The nascent field of chemical biology employs small molecules to study the biology of disease. Scientists are using the methods of chemical biology to understand the principles that underlie cell activity, and, ultimately, to develop new pharmaceuticals and therapies.
The breadth of topics presented by seven researchers during a symposium hosted by the Hunter College Center for Study of Gene Structure and Function reveals just how expansive the field of chemical biology has become. Topics ranged from how RNA interference could be used to prevent HIV infection, to how traditional healers in the Amazon concoct a termite-derived Viagra-like compound, to how a private company is using a new technology platform to redesign chemical entities to interact more flexibly with a diseased system.
Use the tabs above to find a meeting report and multimedia from this event.
Sponsorship
The Gene Center is supported by the Research Centers in Minority Institutions program of the National Center for Research Resources, at the National Institutes of Health on grant number G12 RR-03037.
Introduction
The nascent field of chemical biology employs small molecules to study the biology of disease. Scientists are using the methods of chemical biology to understand the principles that underlie cell activity, and, ultimately, to develop new pharmaceuticals and therapies.
The breadth of topics presented by seven researchers during a recent symposium hosted by the Hunter College Center for Study of Gene Structure and Function reveals just how expansive the field of chemical biology has become. Topics ranged from how RNA interference could be used to prevent HIV infection, to how traditional healers in the Amazon concoct a termite-derived Viagra-like compound, to how a private company is using a new technology platform to redesign chemical entities to interact more flexibly with a diseased system.
Two lecturers—Eloy Rodriguez and Akira Kawamura—discussed the application of chemical biology to the study of herbal medicines or natural products. As Cornell University professor Rodriguez noted, “Natural products have been refined over billions of years of biological selection. We can regard natural products as a combinatory library made by nature and it’s not just a diverse library of compounds, but it’s also biologically already selected.” His research in Amazonian and Caribbean ecosystems has turned up evidence of how chemical compounds cascade through a food chain to protect animals from disease.
Hunter’s own Kawamura showed how his laboratory is employing DNA microarray technology to screen the compounds in traditional Japanese Kampo medicines.
In a twist on that theme, Jose Salas of Spain’s University of Oviedo presented research into “unnatural natural products”—improvements upon natural products that can be generated by re-engineering the pathways of microorganisms that produce bioactive compounds. Salas’ team studies the biosynthetic pathways of natural products in order to engineer biosynthetic pathways for different antitumor drugs.
On the same re-engineering tack, Surface Logix cofounder Carmichael Roberts explained a novel nanotechnology developed at Harvard that his company is using to improve upon existing chemical entities.
Judy Lieberman of Harvard Medical School described how her lab is developing new methods for preventing and possibly treating HIV infection with short interfering RNAs, or siRNAs.
And Scripps Institute researcher Benjamin Cravatt presented his lab’s work into activity-based protein profiling, a technology involving the use of chemical probes to bind and label enzymes in order to characterize their function.
The event keynote came from the father of chemical biology himself, Harvard chemist Stuart Schreiber who gave a state-of-the-discipline address that explained the global goals of chemical biologists and explained his significant contribution to the field: diversity-oriented synthesis.
The Current State of Chemical Genetics
Speaker:
Stuart L. Schreiber, Harvard University
Highlights
- Pioneers in the emerging field of chemical biology aim to create an inventory of all naturally occurring small molecules and a collection of small molecules comprising a small-molecule modulator for each individual function of all human macromolecules.
- Synthetic chemists worldwide are collaborating to create and screen sets of compounds derived by a method called diversity-oriented synthesis.
- More than a million diversity-oriented synthesis compounds will be made publicly searchable according to structure or substructure similarity through the new ChemBank online repository.
What is chemical biology?
Stuart Schreiber is widely acclaimed for defining a new field of chemistry called chemical biology, which he describes as “the study of small molecules in the context of living systems.” Advances in chemistry have enabled the use of small-molecule modulators to explore biology and medicine in a systematic rather than an ad hoc way, Schreiber says.
Chemical biology expands the central dogma of biology—which holds at its core the relationships between macromolecules, DNA, RNA, and proteins—to consider the small molecules that are involved in memory and cognition. “Right now, in your brain, there are small molecules that are enabling you to process information, sensing and signaling,” Schreiber says. “They’re involved in the origins of life.”
Advances in chemistry have enabled the use of small-molecule modulators to explore biology and medicine systematically.
Schreiber defines two goals of chemical biology: “First, to complete the inventory of all naturally occurring small molecules and identify a small-molecule modulator for the individual functions of all the macromolecules; and second, to probe cell circuitry and disease biology in a way that will bridge the long-lamented gulf between basic and clinical research.”
Toward the goal of building a more comprehensive collection of small molecules, Schreiber and his team are promoting three tools in tandem: diversity-oriented synthesis, multidimensional screening, and ChemBank, a public data repository and in silico environment for analysis of small molecules and assay measurements.
As opposed to the traditional target-oriented synthesis, which screens small molecules for their ability to bind to a selected protein target, diversity-oriented synthesis, or DOS, screens small molecules without regard for a target but assessing the ability to modulate a biological pathway. With DOS, chemical biologists are able to form complex and diverse small molecules efficiently, in a few high-yield steps. Schreiber says there now exist DOS-derived small molecules with rare and specific influence on living systems, but that the technique remains to be perfected. Ultimately, to yield molecules that target specific biological pathways, diversity-oriented synthesis must be augmented by information science, he says.
There is another big distinction between diversity- and target-oriented synthesis, Schreiber notes. “In target-oriented synthesis, a stereochemical array is defined for you generally by nature in the form of defining the structure of a natural product, and you aim to devise a synthesis that yields that stereoisomer. In diversity synthesis, you aim to devise the synthesis that systematically permutes every possible stereocenter.”
Skeletal diversity
By combining DOS organic chemistry with specific computational methods (which he hopes to make accessible online through ChemBank later this year), Schreiber is generating small molecules that could become probes for comprehensively exploring the function of all the protein class of macromolecules.
His intent is to generate a distributed collection of small molecules having stereochemical diversity. Schreiber notes that starting from a single compound, there are a number of ways, in just three or four steps, to obtain products with completely different atomic skeletons.
There are a number of ways to obtain products with completely different atomic skeletons.
One is a differentiating process that he says is “not unlike an undifferentiated cell receiving signals and becoming a more differentiated cell.” Certain organic functionalities have the ability to undergo many kinds of transformations and generate many different skeletons, Schreiber says. One example is an unsaturated alkynal boronic acid that under oxidizing conditions yields an amino, and under hydroxy alkylating conditions yields an alanine.
Another skeleton-generating method is a folding process, called so for the way its polypeptide chain folds to its three-dimensional structure when exposed to a “folding buffer.”
And, Schreiber says, the ability to make skeletons combinatorially came about when workers in his lab realized that by putting different substituents onto furan rings, and tweaking the electrophilicity of the intermediate, they could generate additional skeletons through a single folding condition.
In the past year, Schreiber has coordinated synthetic chemists worldwide to use these methods to generate collections of compounds that he calls “DOS diversity sets.” The scientists have generated three sets so far, and Schreiber says they will begin screening the compounds this April.
Small-molecule screening
Through screening of these commercially available, flat, drug-like compounds, Schreiber says his group has been able to find with “concerning frequency”— in more than 300 compounds—small molecules that disrupt or stabilize microtubules. With DOS compounds, no such compounds were found. Since these are known to be general toxins, their absence in a DOS diversity set is encouraging, especially since the same set has yielded many compounds having highly specific and desirable properties.
Schreiber insists on making all of the compounds developed in his lab widely available without intellectual property restrictions. Though his research is not focused on drug discovery, he says, it “opens up opportunities for drug discovery.”
For instance, pharmaceutical companies are now developing cancer therapies with the same target as monastrol, a compound discovered in Schreiber’s lab that blocks cell division.
