Coexistence of the K65R/L74V and/or K65R/T215Y mutations on the same HIV-1 genome

Henry, Mireille; Tourres, Christian; Colson, Philippe; Ravaux, Isabelle; Poizot‐Martin, Isabelle; Tamalet, Catherine

doi:10.1016/j.jcv.2006.07.001

Cited by 14 publications

(13 citation statements)

References 12 publications

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“…11 Both K65R and L74V are known to be associated with diminished viral replication capacity, and only on rare occasions have the two mutations been shown to be located on the same viral genome. 12,13 The level of detection of HIV-1 drug resistance mutations by direct polymerase chain reaction (PCR) population sequencing currently utilized for patient management is about 20%. This level of detection may not always provide adequate information for the optimal treatment decisions, as lowfrequency subpopulations (<20%) of drug-resistant variants may have clinical relevance.…”

Section: Introductionmentioning

confidence: 99%

“…Another comparison between commercially available population sequencing results and a more sensitive research analysis method examined whether K65R might occur as a low-frequency subpopulation in plasma from subjects failing a first-line ABC/ 3TC-containing HAART regimen and population sequence indicating high-abundance L74V at the time of virologic failure; this study utilized a research RT K65R-selective PCR method to detect low-frequency quasispecies at that single position. [11][12][13][14] Although a relatively small number of subjects (four) was evaluated, K65R was detected in 1/720 clones for one of the four subjects studied at virologic failure. These data suggest that low-frequency quasispecies harboring K65R may occur relatively uncommonly (and at very low population densities) in patients experiencing virologic failure on first-line HAART regimens.…”

Section: Introductionmentioning

confidence: 99%

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Tenofovir (TDF)-Selected or Abacavir (ABC)-Selected Low-Frequency HIV Type 1 Subpopulations During Failure with Persistent Viremia as Detected by Ultradeep Pyrosequencing

D’Aquila

Geretti

Horton

et al. 2011

AIDS Research and Human Retroviruses

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Detection of drug resistance is critical for determining antiretroviral treatment options. Ultradeep pyrosequencing (UDPS; 454 Life Sciences) is capable of detecting virus variant subpopulations with much greater sensitivity than population sequencing, which typically has a detection limit around 20%. UDPS of the HIV-1 reverse transcriptase (RT) (amino acids 56-120) was performed to detect the key mutations K65R and L74V associated with tenofovir and abacavir use. Plasma specimens from subjects with persistent rebound viremia following suppression on tenofovir (n = 8) or abacavir (n = 9)-based therapy were studied. Samples from a subject treated with zidovudine/lamivudine/efavirenz with a similar loss of virologic response served as a control. HIV-1 plasma RNA was ≥3.68 log(10) copies/ml at all time points sequenced. The median number of UDPS sequences analyzed/time point was 33,246. Among the eight tenofovir-treated subjects, three showed high-frequency (>20%) RT K65R at the time of failure, whereas one showed low-frequency (<20%) L74V; no low-frequency K65R was detected in these subjects. Among the nine abacavir-treated subjects, three showed low-frequency K65R; no L74V was detected in these patients. No K65R or L74V was detected in the samples from the control subject. At failure, other RT mutations were detected, including low-frequency NNRTI-resistant species detected at ≥1 time point in nine subjects; the key NNRTI mutation K103N, however, was always observed at >20% frequency. Although UDPS is useful in the detection of low-frequency subpopulations with transmitted resistance in antiviral-naive patients, it may have less utility in treatment-experienced patients with persistent viremia on therapy.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Tenofovir (TDF)-Selected or Abacavir (ABC)-Selected Low-Frequency HIV Type 1 Subpopulations During Failure with Persistent Viremia as Detected by Ultradeep Pyrosequencing

D’Aquila

Geretti

Horton

et al. 2011

AIDS Research and Human Retroviruses

View full text Add to dashboard Cite

show abstract

“…The resistance mutations L74V and K65R both participate in crucial hydrogen bond networks at the RT polymerase active site which stabilize the incoming deoxynucleoside triphosphate (dNTP) (10,14,41); however, these mutations rarely occur together in the clinical setting (13,44). The Q151M mutation changes the hydrogen bond network that involves the 3Ј-OH group of dNTP, thus reducing the incorporation of NRTIs, which lack a 3Ј-OH group (14,35).…”

mentioning

confidence: 99%

4′- C -Methyl-2′-Deoxyadenosine and 4′- C -Ethyl-2′-Deoxyadenosine Inhibit HIV-1 Replication

Boyer

Siddiqui³

et al. 2011

Antimicrob Agents Chemother

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It is important to develop new anti-HIV drugs that are effective against the existing drug-resistant mutants. Because the excision mechanism is an important pathway for resistance to nucleoside analogs, we are preparing analogs that retain a 3-OH and can be extended after they are incorporated by the viral reverse transcriptase. We show that 4-C-alkyl-deoxyadenosine (4-C-alkyl-dA) compounds can be phosphorylated in cultured cells and can inhibit the replication of HIV-1 vectors: 4-C-methyl-and 4-C-ethyl-dA show both efficacy and selectivity against HIV-1. The compounds are also effective against viruses that replicate using reverse transcriptases (RTs) that carry nucleoside reverse transcriptase inhibitor resistance mutations, with the exception of the M184V mutant. Analysis of viral DNA synthesis in infected cells showed that viral DNA synthesis is blocked by the incorporation of either 4-C-methyl-or 4-C-ethyl-2-deoxyadenosine. In vitro experiments with purified HIV-1 RT showed that 4-C-methyl-2-dATP can compete with dATP and that incorporation of the analog causes pausing in DNA synthesis. The 4-C-ethyl compound also competes with dATP and shows a differential ability to block DNA synthesis on RNA and DNA templates. Experiments that measure the ability of the compounds to block DNA synthesis in infected cells suggest that this differential block to DNA synthesis also occurs in infected cells.

