Nucleosides for the treatment of respiratory RNA virus infections

Jordan, Paul C; Stevens, Sarah K; Deval, Jerome

What are the molecular determinants of polymerase selectivity against nucleotide analogs? We currently do not understand well how specific changes made in nucleotide analogs alter their recognition as substrates for RNA synthesis, and how substrate selectivity differs among positive and negative strand RNA virus polymerases. For example, many 2 0 -modified nucleotide analogs are known to inhibit HCV polymerase, often with an antiviral spectrum extended to flaviviruses and picornaviruses. However, there is no clear mechanistic basis to explain why none of these compounds inhibit (-)ssRNA viruses, or even other (þ)ssRNA viruses such as CoV. Could the exonuclease/proofreading activity of CoV polymerases excise chain terminators and resume RNA synthesis? Are there specific amino acid within the active site of (-)ssRNA virus polymerases responsible for the discrimination of 2 0 -C-methyl nucleotides? These hypotheses have not been tested, in part, due to the difficulty to conduct biochemical and structural studies on viral polymerases from respiratory viruses. Until recently, the production of soluble, pure viral protein targets has been limiting, especially in the case of large protein complexes. As mentioned earlier in this review, the development of robust expression systems for influenza polymerase trimer, as an example, have made it possible to use X-ray crystallography and potentially cryo-electron microscopy to provide molecular visualization of binding pockets for small molecule inhibitors, entry and exit channels for substrate(s), and potential new ways to disrupt domain interactions. These structural insights will tremendously aid the development of new drugs as well as to further elucidate the mechanisms of action and binding of existing drugs to their protein targets. Molecular modeling is also a useful approach that we and others have used to rationalize the differences in selectivity of lumicitabine against RSV and HCV polymerase. 159 More studies such as these ones will be needed to rationally design new nucleotide analogs targeting respiratory virus polymerases.