And the compound tubacin, the tubulin deacetylase (HDAC6) inhibitor discovered by his group through a multidimensional chemical genetic screen of more than 7,000 small molecules, has been made available to a large number of researchers, ranging from neuroscientists interested in cell motility of neurons, to people working in areas that Schreiber says he “didn’t even know existed.”
Pharmaceutical companies are developing cancer therapies with the same target as monastrol.
In an example of how discoveries from chemical biology bridge basic and clinical research, Schreiber says a study that was published last year showed how tubacin affects the function of aggresomes—the region of the cell where protein aggregates collect. In another example, his lab’s investigation into a molecule related to protein degradation has led to an approach to the clinical treatment for multiple myeloma, an incurable malignancy associated with massive overproduction of protein cells.
Compounds in the public domain
ChemBank, a new data repository for structures and assay measurements, is the vehicle through which Schreiber will make about a million DOS compounds publicly available and searchable according to structure or substructure similarity. Demonstrating how assay measurements could be accessed, Schreiber says ChemBank offers a heat map generated from raw data that enables a user to click over the map and obtain the structure that has the effect denoted by color intensity on the map.
Schreiber says he views screening as a multidimensional undertaking: “Every experiment involves selecting from different cell lines and cell states, selecting different assay measurements, different small molecules, and different concentrations. Increasingly, what we’re trying to do is perform a screening like this in a multidimensional node where we vary all of those parameters simultaneously.”
Like GenBank, the public repository of genome sequence data, Schreiber says ChemBank will provide a large matrix of data with tools that can be applied for statistical analysis and pattern detection.
Engineering Biosynthetic Pathways for Natural Products
Speaker:
Jose Salas, University of Oviedo, Spain
Highlights
- So-called “unnatural natural products” are improvements on natural products that can be generated by re-engineering the pathways of microorganisms that produce bioactive compounds.
- Bacteria known as actinomycetes produce 93 percent of antitumor compounds.
- A biosynthetic improvement on the natural product mithramycin is 80 times more potent as an antitumor compound than the original.
- A new cassette plasmid system can be used to produce novel deoxysugars in a producing organism and to alter the sugar profile of natural products.
Natural-product machines
Microbes, plants, and animals produce a large variety of biologically active natural products, many of which have clinical, veterinary or agricultural applications.
Actinomycetes are the most important bacterial group synthesizing natural products. Of natural products generated by microorganisms, actinomycetes are responsible for 60 percent of antifungals and 64 percent of all bioactive natural products. They are of even greater importance to the production of antitumor compounds, of which they are responsible for 93 percent.
Bioactive compounds produced by actinomycetes.
Specifically, these natural products include the antibacterials erythromycin, tetracyclines, and gentamicin; the antitumor compounds doxorubicin, mithramycin, and bleomycin; and also antifungals, insecticides, antiparasite compounds, and herbicides.
Jose Salas, whose lab at University of Oviedo studies the biosynthetic pathways of natural products, is especially interested in engineering biosynthetic pathways for different antitumor drugs produced by actinomycetes, and in particular streptomycetes.
Classically, strategies for isolating bioactive compounds from microorganisms have been implemented through pharmaceutical company and academic screening programs. Strains are improved through mutagenesis to select mutants with high titers for production, and organic chemists contribute to the process either by generating synthetic compounds or by tweaking the fermentation process to generate semisynthetic compounds.
Combinatorial biosynthesis
In the last few years, the development of new strategies has enabled scientists to manipulate microorganisms to produce what Salas called "unnatural natural products." His lab is pursuing an approach called combinatorial biosynthesis to get microorganisms that typically produce bioactive compounds to generate novel compounds in place of their normal products.
Combinatorial biosynthesis.
Salas said that DNA recombinant technology is allowing the genetic engineering of natural-product biosynthesis gene clusters that will result either in improved production yields or in the generation of novel derivatives with higher bioactivity, lower toxicity, or better therapeutic indexes. Salas's lab and others have characterized clusters of genes involved in the synthesis of several compounds and have used combinatorial synthesis to generate new derivatives.
How does combinatorial biosynthesis work? Salas described the process, beginning with a microorganism that is known to produce a bioactive compound. Through genetic or metabolic engineering, recombinant strains can be generated in which specific genes of the biosynthetic cluster are inactivated. Blocking production of the bioactive compound in this fashion can force the microorganism to generate a novel bioactive compound instead. Researchers can then spread a set of genes from another biosynthetic cluster to block production of the native compound to result in a new compound.
Novel antitumor compounds
The antitumor compounds that Salas is focused on are derived from several sources: borrelidin, an angiogenesis inhibitor; mithramycin and chromomycin, two DNA replication and transcription inhibitors; and two indolocarbazole alkaloids, rebeccamycin, a DNA topoisomerase inhibitor, and staurosporine, a protein kinase inhibitor.
Salas discussed how his lab has developed novel bioactive compounds related to some of these agents. In one example, he showed how the scientists worked with the organism that produces mithramycin to inactivate the mtmW gene, resulting in the accumulation of a novel compound, mithramycin SK, that has a side chain with one less carbon than mithramycin's and is 80 times more potent as an antitumor agent, with strong activity against melanoma and colon cancer cells.
Borrelidin was discovered to have antibacterial properties more than 50 years ago. Later it was shown to exert antiangiogenesis activity by inducing apoptosis. By analyzing the chemical structure of borrelidin, Salas could predict how it is synthesized by a microorganism and thereby develop a biosynthetic gene cluster.
By analyzing a group of genes, borA1-borA6, encoding the subunits of a modular polyketide synthase (PKS) protein that helps synthesize borrelidin, Salas and his colleagues discovered that the genes contain large DNA repeat sequences and that the number of genes is smaller than the number of protein modules predicted. They showed that mutants with in-frame translational fusions between different borA PKS genes still produce borrelidin, indicating that a PKS module was being used iteratively.
Cassette.
By interfering with the genes involved in cyclopentane dicarboxylic acid biosynthesis genes and in nitrile formation, two other processes required for the synthesis of borrelidin, Salas' group was able to induce the biosynthesis of two novel compounds: 12-desnitrile-12-methyl-borrelidin (12-DNFB) and 12-desnitrile-12-carboxyl-borrelidin (12-DNMB). Both have biological activity far exceeding borrelidin's as an inhibitor of tumor growth and chemotaxis.
Salas's group has also developed novel compounds derived from the organisms that form rebeccamycin and staurosporine. The group has used combinatorial biosynthesis to generate 32 different indolocarbazoles compounds.
Glycosylation
Salas also explained how his lab is altering the glycosylation pattern of some bioactive compounds. Because many natural products are glycosylated compounds, Salas said his lab has also developed "plug and play" cassette plasmid systems as a tool for producing novel deoxysugars in a producing organism. By taking advantage of these plasmids and of the "substrate flexibility" of glycosyltransferases, they are able to alter the sugar profile of natural products.
Genomic Screening of Natural Products
Speaker:
Akira Kawamura, Hunter College, CUNY
Highlights
- Conventional approaches to investigating natural-product compounds rely on cytotoxicity studies, but these overlook the many natural compounds that are effective in treating disease without being cytotoxic.
- Testing a natural compound against cells on a DNA microarray can reveal whether it affects gene regulation.
- These methods are providing researchers with new information about the bioactivity of herbal medicines.
Beyond cytotoxicity
Akira Kawamura’s lab, at the department of chemistry at Hunter College of the City University of New York, is taking new approaches to the investigation of molecules from natural resources—organisms including medicinal plants, microorganisms, and various marine organisms—in an effort to find novel organic compounds.
Natural products are rich resources of molecular probes for biomedical researchers, but much of what is presently known about natural-product compounds is based on phenotypic activities. Traditional methods for identifying bioactive molecules rely on a fractionation and subfractionation procedure that screens for cytotoxicity.
