s e i e η c e / t e c h η ο I o g y fàv.

COMBINATORIAL CARBOHYDRATES Prospects are looking sweet for use of sugar-based combinatorial libraries to find novel bioactive substances

Stu Borman C&EN Washington

C arbohydrates play a vital role in mo­lecular recognition, cell signaling, biomolecular transport, the immune

system, and, in fact, in virtually every es­sential biological process. So it's ironic that when it comes to combinatorial chemis­try—the synthesis of collections of varied molecules and the subsequent identifica­tion of those with useful properties—sug­ar-based molecules are about the last thing researchers think about.

This is largely because sugar-based compounds are darned ornery from a synthetic standpoint. Nevertheless, sev­eral research groups are beginning to de­velop increasingly useful strategies for producing combinatorial libraries (collec­tions of diverse compounds) based on these difficult and contrary molecules.

An example of this trend is an idea of using molecular ^ — — "scaffolds" for the display of organic functional groups. The idea, conceived in the early 1980s and reintroduced later in that decade, now is being translated into a novel combi­natorial strategy for carbohy­drate-based drug discovery.

The concept of varying mo­lecular structure and proper­ties systematically by decorat­ing molecular scaffolds with a variety of functional groups stems from research efforts in the 1970s and '80s to design peptidomimetics, compounds that mimic structural features of peptides but often have better potential as drugs. Pep­tides themselves aren't always the best drug candidates. They tend to get broken down by proteases in the gut and bloodstream and frequently have trouble entering the cells

in which they're needed—problems re­lated in part to their amide backbone structures.

A strategy to design peptidomimetics by totally discarding their problematical amide backbones and replacing them with novel structures—while still retaining the amino acid side chains required for recep­tor binding—was first proposed in 1980 by Patrick S. Farmer of the College of Phar­macy at Dalhousie University in Halifax, Nova Scotia. He suggested cyclohexane as the scaffold but was unsuccessful at dem­onstrating the concept experimentally.

In 1986, Patrice C. Bélanger and Claude Dufresne of the medicinal chem­istry department at Merck Frosst Canada, Pointe-Claire/Dorval, Quebec, first suc­cessfully employed this strategy. Bé­langer and Dufresne replaced the pep­tide backbone by a nonpeptide frame­work—a bicyclo[2.2.2]octane scaffold—

Subtle structural changes affect receptor binding

0(CH2)5NH2

OCHo

'-0(CH2)5NH2 RO' R =benzyl Source: Hirschmann and coworkers [J. Med. Chem., 41,1382 (1998)]

Sugar-based peptidomimetic at lower right binds somato­statin receptors by a different binding mode than related peptidomimetics (such as the other three shown). Moreover, acetylating the primary amino group of the compound at up­per left greatly enhances its affinity at the substance Ρ recep­tor but eliminates binding at somatostatin receptors--show-ing that the bioacthdty of these peptidomimetics is sensitive to relatively minor structural modifications.

and decorated it with amino acid side chains.

In the late 1980s, the groups of me­dicinal chemist Gary L. Olson, then at Hoffmann-La Roche, Nutley, N.J., and chemistry professor Ralph F. Hirschmann of the University of Pennsylvania indepen­dently explored the concept. Olson em­ployed a cyclohexane scaffold, whereas Hirschmann, working in collaboration with K. C. Nicolaou, then a chemistry pro­fessor at Penn, used β-D-glucose. After 1989, when Nicolaou moved to Scripps Research Institute in La Jolla, Calif., Penn chemistry professor Amos B. Smith III ex­tended the collaboration with Hirsch­mann to studies on diverse monosaccha­ride scaffolds.

"Hirschmann's putting substituents on the oxygens of carbohydrate scaffolds is one of the best ideas in carbohydrate chemistry in recent times," says chemistry professor Daniel E. Kahne of Princeton University, who specializes in research on bioactive oligosaccharides and glycoconju-gates. "It's really clever. Hirschmann and coworkers said sugars ought to be able to present side chains as well as peptides, and they showed that it worked. It was just fantastic—really creative."

Later on, Hirschmann, Nicolaou, and Smith at Penn; biologists Catherine D. Strader and Margaret A. Cascieri at Merck Research Laboratories (Rahway, N.J.); biol­ogy professor Wylie W. Vale at Salk Insti­tute (La Jolla, Calif.); biochemist Laurie T.

Maechler at MDS Panlabs ——••— (Bothell, Wash.); and cowork­

ers showed that derivatized monosaccharides were sur­prisingly good at binding a va­riety of biological targets [/. Am. Chem. Soc, 114, 9217 (1992)].