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“…Previous studies analyzing the risks and incidence of K65R and L74V mutations in the largest single clinic cohort in Europe (The Chelsea and Westminster HIV cohort) have demonstrated that the risk of developing L74V or K65R mutation during HAART was 4.5 and 2.8 cases per 100 person/year, respectively [26]. The decreased frequency of selection of K65R and L74V and the rare occurrence of K65R+L74V on the same HIV genome [6,27] may be related to the observed attenuation of the virus in the presence of these mutations [5,7,12,15]. …”

Section: Resultsmentioning

confidence: 99%

“…These analyses provided the potential mechanism for the extreme rarity of the double mutant in HIV-infected patients. Similar to K65R+L74V, K65R+L74I is also rarely observed in the absence of other mutations [6-8]. Structurally, valine has two methyl groups, whereas isoleucine's branches are one methyl and one ethyl group.…”

Section: Introductionmentioning

confidence: 99%

A Leu to Ile but not Leu to Val change at HIV-1 reverse transcriptase codon 74 in the background of K65R mutation leads to an increased processivity of K65R+L74I enzyme and a replication competent virus

Chunduri

Rimland

Nurpeisov

et al. 2011

Virol J

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BackgroundThe major hurdle in the treatment of Human Immunodeficiency virus type 1 (HIV-1) includes the development of drug resistance-associated mutations in the target regions of the virus. Since reverse transcriptase (RT) is essential for HIV-1 replication, several nucleoside analogues have been developed to target RT of the virus. Clinical studies have shown that mutations at RT codon 65 and 74 which are located in β3-β4 linkage group of finger sub-domain of RT are selected during treatment with several RT inhibitors, including didanosine, deoxycytidine, abacavir and tenofovir. Interestingly, the co-selection of K65R and L74V is rare in clinical settings. We have previously shown that K65R and L74V are incompatible and a R→K reversion occurs at codon 65 during replication of the virus. Analysis of the HIV resistance database has revealed that similar to K65R+L74V, the double mutant K65R+L74I is also rare. We sought to compare the impact of L→V versus L→I change at codon 74 in the background of K65R mutation, on the replication of doubly mutant viruses.MethodsProviral clones containing K65R, L74V, L74I, K65R+L74V and K65R+L74I RT mutations were created in pNL4-3 backbone and viruses were produced in 293T cells. Replication efficiencies of all the viruses were compared in peripheral blood mononuclear (PBM) cells in the absence of selection pressure. Replication capacity (RC) of mutant viruses in relation to wild type was calculated on the basis of antigen p24 production and RT activity, and paired analysis by student t-test was performed among RCs of doubly mutant viruses. Reversion at RT codons 65 and 74 was monitored during replication in PBM cells. In vitro processivity of mutant RTs was measured to analyze the impact of amino acid changes at RT codon 74.ResultsReplication kinetics plot showed that all of the mutant viruses were attenuated as compared to wild type (WT) virus. Although attenuated in comparison to WT virus and single point mutants K65R, L74V and L74I; the double mutant K65R+L74I replicated efficiently in comparison to K65R+L74V mutant. The increased replication capacity of K65R+L74I viruses in comparison to K65R+L74V viruses was significant at multiplicity of infection 0.01 (p = 0.0004). Direct sequencing and sequencing after population cloning showed a more pronounced reversion at codon 65 in viruses containing K65R+L74V mutations in comparison to viruses with K65R+L74I mutations. In vitro processivity assays showed increased processivity of RT containing K65R+L74I in comparison to K65R+L74V RT.ConclusionsThe improved replication kinetics of K65R+L74I virus in comparison to K65R+L74V viruses was due to an increase in the processivity of RT containing K65R+L74I mutations. These observations support the rationale behind structural functional analysis to understand the interactions among unique RT mutations that may emerge during the treatment with specific drug regimens.

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Coexistence of the K65R/L74V and/or K65R/T215Y mutations on the same HIV-1 genome

Cited by 14 publications

References 12 publications

Tenofovir (TDF)-Selected or Abacavir (ABC)-Selected Low-Frequency HIV Type 1 Subpopulations During Failure with Persistent Viremia as Detected by Ultradeep Pyrosequencing

Tenofovir (TDF)-Selected or Abacavir (ABC)-Selected Low-Frequency HIV Type 1 Subpopulations During Failure with Persistent Viremia as Detected by Ultradeep Pyrosequencing

4′- C -Methyl-2′-Deoxyadenosine and 4′- C -Ethyl-2′-Deoxyadenosine Inhibit HIV-1 Replication

A Leu to Ile but not Leu to Val change at HIV-1 reverse transcriptase codon 74 in the background of K65R mutation leads to an increased processivity of K65R+L74I enzyme and a replication competent virus

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