Progress and successes

• The structures of picornaviruses, like rhinovirus and poliovirus, form a basis for the development of tightly-binding ligands which prevent infection - presumably by blocking the uncoating process of the virus. For the rhinoviruses, responsible for the common cold, the great variability of the virus poses a tremendous challenge for the design of a broad-spectrum drug. This variability is seen not only in the antigenic regions but also in the ligand-binding pocket. As well as the potential for designing a drug that inhibits viral replication, structure-based drug design offers the possibility of designing ligands that stabilize the virus coat, with potential application for poliovirus vaccines. Full implementation of the polio vaccination program in third world countries is hampered by the thermolability of the virus used in the vaccine, which requires the vaccine to be kept at low temperatures, without a single interruption, from manufacturer to injection in the patient. A ligand which would stabilize the virus might be of tremendous value in allowing the vaccine to be exposed to higher temperatures and thus to be more easily used in tropical areas.
  • Kim K.H. et al , Pevear D.C. : 1993, A comparison of the anti-rhinoviral drug binding pocket in HRV14 and HRV1A. J. Mol. Biol. 230: 206-227.
  • Caliguiri L.A. , McSharry J.J. , Lawrence G.W. : 1980, Effect of arildone on modifications of poliovirus in vitro.Virology 105: 86-93.

• Dihydrofolate reductase (DHFR) was the first target protein solved in complex with a drug - even though the complex was a cancer drug, methotrexate, bound to a bacterial enzyme, it was an important first step. Three-dimensional structures of dihydrofolate reductase have been the basis for the design of several improved inhibitors. Both DHFR and thymidylate synthase (TS), another enzyme involved in folate metabolism, are excellent targets for drug design since they are crucial for pyrimidine and purine synthesis. In the search for new cancer chemotherapeutics a TS inhibitor with K_i = 960 nM was rationally changed into one with K_i = 15 nM; the compound is now in clinical evaluation. Recently, the three-dimensional structure of a bifunctional DHFR°V thymidylate synthase from the protozoon parasite Leishmania major has been unravelled, providing an opportunity for a two-pronged attack for drug design for the treatment of leishmaniasis.
  • Kuyper L.F. et al , Goodford P.J. : 1985, Receptor-bound design of dihydrofolate reductase inhibitors: comparison of crystallographically determined enzyme binding with enzyme affinity in a series of carboxy-substituted trimethoprim analogues. J. Med. Chem. 28: 303-311.
  • Webber S.E. et al , Bartlett C.A. : 1993, Design of thymidylate synthase inhibitors using protein crystal structures: the synthesis and biological evaluation of a novel class of 5-substituted quinazolinones. J. Med. Chem. 36: 733-746.
  • Knighton D.R. et al , Matthews D.A. : 1994, Structure and kinetic channelling in bifunctional dihydrofolate reductase thymidylatesynthase. Nature Struct. Biol. 1: 186-194.

   

• A stunning example of successful improvement of a lead compound has been published in the case of influenza virus neuraminidase. The investigators used GRID to find a favorable position for adding a positively charged substituent to the sialidase inhibitor 2-deoxy-2,3-didehydro-D-N-acetylneuraminic acid (Neu5Ac2en). This modified saccharide not only binds over three orders of magnitude better than the parent compound, but is also effective in vitro and in vivo against virus infection.
  • Rational design of potent sialidase-based inhibitors of influenza virus replication. Nature 363: 418-423.

   

• Structural studies on elastase have revealed the erratic mode of binding of closely related inhibitors. Three modifications of the same parent ligand molecule led to surprisingly different binding modes of the three daughter molecules.
  • Mattos C., Rasmussen B., Ding X., Petsko G.A., Ringe D. : 1994, Analogous inhibitors of elastase do not always bind analogously. Nature Struct. Biol. 1: 55-58.

• Thrombin provides an example where natural compounds have been used as leads in a structure-based drug optimization process. The three-dimensional structure of the leech inhibitor hirudin has been the basis for the development of compounds such as 'hirulog', a potent inhibitor of thrombin. Fragments of the natural target of thrombin, fibrinogen, have also been incorporated into high-affinity ligands. Surprisingly, oligonucleotides can also be found that inhibit the activity of thrombin. The binding mode has been unravelled in a three-dimensional structure and is likely to allow the development of yet another family of thrombin inhibitors.
  • Nakanishi H. et al , Kahn M. : 1992, Peptide mimetics of the thrombin-bound structure of fibrinopeptide A. Proc. Natl. Acad. Sci. USA. 89: 1705-1709.
  • Padmanabhan K. , Padmanabhan K.P. , Ferrara J.D. , Sadler J.E. , Tulinsky A. : 1993, The structure of alpha-thrombin inhibited by a 15-mer single-stranded DNA aptamer. J. Biol. Chem. 268: 17651-17654.
  • Lam P.Y.S. et al , Erickson-Viitanen S. : 1994, Rational design of potent, bioavailable, nonpeptide cyclic ureas as HIV protease inhibitors. Science 263: 380-384.

   

• Knowledge of the three-dimensional structure of HIV protease, a small but crucial protein from the AIDS virus, has led to structure-based design of nanomolar, bioavailable, non-peptide inhibitors. Few proteins have been crystallographically studied more frequently than this protease - there are now very many known inhibitor-protease complexes. Interestingly, this goal-oriented research also provides an extremely precious database of known structures of protein-ligand complexes with associated binding constants. This is a gold-mine for the fundamental research of theoretical biophysicists, and will allow them to sharpen their theoretical and computational tools. Eventually this will of course be beneficial for improving the scoring tools in the ligand design process.
  • Lam P.Y.S. et al , Erickson-Viitanen S. : 1994, Rational design of potent, bioavailable, nonpeptide cyclic ureas as HIV protease inhibitors. Science 263: 380°V384.