Organic molecules that have unique biological properties are overlooked by this kind of screening.
But as Kawamura pointed out, if you use cytotoxic assays to look at and guide the purification of natural products, you’re going to end up with only cytotoxic compounds. Many organic molecules that have unique biological properties, yet are devoid of strong cytotoxicity, are overlooked by this kind of screening. “In the long history of natural-products chemistry, people probably missed a lot of interesting compounds in the initial stage of screening because of a lack of cytotoxicity,” he said.
Putting herbs on chips
Kawamura’s lab is using DNA microarray technology to identify interesting compounds in natural products that have been previously unexplored. His hypothesis is that the information derived from microarray gene-expression profiling will expand on what can be learned from cytotoxicity screens.
In the first phase of this approach, his team uses gene-expression profiling to identify targets of a natural product. Sprinkling an herbal medicine mixture of interest onto human cells that are hybridized to an array triggers an up- or down-regulation of certain genes. The results reveal which genes’ expression can be affected by treatment with the particular product.
Once differentially expressed genes are identified through these studies, the lab can use more versatile and less expensive methods to guide the purification of active components. With target genes identified, Kawamura fractionates the original mixture and then cultures the cells of interest on a 96-well plate, treats the cells with the fractions of interest, and uses reverse-transcriptase polymerase chain reaction (RT-PCR) to identify the fractions that are responsible for the regulation of those genes.
To reduce the cost and potential sources of errors in the RT-PCR screening, Kawamura’s team developed a simpler, less expensive screening method: by skipping the purification step, starting from cell lysate, and going directly go to cDNA synthesis, he reduces the number of experimental steps.
Screening compounds in Kampo medicines
With this protocol in hand, the lab began investigations into a variety of Japanese herbal medicines known as Kampo. One such medicine, keishibukuryogan (keishi), is used to treat conditions related to blood vessels. Kawamura’s lab conducted gene expression profiles by screening keishi against human endothelial cells on a microarray. To test the validity of the genomic technology approach, the lab compared cytotoxicity and RT-PCR assays to identify novel fractions, using background controls to eliminate false positives.
The lab compared cytotoxicity and RT-PCR assays to identify novel fractions.
Demonstrating the comparison between the two screenings, Kawamura noted that some of the fractions are the same, but that many other fractions were detected only by the RT-PCR procedure, proving his hypothesis that a DNA microarray profile can provide access to unique fractions that cannot be detected by the cytotoxicity assay. Among these fractions, there are three that Kawamura’s lab is now particularly interested in for further studies.
Kawamura said his studies are “proof of concept.” Because natural products can now be screened with the DNA microarray technology, he noted, “We can now look at the natural products from a very different angle and identify fractions.”
“In our fractionation study, a screening study, we identified fractions to screening and it set us in a unique, a new position to do the further fractionation and screening,” said Kawamura. “This of course opens up an opportunity for the identification of novel compounds and biological properties.”
Biomedicinal Chemistry of Tropical Small Molecules
Speaker:
Eloy Rodriguez, Cornell University
Highlights
- Extraordinary chemodiversity existing among plants and insects of the Amazon and Caribbean forests provides templates for the synthesis of unique and complex natural products.
- The discovery of “chemical defense cascades” explains how plants become the source of compounds that are present in insects and birds.
- Natural products found in the Amazon and even in New York show promise as treatments for modern infectious diseases.
Chemodiversity frontiers
Cornell University biological chemist and professor of environmental studies Eloy Rodriguez called the Amazon rainforest and Caribbean cloud forests and coral reefs “the last frontiers of an extraordinary chemodiversity in the natural world.” Modern pharmaceutical drugs are produced primarily through synthetic combinatorial chemistry, but these natural environments, where Rodriguez maintains research laboratories, provide templates for the synthesis of unique and complex natural products, he said.
Molecules are nature’s medium for communicating, argued Rodriguez, who is known for having established the discipline of zoopharmacognosy to describe the study of animals’ observed ability to self-medicate and to protect themselves from disease with natural products. Now, his research in remote tropical forests has uncovered chemical compounds that function in insect-insect mating and repulsion, and others that are secreted by birds to kill mites, bacteria, and viruses. Rodriguez and his colleagues have also discovered that certain molecules alter gene expression, especially in the production of active polypeptides.
Any given tree in the forest is host to as many as a half million or more natural products, Rodriguez said. These are found in the seeds, the leaves, the flowers, the roots, the stems, the bark, and associated fungi, symbionts, and insects. One insect alone can make 125 compounds, he notes. “You get 1,000 of them, you’ve got 125,000 compounds! There is an incredible amount of chemistry being produced and. . . these are compounds that must have some function, and this is what’s intriguing about going into the forest.”
Chemical defense cascades
Rodriguez has coined the term “chemical defense cascade” to describe the way in which plants become the source of compounds that are present in some insects and birds. One example is a discovery Rodriguez and associates made by conducting mass spectrometry analysis of secretions coating the feathers of a tropical bird in the Amazon.
Chemical defense cascades.
An alkaloid in the secretion, which also contains antibiotic fatty acids that protect the bird against mites and bacterial infection, resembles an alkaloid produced by a local plant. But the bird doesn’t feed on the plant or even on seeds—it feeds primarily on ants. And the compound is one that, biosynthetically, the ant could not have produced. As it turns out, the ants that this bird eats acquire liquid from aphids, which take this very compound from plants.
In this way, the chemical compound cascades from plant to aphid to ant to bird and then is employed by the bird as a defense mechanism, Rodriguez said. “Now we can see that plants are really central to the production of certain chemical defensive systems through this cascade effect.”
Insect and plant compounds
In fact, insects produce many kinds of compounds. Some, such as hydrogen cyanide generated by a millipede, are also produced independently by insects and plants. Others, such as pheromones produced by ants and other insects, or compounds stored by caterpillars and incorporated into butterfly wings, are likely derived from the diet, Rodriguez said.
Cacique yellow feathers with lipids.
Even when a plant- and an insect-generated compound are identical, both could be producing the compound independently using very different biosynthetic pathways. Rodriguez hypothesizes that some more complicated compounds found in plants, where the biosynthesized compounds look identical to those from fungi, could be results of the incorporation of a fungal genome into a plant genome.
Termites for potency, Clitoria for malaria
In another example of how synthetic compounds can mimic those found in nature, Rodriguez described a Viagra-like medicinal cocktail that the women of an Amazon tribe concoct by grinding ants, termites, and hot fruits. The mixture contains amines, testosterone, and diverse steroidal chemicals. “Whatever is going on here, it’s very effective and it works,” said Rodriguez, “and what really fascinates me is how the heck they came up with this mixture! They’re very particular about the termite species. You cannot use just any termite.”
Rodriguez said that he is particularly interested now in compounds from plants that could be useful in controlling diseases such as malaria, dengue fever, and Helicobacter pylori infection. An Amazonian plant genus called Clitoria that his lab is studying has a chemical makeup with activity similar to that of artemisinin, a drug used to treat malaria, as well as of a terpenoid produced by the sunflower family. The fruit of another plant that grows wild in New York has a history of being prescribed by American Mohawk Indians for eradicating ulcers. Rodriguez is studying it as a potential inhibitor of H. pylori, a bacterium involved in ulcer formation and gastric cancer.
“It’s interesting how these molecules can have new roles once you discover new ways to look at their activity,” he said.
Better Pharmaceuticals Through Biophysical Chemistry
Speaker:
Carmichael S. Roberts, Surface Logix, Inc.
Highlights
- A technology originally developed at Harvard University is being used to improve on existing chemical entities.
- Pharmacomers can be designed to minimize nonspecific interactions with proteins to improve the distribution of drugs to desired receptors and enzymes in a specific tissue or organ.
- Several compounds that were created with the platform in the last 18 to 24 months are now moving toward clinical trials.