The researchers synthe­sized a series of glucose-based peptidomimetics that they be­lieved would bind to recep­tors for the peptide hormone somatostatin, and some of the compounds did so. But two of the peptidomimetics also bound to other targets, in­cluding the receptor for sub­stance P, a peptide involved in pain transmission—an unex­pected and surprising finding because somatostatin doesn't bind the substance Ρ receptor and substance Ρ doesn't bind somatostatin receptors. In ad­dition, these two peptidomi­metics bound to the β2-adren-

JULY 20, 1998 C&EN 49

science/technology

Smith: diverse monosaccharide scaffolds

Hirschmann: sugars are privileged platforms

Sofia: pharmacophore mapping libraries

ergic receptor, whose endogenous ligand (the catecholamine adrenaline) is not even a peptide.

Based on these findings, Hirschmann, Smith, and coworkers speculated that so­matostatin and substance Ρ receptors had some heretofore unrecognized features in common. They later demonstrated this ex­perimentally by readily converting a cyclic hexapeptide that acts as a highly potent and selective somatostatin receptor ligand

into a similarly potent and selective sub­stance Ρ receptor ligand \J. Med. Chem., 39, 2441 (1996)]. Since then, they have also shown that subtle changes in a sugar scaffold—such as going from D-glucose to L-glucose or L-mannose—can lead to a sub­stantial change in the biological profile of a sugar-based peptidomimetic.

"This work represented the first clue that sugars represent 'privileged plat­forms'—meaning that they can display

affinity to diverse receptors," says Hirschmann. "That is, there is some­thing about the sugar scaffold that allows carbohydrate peptidomimet-ics to interact with all kinds of differ­ent proteins."

And it turns out that when you build combinatorial libraries to screen for biologically active "leads" (drug candidates), that's exactly what you want. Earlier this year Hirschmann, Smith, senior research fellow Susan P. Rohrer of Merck Research Labo­ratories, and coworkers demonstrat­ed the use of carbohydrate scaf­folds to develop a potent receptor ligand for human somatostatin re­

ceptor subtype 4 and showed that selec­tivity for specific receptors can be ob­tained by modifying the substituents around a sugar ring [/. Med. Chem., 41, 1382 (1998)].

One of the most noteworthy results of this research was the discovery that one particular sugar-based peptidomimetic bound somatostatin receptors through a different mode than other sugar-based peptidomimetics. This was unexpected

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because compounds of the same structur­al class most frequently bind a target in the same way. "But in fact you can't assume that this is the case, and that's the point of this whole business," says Hirschmann. He notes that the alternate binding mode phe­nomenon had been previously demon­strated for protease inhibitors by the groups of professor of chemistry and phar­macy Daniel H. Rich of the University of Wisconsin, Madison, and chemistry profes­sor Dagmar Ringe of Brandeis University, Waltham, Mass.

Researchers at Intercardia Research Laboratories (formerly Transcell Tech­nologies), Cranbury, N.J., have been im­plementing and expanding on the con­cepts devised by Hirschmann and co­workers. Whereas Hirschmann's group showed how a sugar with added amino acid side chains can potentially bind re­ceptors that the sugar wouldn't normal­ly recognize, Intercardia scientists are us­ing solid-support chemistry to actually make large combinatorial libraries of such compounds.

The Intercardia researchers produce an enormous amount of molecular diversity by functionalizing sugar-based ring sys­tems with different kinds of protecting groups. "We were looking for a strategy that would allow us to use carbohydrates for generating libraries that could be screened against a wide variety of biomo-lecular targets, not just those targets that naturally recognize sugars, and we felt that this primary screening library would need to exhibit small-molecule druglike charac­teristics," says Michael J. Sofia, Intercar-dia's vice president of research and direc­tor of chemistry. Sofia and coworkers re­cently generated such a library based on monosaccharide scaffolds that have three sites where chemical diversity could be in­troduced rapidly \J. Org. Cbem., 63, 2802 (1998)].

"We call these libraries universal phar­macophore mapping libraries," says Sofia, "because an analysis showed that they can access chemical diversity space that is not only wide-ranging but also unique when compared to, let's say, a tripeptide system or a small-molecule database of [noncar-bohydrate] druglike molecules." Intercar-dia is using the libraries to identify anti-infective agents in an internal screening program and also intends to use them in collaborative screening efforts with other companies.

Other groups working on carbohy­drate-based combinatorial strategies in­clude those of Kahne, chemistry professor Ole Hindsgaul at the University of Alberta,

Edmonton, and chemistry professor Chi-Huey Wong of Scripps.

