Polymorphic substitution E157Q in HIV-1 integrase increases R263K-mediated dolutegravir resistance and decreases DNA binding activity

Anstett, Kaitlin; Cutillas, Vincent; Fusco, Robert; Mésplède, Thibault; Wainberg, Mark A.

doi:10.1093/jac/dkw109

Cited by 41 publications

(26 citation statements)

References 23 publications

(33 reference statements)

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“…Whether this situation persists or whether a few individuals who are infected with viruses that can develop de novo resistance against this drug will eventually be identified is the subject of anxious scrutiny. An eventual de novo virological failure with resistance mutations may derive from preexisting rare polymorphisms, including E157Q and others (23, 24). In any case, to date, DTG has demonstrated an exceptional robustness against the emergence of drug resistance mutations in treatment-naive individuals.…”

Section: Discussionmentioning

confidence: 99%

The R263K Dolutegravir Resistance-Associated Substitution Progressively Decreases HIV-1 Integration

Mésplède

Leng

Pham

et al. 2017

mBio

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Human immunodeficiency virus (HIV) infection persists despite decades of active antiretroviral therapy (ART), effectively preventing viral eradication. Treatment decreases plasma viral RNA, but viral DNA persists, mostly integrated within the genome of nucleated blood cells. Viral DNA blood levels correlate with comorbidities and the rapidity of viral rebound following treatment interruption. To date, no intervention aiming at decreasing HIV DNA levels below those attained through ART has been successful. This includes use of some integrase inhibitors either as part of ART or in treatment intensification studies. We have argued that using the integrase inhibitor dolutegravir (DTG) in similar studies may yield better results, but this remains to be studied. In treatment-experienced individuals, the most frequent substitution associated with failure with dolutegravir is R263K in integrase. R263K decreases integration both in cell-free and tissue culture assays. We investigated here how integrated DNA levels evolve over time during prolonged infections with R263K viruses. To investigate a potential defect in reverse transcription with R263K, the levels of reverse transcripts were measured by quantitative PCR. We measured HIV type 1 (HIV-1) integration in Jurkat cells over the course of 4-week infections using Alu-mediated quantitative PCR. The results show that R263K did not decrease reverse transcription. Prolonged infections with R263K mutant viruses led to less HIV-1 integrated DNA over time compared to wild-type viruses. These tissue culture results help to explain the absence of the R263K substitution in most individuals experiencing failure with DTG and support studies aiming at longitudinally measuring the levels of integrated DNA in individuals treated with this drug.

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Section: Discussionmentioning

confidence: 99%

The R263K Dolutegravir Resistance-Associated Substitution Progressively Decreases HIV-1 Integration

Mésplède

Leng

Pham

et al. 2017

mBio

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“…Indeed, rare DTG failures are often associated with the R263K substitution, whereas RAL and EVG select for classical resistance mutations, including mutations at positions G140 plus Q148 or the N155H mutation. We previously reported that the R263K mutation may in large part be incompatible with classical resistance mutations for reasons of decreased replicative fitness and low-level resistance (32)(33)(34). Whether the resistance levels of mutants combining the R263K mutation with classical resistance mutations are also susceptible to acetylation inhibition remains to be investigated.…”

Section: Discussionmentioning

confidence: 99%

“…Further investigation is under way to pinpoint the exact cause of our observed results through use of mass spectrometry analysis and the study of IN-DNA interactions by use of multiple C-terminal mutants. We also previously reported an effect of the R263K substitution on IN DNA binding activity in cell-free assays (11,32). While this may also be considered a mechanism through which the substitution provides resistance, it is important that these studies were performed with recombinant enzymes purified from Escherichia coli that most likely lack proper posttranslational modifications (37).…”

Section: Discussionmentioning

confidence: 99%

HIV-1 Resistance to Dolutegravir Is Affected by Cellular Histone Acetyltransferase Activity

