Where Two Fields Meet

As Dale Boger, Professor, Department of Chemistry, The Scripps Research Institute, La Jolla, California, observes, the term “chemical biology” means a different thing to every person, and has had different meanings throughout the years. He describes chemical biology as the application of chemistry to biology and the biology/chemistry interface. “If I had to attribute a single feature as to why chemists are interested in the chemistry/biology interface, it's when structural information became available on proteins, so the problems of biology could be related to structural features.” Boger's group's interests revolve around making molecules to exploit, predict, and control their properties. “The range of our work is pretty wide,” he explains. It encompasses synthesizing natural products, analogs of natural products, bioorganic and medicinal chemistry, DNA-agent interactions, and the chemistry of anti-tumor and immunomodulatory agents. “Our combinatorial chemistry approach is dictated by the problem we want to solve.”

By synthesizing natural compounds in the lab, the group is able to make a range of derivatives and analogues of those compounds. If the mechanism of action is not known, they try to determine it. If it is known, they try to characterize the nature of the interaction of the molecule with its target.

From Basic to Applied Research

Boger notes that the projects undertaken by his group are not those that would be taken on by pharmaceutical companies. His team will make a compound to answer a “yes or no” question, where obtaining an unambiguous answer is important but the actual answer is not. Pharmaceutical companies, on the other hand, would not invest in the same project if there was a possibility the answer might be “no.” This is not to say that commercial applications don't arise from this work. For example, a large collaboration of groups at Scripps has developed potent inhibitors of a biologic target, fatty acid amide hydrolase (FAAH). This enzyme degrades oleamide, an endogenous sleep-inducing lipid, and anandamide, an endogenous ligand for cannabinoid receptors. The inhibitors serve as tools to clarify the role of endogenous oleamide and anandamide, and may prove to be useful for developing therapeutic agents to treat sleep disorders or pain. Boger thinks that pharmaceutical companies will target this new class of endogenous signaling molecules in an effort to replace the now problematic cyclooxygenase-2 (COX-2) inhibitors. “This work involved beautiful techniques, including proteomewide screening of inhibitors, and not just screening, but optimizing selectivity and using novel designs to inhibit,” he says.

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Boger points out that the focus on the choice of problems to address, and what chemistry to bring to bear on and build function into molecules, is clearly going to have an effect on public health. It will be up to pharmaceutical companies to do the experimental validation in animal models, but he believes that in this case, as well as others, those pharmaceutical companies would never have discovered the target. “About ten years into the fifteen-year-long process, people looked at our process as, if not flaky, then shaky. Then ten years in, they see it as interesting. Still, someone has to identify and validate these targets. There's a huge role for basic science, even bigger today than in the past.”

Inorganic Physiology

“My opinion is that chemical biology is a very broad field. It's synthetic chemistry applied to biology in new ways,” says Thomas V. O'Halloran, Director, Chemistry of Life Processes Institute, Northwestern University, Evanston, Illinois. “What I bring into the vision is inorganic in flavor.” He thinks that the field of bioinorganic chemistry emerged from the study of metabolic enzymes, from the old field of trace element nutrition. He takes an analytic chemistry and medical biology approach, combined into a new field he calls inorganic physiology, defined as how inorganic chemistry is done inside the cell. One problem cells have is how to get transition metals into the correct sites, what he calls the “metalome.” Transition metals were once thought to be trace components in cells. In bacterial cells, the total number of molecules of transition metals like iron or zinc is in millimolar quantities. Other important metals include copper, manganese, and sometimes nickel. All of these are concentrated by cells. “It takes a huge amount of energy to sequester these components, because the cells need to double their metals to divide. The competition for metals is like the commodities market.”

These metals are different from organic cofactors because the cell can't make or destroy them, so the control chemistry must be controlled locally. The cell must be able to distinguish among these metals. O'Halloran observes that it is a real challenge for chemists to design specific chelators, whereas evolution has done it better, with different metalloenzymes that can only use one metal, for example, zinc but not copper. His group has worked out some pathways involving metalloregulatory proteins, proteins that act as metal responsive genetic switches. They have also named and characterized the activity of metallochaperones—diffusible metal ion receptors involved in intracellular metal trafficking that pick up metals and deliver them to their target proteins.

“We focus on new classes of metal receptors that control the physiology of single cells, like bacterial cells, which need to transport and deliver millions of metal cofactors,” O'Halloran says. Mammalian cells, like bacterial cells, have needs for metals that are organ-specific, and these needs are for a particular metal across the periodic chart. For example, the pancreas, above and beyond the normal requirements of any mammalian cell, has an extra need for zinc, which is involved in insulin action.

“I am convinced there is a whole new era of therapy opening up using good medicinal chemistry,” O'Halloran says. “It's a fascinating time to apply emerging technology in chemical science to genetic findings. I am in the middle, understanding mechanisms and building a better picture of how cells do inorganic chemistry.”

New Routes to Discovery

Interests of Linda B. McGown, William Weightman Walker Professor and Head, Department of Chemistry and Chemical Biology, Rensselaer Polytechnic Institute, Troy, New York, and her group, include affinity techniques for protein analysis, analytical applications of aptamers, and the role of genomic G-quadruplex DNA in gene regulation in human diseases, such as diabetes and cancer. Aptamers, nucleic acid sequences that bind to protein targets, have several advantages over antibodies as affinity-binding reagents. They can be easily synthesized and modified chemically, are highly stable, and are reusable, because they can be reversibly folded and unfolded for capture and release of the target protein.

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Image 2. (Click to enlarge)

McGown's group is exploring a new route to discovery of DNA-binding ligands using a platform her group developed for affinity matrix–assisted laser desorption-ionization (MALDI) mass spectroscopy (MS) screening. In this rapid screening method, the affinity binder is immobilized at the probe surface, and biologic samples, for example, cell lysates, blood, or nuclear fractions, are added. If proteins from the samples attach to the affinity binder, the proteins can be concentrated, purified, and analyzed by MALDI-MS. McGown likens it to a reverse SELEX (systematic evolution of ligand by exponential enrichment process) approach in which, rather than screening a combinatorial DNA library for affinity to a target protein, the DNA is designed from genomic sequences and the protein samples are natural biological “combinatorial” libraries of proteins. The DNA-binding ligands differ from aptamers not only in the process of their discovery, but also that they may potentially reveal biologically significant protein-DNA interactions. G-quadruplex structures are thought to form in genomic DNA. McGown is looking at sequences in promoter regions that have been identified from their ability to form G-quadruplexes in vitro. These structures so far have not been seen in vivo, although many researchers hypothesize that they are formed in living human cells. One region of interest she is working on is the insulin-linked polymorphic region (ILPR), a non-coding region in the promoter of the insulin gene. It is found only in primates, is highly polymorphic in humans, and is in a locus highly correlated with susceptibility to type 1 diabetes. McGown's group has discovered that one ILPR variant G-quadruplex binds insulin and insulinlike growth factor 2 (IGF-2) to a similar affinity, but does not recognize IGF-1 to the same degree. “This may be of biologic significance,” she says. “It's a start. We can play around with increasing the sensitivity of binding for insulin over IGF-2, and look for in vivo interactions.” This work may ultimately lead to the discovery of both disease biomarkers and ligands for their detection.