Unusual codon 69 insertions: influence on human immunodeficiency virus type 1 reverse transcriptase drug susceptibility.
BACKGROUND
Multiple amino acid changes in the reverse transcriptase (RT) enzyme of the human immunodeficiency virus 1 (HIV-1) confer simultaneous resistance to most nucleoside RT inhibitors (NRTI). It may take place through different pathways: one of these is the codon 69 insertion, which can involve several 2-amino acid patterns.


METHODS
We are reporting the case of three patients treated with various antiretroviral compounds. For these subjects we have conducted both a genotypical and a phenotypical analysis in order to understand what kind of influence these insertions may have on HIV-1 RT drug susceptibility. Plasma samples from these patients have been extracted and the RT region has been amplified, cloned and sequenced; meanwhile their PBMCs have been separated, cultivated and then tested for drug susceptibility.


RESULTS
Data obtained from the cloning assay showed that the patients had different mutational patterns but constant multiple resistance to NRTI. In particular, they harbored mutations related to Zidovudine (ZDV), 3TC and various NRTIs. Moreover, all three samples had a T69S substitution followed by three different dual amino acid insertions: SG, TG and VG. Several phenotypic experiments revealed that the viruses were resistant to 3TC as well as to ZDV and ABC. Different results were obtained using d4T and ddI.


CONCLUSIONS
In our three patients, all mutation inserts impaired the use of NRTI, particularly ZDV and 3TC. Patient 001 presented a pattern that should not cause a high phenotypic resistance to 3TC per se, and so we can argue that the concomitant presence of the insertion T69S (SG) makes this isolate moderately resistant to this drug. We observed a similar phenomenon in subject 003. d4T was less involved in the resistance generation caused by the RT insertion (in one out of three cases). Moreover, we identified a new 2aa insertion (TG) that has, to the best of our knowledge, never been reported before. A careful survey of novel RT genotypic insertion is thus warranted.
Bulgheroni
Elisabetta
E
Istituto di Malattie Infettive e Tropicali, Università degli Studi, Ospedale Luigi Sacco, Via GB Grassi 74, 20157 Milan, Italy.
Croce
Francesco
F
Citterio
Paola
P
Viganò
Ottavia
O
Visonà
Raffaella
R
Sala
Eugenio
E
Galli
Massimo
M
Rusconi
Stefano
S
D019943:Amino Acid Substitution; D019380:Anti-HIV Agents / Q000494:pharmacology* / Q000627:therapeutic use; D003062:Codon; D024921:Drug Resistance, Multiple, Viral; D004359:Drug Therapy, Combination; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology; D054303:HIV Reverse Transcriptase / Q000187:drug effects* / Q000235:genetics; D015497:HIV-1 / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology* / Q000627:therapeutic use; D017422:Sequence Analysis, DNA
D019380:Anti-HIV Agents; D003062:Codon; D018894:Reverse Transcriptase Inhibitors; D054303:HIV Reverse Transcriptase