Anstett

Brenner

Mésplède

et al. 2017

J Virol

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Integrase strand transfer inhibitors (INSTIs) are the newest class of antiretrovirals to be approved for the treatment of HIV infection. Canonical resistance to these competitive inhibitors develops through substitutions in the integrase active site that disrupt drug-protein interactions. However, resistance against the newest integrase inhibitor, dolutegravir (DTG), is associated with an R263K substitution at the C terminus of integrase that causes resistance through an unknown mechanism. The integrase C-terminal domain is involved in many processes over the course of infection and is posttranslationally modified via acetylation of three lysine residues that are important for enzyme activity, integrase multimerization, and protein-protein interactions. Here we report that regulation of the acetylation of integrase is integral to the replication of HIV in the presence of DTG and that the R263K mutation specifically disrupts this regulation, likely due to enhancement of interactions with the histone deacetylase I complex, as suggested by coimmunoprecipitation assays. Although no detectable differences in the levels of cell-free acetylation of the wild-type (WT) and mutated R263K enzymes were observed, the inhibition of cellular histone acetyltransferase enzymes sensitized the NL4.3 virus to DTG, while NL4.3 was almost completely unaffected. When levels of endogenous acetylation were manipulated in virus-producing cells, inhibitors of acetylation enhanced the replication of NL4.3, whereas inhibition of deacetylation greatly diminished the replication of NL4.3 Taken together, these results point to a pivotal role of acetylation in the resistance mechanism of HIV to some second-generation integrase strand transfer inhibitors, such as DTG. This is, to our knowledge, the first report of the influence of posttranslational modifications on HIV drug resistance. Both viral replication and resistance to second-generation integrase strand transfer inhibitors of both WT and INSTI-resistant HIV strains were differentially affected by acetylation, likely as a result of altered interactions between integrase and the cellular deacetylation machinery. Many "shock and kill" strategies to eradicate HIV manipulate endogenous levels of acetylation in order to reactivate latent HIV. However, our results suggest that some drug-resistant viruses may differentially respond to such stimulation, which may complicate the attainment of this goal. Our future work will further illuminate the mechanisms involved.

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“…It has also been reported in a patient with virological failure on a DTG-containing regimen [28]. It appears to reduce RAL and EVG susceptibility by 2-to 3-fold [29,30] and to increase DTG susceptibility [31]. Seven (13.2%) patients had the HIV-1 integrase mutation L74I or L74M detected in 4 different HIV-1 subtypes, 3 in A1, 1 in B, 2 in C, and 1 in CRF02_AG.…”

Section: Discussionmentioning

confidence: 77%

Resistance-Associated Mutations and Polymorphisms among Integrase Inhibitor-Naïve HIV-1 Patients in Kuwait

Chehadeh¹,

Albaksami²,

John³

et al. 2017

Intervirology

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Objectives: Resistance-associated mutations (RAMs) in the integrase of different HIV-1 subtypes were investigated in a cohort of patients never exposed to integrase strand transfer inhibitors (INSTIs). Methods: The viral RNA was extracted from plasma samples of 53 INSTI-naïve patients, and the integrase genetic region was sequenced and analyzed for subtype assignment and drug resistance. Results: The median viral load at sampling was 5.28 × 10⁴ RNA copies/mL. Bayesian phylogenetic analysis showed 85% of the HIV-1 isolates were non-B subtypes, with a predominance of subtypes C (22.6%) and CRF01_AE (26.4%). A total of 52 and 110 mutations were found in the integrase region of HIV-1 B and non-B subtypes, respectively. Nonpolymorphic INSTI-RAMs were not detected in this study. However, the accessory mutation E157Q was found in 1 patient with CRF02_AG, and the polymorphic mutations L74M/I that may contribute to a reduced susceptibility to INSTIs in the presence of major mutations were observed in 6 (13.3%) patients with non-B subtypes and 1 (12.5%) patient with the B subtype. Polymorphic mutations at positions known to harbor primary and accessory RAMs were also detected in this study. Conclusion: Our results highlight the importance of monitoring the emergence of INSTI-RAMS before and after the initiation of INSTI-based therapy.

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Polymorphic substitution E157Q in HIV-1 integrase increases R263K-mediated dolutegravir resistance and decreases DNA binding activity

Abstract: This study shows that E157Q may act as a compensatory mutation for R263K. Since E157Q is a natural polymorphism present in 1%-10% of HIV-positive individuals, it may be of particular importance for patients receiving INSTI therapy.

Cited by 41 publications

References 23 publications

The R263K Dolutegravir Resistance-Associated Substitution Progressively Decreases HIV-1 Integration

The R263K Dolutegravir Resistance-Associated Substitution Progressively Decreases HIV-1 Integration

HIV-1 Resistance to Dolutegravir Is Affected by Cellular Histone Acetyltransferase Activity

Resistance-Associated Mutations and Polymorphisms among Integrase Inhibitor-Naïve HIV-1 Patients in Kuwait

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