Enhancing pharmacokinetic and pharmacodynamic profiles
Surface Logix is a drug development company that uses a proprietary technology platform to create new chemical entities. Carmichael Roberts, the company's cofounder and president, said the so-called Pharmacomer Technology Platform improves on existing chemical entities by enhancing their pharmacokinetic and pharmacodynamic profiles.
The Pharmacomer Technology Platform improves on existing chemical entities.
Based on research done in the laboratory of George Whitesides at Harvard University's Department of Chemistry and Chemical Biology, where Roberts was a National Science Foundation Fellow, the technology allows scientists to make small-molecule drugs by using biophysical chemistry and nanotechnology to analyze and identify the liabilities of a known drug in a variety of physiological microenvironments.
Surface Logix scientists correct the liabilities by replacing standard medicinal chemistry synthons, or building blocks, with proprietary small-molecule chemistries the company has named Pharmacomers.
Specific Pharmacomers, which have also been analyzed in a variety of physiological microenvironments, are chosen depending on the functionality required to improve the pharmacokinetic or pharmacodynamic liability of the known drug. Pharmacomers are designed using a set of assays, indices, and correlations that Surface Logix has devised. The approach allows the company's scientists to create new chemical entities and new clinical candidates rapidly and efficiently.
Surface Logix pipeline
Therapies for inflammation, cancer, dyslipidemia, and cardiovascular disease (including hypertension and erectile dysfunction) are among those the company has pursued. Roberts said several compounds that were created with the platform in the last 18 to 24 months are now moving toward clinical trials. The company has already filed its first Investigational New Drug (IND) application for its candidate drug to treat erectile dysfunction and expects to start human trials later this year.
What exactly is Pharmacomer Technology? Roberts described a microfabrication technique that uses stamps to mimic the patterns of anything from small molecules to cells. The platform consists of a network of capillaries on the surface of a chip and a stamp that is used to transfer ink to a pattern onto which proteins are placed.
Pharmacomers can be designed to minimize nonspecific interactions with proteins so as to improve the distribution of drugs to desired receptors and enzymes in a specific tissue or organ. They can also increase metabolic stability by minimizing breakdown by catabolic enzymes, enhance membrane permeability to barrier cells and target cells, and improve solubility in relevant biological fluids.
Pharmacomers can be designed to ... improve metabolic stability.
Roberts described how the platform was applied to the pursuit of anti-inflammatory therapies: "Ideally, you'd want a drug that blocks at least one, or ideally maybe multiple events that cause the onset of inflammation," he said. "We looked at all sorts of things on the surface to see what more could we put down that actually does the trick in terms of allowing us to work with blood cells. Some things worked a little bit, others didn't, but we found some really interesting things happen with small-molecule building blocks."
The experiments led the team to realize that there are certain classes of small molecules that can be used as pharmaceutical reagents. "Maybe by themselves they'd be interesting drugs. . . or if you attached them to small molecules that are either drugs today, or compounds that are failing clinical trials, they'd allow us to make new drugs by simply changing the molecular structure and creating a new chemical entity."
Most small-molecule drugs, he said, are very rigid, very inflexible—they're brick-like in structure. "It turns out that the opposite is true for the chemistries that we were looking at on the surfaces. The best surface chemistries that we found were the ones that were not rigid in structure."
Searching the compounds
"The way to think about this is ... the medicinal chemist is designing the molecule to bind tightly and selectively to one protein, but that protein may be sitting in the tissue of the heart, and that protein needs to make it through the stomach, it needs to make it through the intestine, it needs to get absorbed, it needs to make it past all the serum proteins in the blood, it needs to make it to the heart, it then needs to get through the interstitial tissue, and all the barriers and eventually make it to that protein that it wants to bind to."
"The best surface chemistries were the ones that were not rigid in structure."
In all, Surface Logix researchers have applied the Pharmacomer Technology Platform to characterize 1,600 small-molecule drugs that have been approved by the US Food and Drug Administration in the last 30 years. By and large, Roberts said, each of those approved drugs fit into a relatively small chemical space—unlike the bulk of the new Surface Logix chemistries, which fit in spaces traditional drugs weren't in at all, a characteristic referred to as "dynamic conformation." Dynamic conformation is used to describe both the intrinsic and extrinsic nature of molecules. Surface Logix has created thousands of molecules (that is, Pharmacomers) that occupy a wide range of dynamic conformation that can be used to improve the pharmacokinetic and pharmacodynamic profiles of drugs and late-stage candidate compounds.
Building a drug is like designing a building, Roberts said: it's necessary to design the product to withstand the elements by not making it absolutely rigid, by putting in some degree of flexibility and some dynamic conformation.
Building a drug is like designing a building.
Roberts said that the Surface Logix process aims to figure out what part of a drug can be removed because it's not absolutely critical for potency or selectivity. "Let's say we drop a drug from 600 molecular weight down to, say, 450 molecular weight. We would then use our set of assays to figure out what chemistry from these little surface chemistries can we put onto a compound and therefore generate a brand-new molecule that may address a problem that is currently in the industry, as it relates to a class of molecules, or a set of molecules."
The piece removed was almost always the part that made the molecule extremely rigid and brick-like, Roberts said. And when it was replaced with Surface Logix's own chemistry, a little more dynamic conformation was introduced into the molecule.
Activity Based Protein Profiling for Proteomics
Speaker:
Benjamin F. Cravatt, The Scripps Research Institute
Highlights
- Activity-based protein profiling (ABPP) is a technology in which chemical
probes are used to bind and label enzymes in order to characterize their
function.
- ABPP can be used to identify novel disease-associated enzymes and enzyme
activities, such as those that vary in models of human cancer and primary tumor
specimens.
- ABPP is useful as a screen to discover potent and selective reversible
enzyme inhibitors.
- ABPP is currently carried out on gel platforms, but merging the technology
with high-resolution tools such as liquid chromatography mass spectrometry would
permit multiplexing of ABPP probes.
Characterizing enzymes
The field of proteomics aims to characterize protein function on a
system-wide scale. But for several classes of enzymes, conventional proteomics
techniques are not ideal characterization methods.
Strategies used for genomic and proteomic functional characterization measure
changes in the abundance of biomolecules and changes in expression that permit
an inference to be made about changes in protein activity. Most enzyme classes,
however, are regulated by posttranslational mechanisms—they’re produced by the
cell in some form that affects their function after synthesis. They call for a
different approach to functional characterization that measures activity.
Most enzyme classes are regulated by posttranslational
mechanisms.
To address this need, several years ago Benjamin Cravatt's group at the
Scripps Research Institute developed a technology called “activity-based protein
profiling” (ABPP). The strategy uses chemical probes to assess the state of
enzyme active sites in whole proteomes and thereby functionally characterize
various enzyme classes. The probes bind and label enzymes and attach a reporter
tag that gives a readout for the enzyme’s functional state independent of all of
its other forms.
Cravatt noted that the enzyme classes addressed by ABPP fall into two general
categories: enzymes for which active site-directed affinity agents have been
well defined and enzymes for which active site-directed affinity agents have
been lacking.
Today, ABPP has matured since its introduction and is providing new methods
for identifying previously unknown disease-associated enzymes. Cravatt's group
is focused on enzyme activities that vary in models of human cancer and primary
tumor specimens. They also use the technology as a screen to discover potent and
selective reversible enzyme inhibitors.
An exquisite tool
ABPP probes are designed to contain a reactive group that binds to the active
sites of particular enzymes and a reporter tag for the detection and isolation
of these labeled enzymes. The first class of enzymes the laboratory tackled with
this approach was the serine hydrolases, whose genes comprise about 1 percent of
the human genome and which have myriad in vivo functions.
The profiling experiments, conducted in one- and two-dimensional
electrophoretic gels, look for enzymes that are differentially expressed among
the cancer cell lines and label only the active enzymes in each sample.
