Effects of substitutions at the 4' and 2 positions on the bioactivity of 4'-ethynyl-2-fluoro-2'-deoxyadenosine

Antimicrob Agents Chemother. 2013 Dec;57(12):6254-64. doi: 10.1128/AAC.01703-13. Epub 2013 Oct 7.

Abstract

Nucleos(t)ide reverse transcriptase inhibitors (NRTIs) form the backbone of most anti-HIV therapies. We have shown that 4'-ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) is a highly effective NRTI; however, the reasons for the potent antiviral activity of EFdA are not well understood. Here, we use a combination of structural, computational, and biochemical approaches to examine how substitutions in the sugar or adenine rings affect the incorporation of dA-based NRTIs like EFdA into DNA by HIV RT and their susceptibility to deamination by adenosine deaminase (ADA). Nuclear magnetic resonance (NMR) spectroscopy studies of 4'-substituted NRTIs show that ethynyl or cyano groups stabilize the sugar ring in the C-2'-exo/C-3'-endo (north) conformation. Steady-state kinetic analysis of the incorporation of 4'-substituted NRTIs by RT reveals a correlation between the north conformation of the NRTI sugar ring and efficiency of incorporation into the nascent DNA strand. Structural analysis and the kinetics of deamination by ADA demonstrate that 4'-ethynyl and cyano substitutions decrease the susceptibility of adenosine-based compounds to ADA through steric interactions at the active site. However, the major determinant for decreased susceptibility to ADA is the 2-halo substitution, which alters the pKa of N1 on the adenine base. These results provide insight into how NRTI structural attributes affect their antiviral activities through their interactions with the RT and ADA active sites.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Deoxyadenosines / chemistry*
  • Deoxyadenosines / pharmacology
  • HIV Reverse Transcriptase / antagonists & inhibitors
  • HIV Reverse Transcriptase / metabolism
  • HIV-1 / drug effects
  • Humans
  • Kinetics
  • Magnetic Resonance Spectroscopy
  • Molecular Conformation
  • Reverse Transcriptase Inhibitors / chemistry
  • Reverse Transcriptase Inhibitors / pharmacology
  • Structure-Activity Relationship

Substances

  • Deoxyadenosines
  • Reverse Transcriptase Inhibitors
  • HIV Reverse Transcriptase
  • 4'-ethynyl-2-fluoro-2'-deoxyadenosine