Around 1995, Kahne and coworkers constructed the first solid-phase carbohy­drate library, using oligosaccharide solid-phase synthesis chemistry developed earli­er by his group, and Hindsgaul and coworkers developed a "random glycosyl-ation" strategy for making oligosaccharide libraries in solution. More recently, Hinds-gaul's group has been busy synthesizing " carbohybrids "—carbohydrates derivat-ized with organic functional groups—in

an effort to identify novel ligands of carbo­hydrate-binding proteins.

"Oligosaccharides that bind proteins normally bind very weakly," says Hinds­gaul, "so we don't try to mimic oligosac­charides. We decided to just admit that sugars don't have the right stuff for tight binding and to add some nonsugar groups, which is why we call the compounds car­bohybrids." Hindsgaul and coworkers Ulf J. Nilsson and Eric J-L. Fournier recently identified a carbohybrid that inhibits the enzyme β-galactosidase with the highest

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JULY 20, 1998 C&EN 51

s c i e n c e / t e c h n o l o g y

Carbohybrids target carbohydrate-binding proteins

OFt^OR

R 0 -Ο II

OR

Ο II

SCCH3

R = C(CH2)10CH3

Hindsgaul and coworkers modify carbohydrates to generate carbohy­brids, such as structure at bottom—a monosaccharide attached to a small organic ring derivatized with an amino acid.

activity ever achieved for inhibitors of that enzyme, work that will be reported in an upcoming issue of Bioorganic & Medici­nal Chemistry.

Wong and coworkers recently report­ed use of a new protection and deprotec-tion strategy for synthesizing carbohy­drate-based libraries \J. Am. Chem. Soc, 120, 7137 (1998)]. "We have four differ­ent protecting groups for a sugar," says

Hindsgaul: synthesizing carbohybrids

Wong, "and you can selectively deprotect any of these positions for the glycosylation reaction" used to couple carbohydrate units. Wong's group demonstrated the technique by using it to synthesize a 45-member oligosaccharide library as individ­ual molecular entities. The researchers are currently screening the library for com­pounds that bind to lectins and antibodies.

"The application of combinatorial and

e d u c a t i o n

carbohydrate chemistry to the develop­ment of novel pharmaceuticals is still in the early stages," notes Hindsgaul. "The fact that several very different approaches are being used by the various groups in­creases the chances that important new discoveries will be made. In the jargon of combinatorial chemistry, not only 'molec­ular diversity' but also 'research diversity' is at play"^

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Lehigh offers polymer education at a distance At the start of this year, six employees of industrial firms far from Lehigh Universi­ty in Bethlehem, Pa., began studies for master's degrees in polymer science and engineering from the university. These first distance-education M.S. students in Lehigh's Center for Polymer Science & Engineering have been attending classes televised to their workplaces. They will even do research for their theses without ever going to Bethlehem.

The six students took two three-credit courses: a chem­istry course called Organic Polymer Science and a chemi­cal engineering course called Polymer Interfaces. The class sessions were televised from the campus and the encrypt­ed signal decoded from Le­high at each of the work sites.

Students had voice tele­phone and fax links to the Le­high classroom and could also communicate by computer with Lehigh's interactive com­puter. Videotape backups were available when students missed classes because they were traveling. In addition to participating in live classes, students could interact with each other and with students on campus by logging on to chat rooms associated with each course.

The students will eventually amass 24 credits in courses from the departments of chemistry, chemical engineering, me­chanical engineering, materials, or phys­ics, plus six credits of research. Students will do research on nonproprietary top­ics at their workplaces, codirected by their employers and Lehigh professors. The full-time equivalent of four to six months of work may be needed to com­plete their theses.

"While one-third to one-half of all de­greed chemical engineers, chemists, and

materials scientists and engineers are en­gaged in some form of polymer science and engineering at any one time, only a fraction of these people have had as much as one regular polymer course," says Leslie H. Sperling, director of Lehigh's Engineer­ing Polymers Laboratory. "While the situa­tion is slowly improving, polymer educa­tion clearly still lags behind as an academ­ic subject. The objective of this distance

Sperling lectures to Polymer Interfaces students in one of two distance learning classrooms at Lehigh.

education program is to make available to professionals around America a chance to address this problem."

Companies interested in distance edu­cation partnerships with Lehigh are ex­pected to designate support personnel, set aside two classrooms, and acquire the needed television equipment, computers, and phone and fax lines. Lehigh charges no partnership fees.

Persons interested in applying as stu­dents or who have questions about involv­ing their companies should contact Peg Kercsmar, manager of the Office of Dis­tance Education, Lehigh University, 36 Uni­versity Dr., Bethlehem, Pa. 18015; phone: (610) 758-5794, e-mail: mak5@lehigh.edu.

Stephen Stinson

52 JULY 20, 1998 C&EN