Probe-treated proteomes are separated on the gels and then in-gel
fluorescence scanning permits visualization of the labeled enzymes.
The profiling experiments look for enzymes that are differentially expressed among the cancer cell lines.
Even when more abundant, inactive enzymes are not labeled. Although an enzyme
that is expressed in every cell line might not be of great interest, the
activities that show selective distribution among the cell lines might be useful
to define cell phenotype, Cravatt said.
The probes developed in his lab are exquisite tools for understanding how the
cell regulates these enzymes, and they are being put to use for discovering new
disease models and targets. “Especially in our cancer research,” Cravatt said,
“we’re always trying to identify high-fidelity model systems to study.”
Applied to a panel of human breast carcinoma and melanoma cancer cell lines,
enzyme activity profiling proved useful for distinguishing a breast carcinoma
cell from a melanoma cell, or an invasive cell from a noninvasive cell. In some
experiments, Cravatt said, activity-based protein profiling revealed that, even
in simple cell biological models of cancer, the cell can use a variety of
mechanisms that regulate proteins through posttranslational modification.
The goal of Cravatt's lab is to try to identify specific enzymes from these
experiments that would be of interest for more detailed investigation. But,
Cravatt explained, activity-based protein profiling is just one tool for
characterizing enzyme function. “We’re trying to identify enzymes by a sort of
guilt-by-association model that might be involved in disease,” he said. Specific
studies to confirm or refute the roles that enzymes might play would be
required.
Selective inhibitors
Workers in the lab have also applied the ABPP technology to evaluate the
selectivity of inhibitors that target members of large families of enzymes. In
particular, they examined the proteome-wide selectivity of inhibitors of the
brain enzyme fatty acid amide hydrolase (FAAH), which regulate pain and anxiety.
In these experiments, the group was able to rapidly distinguish selective FAAH
inhibitors from those that targeted other hydrolases in the proteome. This
latter set of nonspecific inhibitors could thus be discarded before any time was
spent on animal pharmacology studies.
Challenges
To be sure, though ABPP has seen much technical improvement and has permitted
greater understanding of the role enzymes play in complex physiological and
pathological processes since it was introduced, Cravatt sees several challenges
to advancing the approach.
More probes are needed to address all enzyme and protein
classes.
To begin with, he said, more probes are needed. To address all enzyme and
protein classes will require the synthesis of more structurally diverse probe
libraries. Also, he said, extension of this technology to in vivo applications
must be considered a primary goal. The classical reporter tags are simply too
bulky to migrate through cell membranes and be distributed uniformly in animals.
Cravatt said his group has adapted “tag-free” strategies for ABPP that enable
proteomic experiments to be conducted in living cells and animals.
Cravatt said that the combined efforts of his lab and other academic and
industrial research groups have resulted in the development of probes for more
than a dozen classes of enzymes. These probes are being used to identify novel
activities associated with pathophysiological process such as cancer, metabolic
diseases, and neurological disorders. To date, most of this work has been done
with gel-based detection methods. Future ABPP endeavors include merging the
technology with higher-resolution platforms such as liquid chromatography mass
spectrometry and microarray methods, which should increase the depth of
information content obtained in functional proteomic experiments.
Harnessing RNA Interference for Therapy
Speaker:
Judy Lieberman, CBR Institute for Biomedical Research, Harvard Medical School
Highlights
- RNAi shows promise as a therapy for preventing HIV infection when delivered locally as a microbicide.
- Studies suggest that the HIV virus can be suppressed with an siRNA therapy that targets both a host co-receptor such as CCR5 and HIV genes.
- Targeting siRNAs specifically into tumor cells may also be a successful strategy.
Interfering with SNP diseases
SiRNA can target HIV gene at multiple sites.
RNA interference is an evolutionarily conserved, endogenous mechanism for sequence-specific gene silencing that uses small double-stranded RNAs called small interfering RNAs, or siRNAs, to direct cleavage or prevent translation of homologous mRNAs.
In a nutshell, RNAi works when small, double-stranded pieces of RNA composed of a sequence that is homologous to a targeted gene are able to interfere with the activity of a cell’s messenger RNA.
RNAi shows promise as a therapy for a wide variety of diseases. siRNAs are inexpensive and simple to make, and studies have shown that they have exquisite specificity for their targets and are 100 to 1,000 times more potent than antisense RNAs. In vitro and in vivo tests have demonstrated the success of highly specific targeting in which an siRNA targets a single-nucleotide polymorphism.
RNAi hurdles
For other indications, such as viral infection where there is viral variance or mutation, the high degree of specificity of siRNA might actually be a problem, said Judy Lieberman. But for HIV, which is one of the most variable and mutating genomes that exist, Lieberman’s laboratory at Harvard Medical School has been able either to identify target host genes, such as the co-receptor for HIV, or to choose highly homologous sequences and obtain protection against all the five clades of HIV and all the primary isolates.
The main challenge now for RNAi therapy is finding a way to deliver siRNAs into the cytosol of target cells in vivo without instigating off-target effects, Lieberman said. “We’ve found that we can get very good local delivery at mucosal surfaces. We’ve also found that we can deliver these molecules very efficiently into organs. . . such as the kidney, just by locally injecting the molecules into the veins draining the organs,” she said.
Lieberman’s team has also developed a method for cell-specific delivery, by which researchers can take advantage of cell surface receptors to deliver siRNAs only to cells they want to target. “For drug delivery, that’s very attractive because it means that... you don’t have to worry so much about toxicity if you’re getting the drug only where you want it to be,” she said.
Her lab was also able to protect mice from autoimmune hepatitis by hydrodynamic tail vein injection of siRNAs targeting the gene Fas, but this delivery method, she predicts, is unlikely to be adaptable for human use.
Targeting HIV
Lieberman said that her lab began exploring RNAi to see if it could be used in vitro to prevent or treat HIV infection. In fact, her lab and others have now targeted virtually every gene in HIV with RNAi. Her group has also been able to successfully target the viral host receptors and co-receptors. And they discovered that, in much the same way that drug therapy is made more effective by targeting the protease and the reverse transcriptase steps at the same time, RNAi is most effective if it targets a host co-receptor such as CCR5 and an HIV gene, Lieberman said.
Synergistic suppression of HIV replication by targeting CCR5 and P24 in primary macrophages.
By targeting the host co-receptor macrophage CCR5, Lieberman said, the group achieved very good, though not complete, suppression of HIV. Targeting an HIV gene directly achieved similarly very good but not complete infection. But targeting both resulted in complete suppression of the virus. And, Lieberman said, the silencing effect lasted much longer than expected—as long as three weeks.
Those results led Lieberman to investigate the potential of a microbicide to prevent HIV infection via sexual transmission. For delivery, Lieberman said, “We found the very simple solution that we could mix our siRNAs with a transfection reagent and get incredibly efficient introduction of siRNAs into cells.”
Though mice cannot be infected with HIV, Lieberman’s lab is testing the efficacy of delivering siRNA as a microbicide to female mice infected with the herpes simplex virus.
Systemic delivery
Systemic delivery is a bigger challenge that Lieberman’s group is tackling. In an experiment using a non-physiologically-relevant hydrodynamic injection, they targeted the host response to viral infection that leads to hepatitis. When there’s inflammation or infection in the liver, Lieberman explained, liver cells upregulate the Fas receptor. When activated lymphocytes move into the liver, they trigger apoptosis of the liver cells.
Efficient delivery of fluorescent siRNAs into liver cells.
In the case of hepatitis B and C, it’s the immune response, not the virus, that kills liver cells. Because, again, mice cannot be infected with hepatitis B or C, the lab looked to several artificial models, activating lymphocytes in vivo to go into the liver and cause hepatitis.
They achieved about 90 percent transduction of the siRNAs into liver cells in vivo with no obvious toxicity. Specific gene silencing lasted for more than 10 days. Results indicate that this treatment can be started even after destruction of the liver has begun.
