In vitro study of resistance-associated genotypic mutations to nucleoside analogs

Nucleosides Nucleotides Nucleic Acids. May-Aug 2003;22(5-8):991-4. doi: 10.1081/NCN-120022720.

Abstract

In spite of a rather long period of investigations, the problem of HIV drug resistance remains unsolved, and more that, at present HIV-1 mutants resistant to all known nucleoside inhibitors being used in clinical therapy against the human immunodeficiency syndrome are discovered. In this study we selected HIV-1 mutants resistant to the nucleoside inhibitors of HIV reverse transcriptase (NRTI): 3'-azido-2',3'-dideoxythymidine (AZT), 5'-phosphit 3'-azido-2',3'-dideoxythymidine (ph-AZT), dideoxyinosine (ddI) and didehydrodeoxythymidine (d4T). Selection of resistant mutants was carried out by gradually increasing of drug concentration in the culture medium during propagation of the HIV-1EVK on fresh MT-4 cells. Phenotypic resistance was defined as an increase in ID50 of 160-fold for AZT, 8 for ph-AZT, 10 for ddI, 7 for d4T. In comparison studies it was determined that the viral resistance to these drugs was appeared variously in a similar conditions and duration of selection. The nucleotide sequences of the RT region of the HIV-1 variants were compared with the HIV-1EVK from "0" passage. For some of selected HIV-1 mutants NRTI resistance mutations were detected. Selected AZT resistant variants contained amino acid substitutions in positions D67A and K70R. Our studies was not revealed substitution at position 75 for ph-AZT resistant variants, whereas substitution at position L214F have been observed in both experiments using AZT and ph-AZT. Selected d4T resistant mutants contained amino acid substitutions in positions N54D and P52R. Selected ddI resistant mutants contained only one amino acid substitution in position P143S. Collection of drug-resistant mutants should prove to be a convenient tool for rapid investigations a new antiretroviral agents on cross drug-resistance.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Didanosine / chemistry
  • Didanosine / pharmacology
  • Drug Resistance, Viral / genetics*
  • Genetic Variation
  • Genotype
  • HIV Reverse Transcriptase / metabolism
  • HIV-1 / drug effects*
  • HIV-1 / enzymology
  • HIV-1 / genetics*
  • Humans
  • Mutation*
  • Nucleosides / chemistry
  • Nucleosides / pharmacology*
  • Reverse Transcriptase Inhibitors / chemistry
  • Reverse Transcriptase Inhibitors / pharmacology
  • Stavudine / chemistry
  • Stavudine / pharmacology
  • Zidovudine / chemistry
  • Zidovudine / pharmacology*

Substances

  • Nucleosides
  • Reverse Transcriptase Inhibitors
  • Zidovudine
  • Stavudine
  • HIV Reverse Transcriptase
  • Didanosine