Open Questions
The Current State of Chemical Genetics
How many possible DOS diversity sets can be generated?
What are the best approaches for performing multidimensional screenings of DOS compounds to permit selecting from different cell lines and cell states, different assay measurements, different small-molecules, and different concentrations?
Can the efforts of synthetic chemists worldwide be coordinated so that methods for generating DOS compounds are standardized?
Engineering Biosynthetic Pathways for Natural Products
Are we missing novel active compounds produced by combinatorial biosynthesis
because they kill the recombinant strain?
Can we alter the active site of a glycosyltransferase to accept different
substrate aglycones?
Will combinatorial biosynthesis replace other strategies for generating
novel bioactive compounds?
Genomic Screening of Natural Products
What are the active compounds in Kampo medicines?
How can we examine synergistic effects among Kampo components?
Can we use genes with low abundance protein products for screening compounds?
Biomedicinal Chemistry of Tropical Small Molecules
How were the medicinal uses of natural compounds originally discovered by
forest dwellers?
When the chemistry of plant- and insect-generated compounds are alike, how
can we determine whether they have cascaded through an ecosystem or been
produced independently?
Can the hundreds of thousands of compounds generated by nature be
systematically screened?
Activity-Based Protein Profiling: Chemical Strategies for Functional
Can ABPP technology be merged with high-resolution tools such as liquid
chromatography mass spectrometry to permit multiplexing?
How can the ABPP technology best be extended to in vivo
applications?
Can activity-based protein profiling probes be developed for all enzyme
classes?
Harnessing RNA Interference for Therapy
What is the most effective method of delivering siRNAs into the cytosol of target cells in vivo without instigating off-target effects?
Could a microbicide be effective method for introducing siRNAs to prevent HIV infection via sexual transmission?
Why does the silencing effect of siRNAs on HIV last longer when both the host co-receptor and the gene are targeted?
Web Sites
Schreiber laboratory homepage
Includes links to recent articles describing his work in The Scientist, Drug Discovery Today, Molecular Cell, and Chemical & Engineering News as well as streaming video from several lectures.
ChemBank
A freely available collection of data about small molecules and resources for studying their properties, especially their effects on biology. It is being developed to assist biologists who wish to identify small molecules that can be used to perturb a particular biological system and chemists designing novel compounds or libraries, and to serve as a source of data for cheminformatic analyses.
What is Chemical Genetics?
As defined by the Howard Hughes Medical Institute's Biointeractive website.
ICCB-CGI
The homepage of the Initiative for Chemical Genetics at the Harvard Medical School Institute of Chemical and Cell Biology.
Target-Oriented and Diversity-Oriented Organic Synthesis in Drug Discovery
PDF of a review article by Stuart Schreiber that appeared in March 17, 2000 edition of Science describing diversity-oriented synthesis for drug discovery.
Cornell University Plant Biology Department homepage
Includes links to the L.H. Bailey Hortorium, which hosts an 845,000-specimen herbarium.
EsBaran Field Station
The home page of the Amazonian research station directed by Eloy Rodriguez dedicated to education, conservation, and the discovery of novel medicinal compounds from applied field chemoecology.
Biodiversity and Biological Collections Web Server
Devoted to information of interest to systematists and other biologists of the organismic kind. Hosted by Julian Humpheries at the University of New Orleans, it includes information about specimens in biological collections, taxonomic authority files, directories of biologists, reports by various standards bodies, an archive of the Taxacom, MUSE-L and CICHLID-L listservs, and access to online journals.
Medicinal Plants page of the Environmental and Natural Resource Management project
Hosted by the Canadian International Development and Research Center, which supports research projects, networks, and other activities on medicinal plants and natural products in Asia, Africa, and Latin America and the Caribbean. Program initiatives include one on the Sustainable Use of Biodiversity Ecosystem Approaches to Human Health Small, Medium, and Micro Enterprise Innovation and Technology.
American Botanical Council's Medicinal Plant Information page
Hosts links to numerous database and plant research information resources.
The World of Kampo
Information about traditional Japanese traditional herbal medicine and a link to Kampo Today, an English-language periodical devoted to Kampo and Kampo therapeutics.
Lieberman laboratory homepage
RNAi News
Weekly paid subscription newsletter of RNAi technology, research, and business.
RNAi Net
Free news and features about RNAi technologies and research.
siRNA Patents
Website hosted by patent holder the Massachusetts Institute of Technology, with contact information for licensing siRNA technology.
Silencing Genes in HIV
Article describing the work of Judy Lieberman and others in applying RNAi to the suppression of HIV.
Cravatt laboratory homepage
Surface Logix homepage
Whitesides Research Group
Homepage of the lab that developed the technology behind Surface Logix platform technology.
Books
F. Darvas, Guttman, A. & Dorman, G., Eds. 2004. Chemical Genomics. Marcel Dekker, New York, NY.
C. M. Dobson, Gerrard, J. A. & Pratt, A.J. 2002. Foundations of Chemical Biology. Oxford University Press, Oxford, UK.
D. Engelke. 2004. RNA Interference (RNAi): The Nuts & Bolts of siRNA Technology. DNA Press, Eagleville, PA.
G. Hannon. 2003. RNAi: A Guide to Gene Silencing. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
H. Waldmann, & Janning, P. 2004. Chemical Biology: A Practical Course. John Wiley & Sons, Hoboken, NJ.
Journal Articles
The Current State of Chemical Genetics
P. R. Andreana, Liu, C. L., & Schreiber, S. L. Stereochemical control of the Passerini reaction. Org. Lett. 6: 4231-4233. Full Text
S. Schreiber. 2004. Q & A: Stuart L. Schreiber. Curr. Biol. 14: R292-R293. Full Text
B. E. Bernstein, Humphrey, E. H., Liu, C. L. et al. 2004. The use of chromatin immunoprecipitation assays in genome-wide analyses of histone modifications. Methods Enzymol. 376: 350-361. Full Text
B. E. Bernstein, Kamal, M., Lindblad-Toh, K. et al. 2005. Genomic maps and comparative analysis of histone modifications in human and mouse. Cell 120: 169-181.
B. E. Bernstein, Liu, C. L., Humphrey, E. H. et al. 2004. Global determinants of nucleosome occupancy in yeast. Genome Biol. 5: R62. Full Text | Supplementary Data
M. D. Burke, Berger, E. M. & Schreiber, S. L. 2004. A synthesis strategy yielding skeletally diverse small molecules combinatorially. J. Am. Chem. Soc. 126: 14095-14104. Full Text
M. D. Burke, & Schreiber, S. L. 2004. A planning strategy for diversity-oriented synthesis. Angew. Chemie Int. Ed. Engl. 43: 46-58. Full Text
R. A. Butcher, & Schreiber, S. L. 2004. Identification of Ald6 as the target of a class of small molecule suppressors of FK506 and their use in network dissection. Proc. Natl. Acad. Sci. USA 101: 7868-7873. Full Text
S. J. Haggarty, Wong, J. C., Clemons, P. A. et al. 2004. Mapping chemical space using molecular descriptors and chemical genetics: deacetylase inhibitors. Comb. Chem. High Throughput Screen. 7: 669-676. Full Text
J. Huang, Zhu, H., Haggarty, S. J. et al. 2004. Finding new genetic modifiers of the target-of-rapamycin (TOR) signaling pathway through chemical genetics and proteome chips. Proc. Natl. Acad. Sci. USA 101: 16594-16599. Full Text
E. L. Humphrey, Shamji, A. F., Bernstein, B. E. et al. 2004. Rpd3p relocation mediates the transcriptional response to rapamycin in yeast. Chem. Biol. 11: 295-299. Full Text | Supplementary Data
Y.-K. Kim, Arai, M. A., Arai, T. et al. 2004. Relationship of stereochemical and skeletal diversity of small molecules to cellular measurement space. J. Am. Chem. Soc. 126: 14740-14745. Full Text
S. Krishnan, & Schreiber, S. L. 2004. Syntheses of stereochemicaly diverse nine-membered ring-containing biaryls. Org. Lett. 6: 4021-4024. Full Text
M.M.-C. Lo, Neumann, C. S., Nagayama, S. et al. 2004. A library of spirooxindoles based on a stereoselective three-component coupling reaction. J. Am. Chem. Soc. 126: 16077-16086. Full Text
P. Nghiem, Kim, Y.-S. & Schreiber, S. L. 2004. Protein kinases as targets in cancer therapy: validated and emerging approaches. In C. Brenner and Duggan, D., Eds.: 293-315. Oncogenomics: Molecular Approaches to Cancer. John Wiley & Sons, Hoboken, NJ.
H. Oguri, & Schreiber, S. L. 2005. Skeletal diversity via a folding pathway: synthesis of indole alkaloid-like skeletons. Org. Lett. 7: 47-50. Full Text
R. T. Peterson, Shaw, S. Y, Peterson, T. A. et al. 2004. Chemical suppression of a genetic mutation in a zebrafish model of aortic coarctation. Nat. Biotechnol. 22:595-599. Full Text
D. R. Schmidt, Kwon, O. & Schreiber, S. L. 2004. Macrolactones in diversity-oriented synthesis: preparation of a pilot library and exploration of factors controlling macrocyclization. J. Comb. Chem. 6: 286-292. Full Text
S. Schreiber. 2001. Target-oriented and diversity-oriented organic synthesis in drug discovery. Science 287: 1964-1969. Full Text
S. J. Taylor, Taylor, A. M. & Schreiber, S. L. 2004. Synthetic strategy toward skeletal diversity via solid-supported, otherwise unstable reactive intermediates. Angew. Chemie Int. Ed. Engl. 43: 1681-1685. Full Text
J. C. Wong, Sternson, S. M., Louca, J. B. et al. 2004. Modular synthesis and preliminary biological evaluation of stereochemically and skeletally diverse 1,3-dioxanes. Chem. Biol. 11: 1279-1291. Full Text
Engineering Biosynthetic Pathways for Natural Products to Generate Novel Antitumor Derivatives
C. Méndez & Salas, J. A. 2001. Altering the glycosylation pattern of bioactive compounds. Trends Biotechnol. 19: 449-456.
C. Mendez & Salas, J. A. 2005. Engineering glycosylation in bioactive compounds by combinatorial biosynthesis. Ernst Schering Res. Found. Workshop 51: 127-46.
C. Méndez, Weitnauer, G., Bechthold, A. et al. 2000. Structure alteration of polyketides by recombinant DNA technology in producer organisms—prospects for the generation of novel pharmaceutical drugs. Curr. Pharmaceutical Biotechnol. 1: 355-395.
J. A. Salas & Méndez, C. 1998. Genetic manipulation of antitumor-agent biosynthesis to produce novel drugs. Trends Biotechnol. 16: 475-482.
Biomedicinal Chemistry of Tropical Small Molecules with Life-Altering Activities
Aregullin, M. & E. Rodriguez. 2000. Hydrophyllaceae. In J. Avalos, & Maibach, H., Eds.: 187-199. Dermatological Botany. CRC Press LLC, Boca Raton, FL.
Berry, J., McFerren, M. & Rodriguez, E. 1996. Zoopharmacognosy. A biorational approach to chemical prospecting. In D. Gustine, & Flores, H., Eds.: Phytochemicals in Health. Pennsylvania State University Press, University Park, PA.
Freeman, F., M. Aregulin & E. Rodriguez. 1993. Naturally occurring 1,2-dithiins. Rev. Heteroatom Chem. 9: 1-15.
Rodriguez, E. & J. West. 1994. The tropical rain forest: a rich source of natural medicines. Proceedings of the NIGMS-NIH Symposium, Atlanta, Georgia.
Rodriguez, E. & R. Wrangham. 1993. Zoopharmacognosy: the use of medicinal plants by animals. In H. Stafford, & Downum, K., Eds.: 89-105. Recent Advances in Phytochemistry. Plenum Press, New York, NY.
Activity Based Protein Profiling for Proteomics
G. C. Adam, Burbaum, J., Kozarich, J. W. et al. 2004. Mapping enzyme active sites in complex proteomes. J. Am. Chem. Soc. 126: 1363-1368. Full Text
K. T. Barglow, & Cravatt, B. F. 2004. Discovering disease-associated enzymes by proteome reactivity profiling. Chem. Biol. 11: 1523-1531. Full Text
N. Jessani, & Cravatt, B. F. 2004. The development and application of methods for activity-based protein profiling. Curr. Opin. Chem. Biol. 8: 54-59. Full Text
N. Jessani, Humphrey, M., McDonald, W. H. et al. 2004. Carcinoma and stromal enzyme activity profiles associated with breast tumor growth in vivo. Proc. Natl. Acad. Sci. USA 101: 13756-13761. Full Text
A. Saghatelian, Jessani, N., Joseph, A. et al. 2004. Activity-based probes for the proteomic profiling of metalloproteases. Proc. Natl. Acad. Sci. USA 101: 10000-10005. Full Text
S. A. Sieber, Mondala, T. S., Head, S. R. et al. 2004. Microarray platform for profiling enzyme activities in complex proteomes. J. Am. Chem. Soc. 126: 15640-15641. Full Text
A. E. Speers, & Cravatt, B. F. 2004. Profiling enzyme activities in vivo using click chemistry methods. Chem Biol. 11: 535-546. Full Text
A. E. Speers, & Cravatt, B. F. 2004. Chemical strategies for activity-based proteomics. Chembiochem 5: 41-47. Full Text
Harnessing RNA Interference for Therapy
J. Lieberman & Dykxhoorn, D. 2005. The Silent Revolution: RNA interference as a basic biology research tool and therapeutic. Ann Rev Med 401-423. [PDF]
C.D. Novina, Murray, M.F., Dykxhoorn, D.M., et al. 2002. siRNA-directed inhibition of HIV-1 infection. Nature Med 8:681-686.
E. Song, Lee S.-K., Wang J., et al. 2003. RNA interference targeting fax protects mice from fulminant hepatitis. Nature Med 9:347-351.
E. Song, Lee S.-K., Dykxhoorn D.M., et al. 2003. Sustained Small Interfering RNA-Mediated Human Immunodeficiency Virus Type 1 Inhibition in Primary Macrophages. J Virol 77:7174-7181. Full Text
P. Hamar, Song E., Kokeny G., et al. 2004. Short interfering RNA targeting Fas protects mice against renal ischernia-reperfusion injury. Proc Natl Acad USA 101:14883-14888.
J.Lieberman. 2003. Shooting the Messenger: Harnessing RNA Interference to Combat HIV Infection. Summary by Tim Horn; edited by Veronica Miller. The PRN Notebook 8:11-14.
J. Lieberman, Song E., Lee S.-K. & Shankar P. 2003. Interfering with disease: Opportunities and roadblocks to harnessing RNA interference. Trends Mol Med 9:397-403.
E. Song, Stern P., Palliser D., et al. 2004. RNA Interference in Animal Models. In M Sohail, editor. Gene Silencing by RNA Interference: Technology and Application, London: CRC Press.
P. Shankar & Lieberman J. 2005. RNAi and HIV: from here to therapy. In S. Butera, ed. HIV Chemotherapy: A Critical Review. Horizon Scientific Press. In press.
D. Dykxhoorn & Lieberman J. 2005. The Silent Revolution: RNA interference as basic biology, research tool and therapeutic. Ann Rev Med 56:401-423.
P. Shankar, Manjunath N. & Lieberman J. 2005. The Prospect of Silencing Disease Using RNA Interference. JAMA 293:1367-1373.
Keynote Speaker
Stuart L. Schreiber, PhD
Harvard University
email | web
site | publications
Stuart L. Schreiber is an investigator at the Howard Hughes
Medical Institute and is Morris Loeb Professor and chair of the department of
chemistry and chemical biology at Harvard University. He is a founder and
director of the Harvard Institute of Chemistry and Cell Biology (ICCB) and of
its affiliated, National Cancer Institute-sponsored Initiative for Chemical
Genetics, and a member of the faculty of the Broad Institute, a joint initiative
of Harvard University and the Massachusetts Institute of Technology. He is also
a member The Rockefeller University Board of Trustees.
Schreiber earned a PhD in organic chemistry at Harvard University before
joining the faculty at Yale University in May of 1981. He returned to Harvard in
1988. He is known for having developed systematic ways to explore biology,
especially disease biology, using small molecules (precursors to therapeutic
drugs that are used as bioprobes) and for his role in the development of the
field of chemical biology. Using his chemical approach, he has discovered
principles that underlie information transfer and storage in cells.
He is a member of the National Academy of Sciences and the American Academy
of Arts & Sciences and is a founder of several successful biotechnology
firms, including Vertex Pharmaceuticals, ARIAD Pharmaceuticals, and Infinity
Pharmaceuticals. To facilitate sharing of information derived from small
molecules, Schreiber and the ICCB launched the online public database ChemBank
in 2003.
Speakers
Benjamin F. Cravatt, PhD
The Scripps Research Institute
email | web site | publications
Benjamin F. Cravatt is a professor at the Skaggs Institute for
Chemical Biology and the departments of cell biology and chemistry at The
Scripps Research Institute, where he directs a research group that develops and
applies an array of biochemical, chemical, and genetic technologies to elucidate
the roles of enzymes in physiological and pathological processes, especially as
pertains to the nervous system and cancer. Cravatt's research group has obtained
fundamental insights into the chemical, biochemical, and physiological workings
of several important mammalian serine hydrolases, including enzymes involved in
the neurobiology of pain and in proteases associated with tumor progression.
Cravatt received a PhD in macromolecular and cellular structure and chemistry
from The Scripps Research Institute in 1996. His honors include a graduate
fellowship from the National Science Foundation (1992-1995), a Searle Scholar
Award (1998-2001), a Technology Review's TR100 Top 100 Young Innovators Award
(2002), the Promega Award for Early Career Life Scientists from the American
Society for Cell Biology (2002), the Eli Lilly Award in Biological Chemistry
(2004), and a Cope Scholar Award (2005).
Akira Kawamura, PhD
Hunter College, CUNY
email | web
site | publications
Akira Kawamura received his BS and MS degrees in organic
chemistry under the guidance of Kazuo Tachibana at the University of Tokyo,
where he worked on the isolation and biochemical characterization of
shark-repelling substances from the Moses sole flat fish. Kawamura received his
PhD in chemistry from Columbia University under the direction of Koji Nakanishi
and Nina Berova in 1999 for his microscale structural analyses on sphingolipids,
brassinosteroids, and endogenous ouabain.
Before joining Hunter College as an assistant professor, Kawamura spent his
postdoctoral stint in the group of Peter G. Schultz at the Scripps Research
Institute, where he conducted genomic profiling of natural products and various
biological systems, including angiogenesis, cellular density arrest, and
hypertension from 1999 to 2002.
Judy Lieberman, MD, PhD
CBR Institute for Biomedical Research, Harvard Medical School
email | web site |
publications
Judy Lieberman, senior investigator at the Harvard Medical
School Center for Blood Research and professor in pediatrics, received her PhD
in theoretical physics at Rockefeller University and an MD in the joint
Harvard-Massachusetts Institute of Technology Program in Health, Science and
Technology. She trained in internal medicine and hematology/oncology at New
England Medical Center, and learned about T cell immunology in the laboratory of
Herman Eisen in the Center for Cancer Research at MIT.
Lieberman's laboratory studies cytotoxic T lymphocytes (CTLs), key cells in
the immune defense against viral infection and cancer, and their role in
antiviral immunity. A major focus is studying the molecular pathways used by
CTLs to induce cell death. Lieberman and her coworkers have identified a novel
caspase-independent apoptotic pathway induced by the CTL protease granzyme A.
Other work centers on understanding how CTL function is regulated.
Her laboratory has been characterizing the CTL response to HIV-1, the virus
that causes AIDS, and trying to understand why it does not ultimately control
the virus. She is also working to develop an oral HIV vaccine using Listeria
monocytogenes as a vector. Her interest in HIV led to recent work on
harnessing RNA interference (RNAi) to suppress HIV infection. Her laboratory is
working on translating RNAi for therapeutic use against HIV and other
indications. She was the first to demonstrate in an animal model that RNAi could
protect animals from disease.
Carmichael S. Roberts, PhD
Surface Logix, Inc.
email | web site
Carmichael C. Roberts is cofounder and president of Surface
Logix, Inc., a drug optimization company focused on combining proprietary
small-molecule chemistries with known biologically relevant pharmacophores to
create new chemical entities with improved pharmacodynamic and pharmacokinetic
properties. Previously, Roberts worked in business development at GelTex
Pharmaceuticals, which was acquired by Genzyme, where he designed business
development strategies for a preclinical product line for infectious disease and
obesity.
Roberts received a PhD in organic chemistry from Duke University and was a
National Science Foundation Fellow at Harvard University's department of
chemistry and chemical biology under Professor George Whitesides. He holds an
MBA from the Massachusetts Institute of Technology Sloan School of
Management.
Eloy Rodriguez, PhD
Cornell University
email | web
site | publications
Eloy Rodriguez describes himself as "a biological chemist
interested in the organic chemistry of natural drugs from plants, insects, and
fungi." He is the James A. Perkins Professor of Environmental Studies at Cornell
University where he specializes in the chemical properties of tropical and
desert plants. He developed and directs both the Punta Cana Cornell Biodiversity
Center in the Dominican Republic and the university's EsBaran Amazon Field
Station and Laboratory in Peru. Since 2000 he has also been associate director
of the Cornell's Weill Medical College Center for Complementary and Integrative
medicine, located in New York City.
Rodriguez, with Richard Wrangham of Harvard, is known for establishing the
field of and coining the term zoopharmacognosy, the study of animals' observed
ability to self-medicate and to protect themselves from disease. He earned a
doctorate in phytochemistry and plant biology from the University of Texas at
Austin in 1975. His past positions include professor in the School of Medicine
and departments of developmental cell biology and ecology and evolutionary
biology at the University of California at Irvine. He has published more than
155 articles and two books on a wide variety of topics in plant chemistry.
Jose A. Salas, PhD
University of Oviedo
email | web site | publications
Jose A. Salas is a professor of microbiology at the University
of Oviedo in Spain, where he obtained his PhD in 1980 by conducting studies of
cell differentiation in Streptomyces. His postdoctoral work in the UK at
the University of Cambridge and the University of Leicester looked at molecular
resistance mechanisms in antibiotic-producing actinomycetes.
Salas heads a group of 20 scientists focused on the isolation and
characterization of antibiotic and antitumor biosynthesis gene clusters. The
group, which aims to engineer these biosynthetic pathways to generate novel
derivatives with potential clinical application, has published more than 120
papers and filed 10 patents based on scientific discoveries.
Adrienne Burke
Adrienne Burke is founder of Genomic Society Consulting, which provides market research and communications services to the life sciences industry. She has been a science and health journalist since 1993.
She was the founding editor-in-chief of Genome Technology magazine and
editorial director of GenomeWeb.com. She also served as editor-in-chief of
BioInform, a weekly newsletter for the bioinformatics industry, and
oversaw launches of the newsletters ProteoMonitor and Agricultural
Genomics. Earlier, she was a managing editor of a magazine for occupational
health professionals.
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