Integrase Inhibitor Resistance Research Articles

Integrase variability and susceptibility to HIV integrase inhibitors: impact of subtypes, antiretroviral experience and duration of HIV infection

Little is known about the extent and predictors of polymorphisms potentially influencing the susceptibility to HIV integrase inhibitors (INIs). Genetic sequences of HIV integrase were obtained from INI-naive patients at two European clinics. The 39 amino acid changes at 29 integrase positions so far associated with INI resistance were examined according to HIV clade, prior antiretroviral exposure and duration of HIV infection. Integrase sequences were obtained from 418 patients, 294 (70.3%) infected with clade B and 124 (29.7%) infected with non-B variants (predominantly CRF02, A, C and D). Overall, 40% of patients were antiretroviral experienced and 32.8% were recent seroconverters. The most prevalent INI resistance-associated mutations were V72I (63.9%), V201I (54.8%), T206S (25.4%), I203M (9.8%) and K156N (7.4%). Major INI resistance mutations at positions 66, 92, 143, 148 and 155 were not detected. The mean number of polymorphic sites was greater in non-B than in B variants (2.17 versus 1.59; P < 0.001), and in antiretroviral-experienced than in drug-naive patients (1.89 versus 1.68; P = 0.034), whereas no significant differences were seen comparing recent seroconverters and chronically infected persons. Major INI resistance-associated mutations are very rare, if indeed ever present, in INI-naive patients. However, polymorphisms at positions which may influence the genetic barrier and/or drive the selection of specific INI resistance pathways are common, especially in HIV non-B subtypes.

Effects of HIV type-1 immune selection on susceptability to integrase inhibitor resistance

All site-specific interactions between HIV type-1 (HIV-1) subtype, human leukocyte antigen (HLA)-associated immune selection and integrase inhibitor resistance are not completely understood. We examined naturally occurring polymorphisms in HIV-1 integrase sequences from 342 antiretroviral-naive individuals from the Western Australian HIV Cohort Study and the Swiss HIV Cohort Study. Standard bulk sequencing and sequence-based typing were used to generate integrase sequences and high-resolution HLA genotypes, respectively. Viral residues were examined with respect to drug resistance mutations and CD8(+) T-cell escape mutations. In both predominantly subtype B cohorts, 12 of 38 sites that mediate integrase inhibitor resistance mutations were absolutely conserved, and these included the primary resistance mutations. There were 18 codons with non-primary drug resistance-associated substitutions at rates of up to 58.8% and eight sites with alternative polymorphisms. Five viral residues were potentially subject to dual-drug and HLA-associated immune selection in which both selective pressures either drove the same amino acid substitution (codons 72, 157 and 163) or HLA alleles were associated with an alternative polymorphism that would alter the genetic barrier to resistance (codons 125 and 193). The common polymorphism T125A, which was characteristic of non-subtype B and was also associated with carriage of HLA-B*57/*5801, increased the mutational barrier to the resistance mutation T125K. Primary integrase inhibitor resistance mutations were not detected in the absence of drug exposure in keeping with sites of high constraint. Viral polymorphisms caused by immune selection and/or associated with non-subtype B might alter the genetic barrier to some non-primary resistance-associated mutations.

Induced‐Fit Docking Approach Provides Insight into the Binding Mode and Mechanism of Action of HIV‐1 Integrase Inhibitors

A three-dimensional model of a complex between HIV-1 integrase (IN), viral DNA, and metal ions that we recently built was used as a target for a docking method (induced-fit docking, IFD) that accurately predicts ligand binding modes and concomitant structural changes in the receptor. Six different well-known integrase strand transfer inhibitors (INSTIs): L-708,906, L-731,988, S-1360, L-870,810, raltegravir, and elvitegravir were thus used as ligands for our docking simulations. The obtained IFD results are consistent with the mechanism of action proposed for this class of IN inhibitors, that is, metal chelating/binding agents. This study affords new insight into the possible mechanism of inhibition and binding conformations for INSTIs. The impact on our hypothesis of specific mutations associated with IN inhibitor resistance was also evaluated. All these findings might have implications for integrase-directed HIV-1 drug discovery efforts.

Genetic barriers for integrase inhibitor drug resistance in HIV type-1 B and CRF02_AG subtypes

HIV type-1 (HIV-1) integrase (IN) inhibitor resistance is the consequence of mutations that are selected in the viral IN gene targeted by antiretroviral drugs, such as raltegravir (RAL) and elvitegravir (EVG). The genetic barrier, defined as the number of viral mutations required to overcome the drug-selective pressure, is one of the important factors in the development of drug resistance. The genetic barrier for IN inhibitor resistance was compared between HIV-1 subtype B and HIV-1 subtype CRF02_AG, which is highly prevalent in West Africa and becoming more frequent in developed countries. IN nucleotide sequences from 73 HIV-1 subtype B and 77 HIV-1 subtype CRF02_AG antiretroviral-naive patients were examined at 19 IN amino acid positions implicated in RAL and EVG resistance. The majority (14/19) of the studied positions showed a high degree of conservation of the predominant codon sequences leading to a similar genetic barrier between subtypes B and CRF02_AG. Nevertheless, at positions 140 and 151, the variability between subtypes affected the genetic barrier for the mutations G140C, G140S and V1511 with a higher genetic barrier being calculated for subtype CRF02_AG. The major IN mutations E92Q, Q148K/R/H, N155H and E157Q (implicated in the resistance of IN inhibitors RAL and EVG) are highly conserved between subtypes B and CRF02_AG and display a similar genetic barrier. However, subtype CRFO2_AG showed a higher genetic barrier to acquire mutations 6140S, 6140C and V1511 as compared with subtype B, which could play a role in the resistance to RAL and/or EV6.

Implications of HIV-1 M Group Polymorphisms on Integrase Inhibitor Efficacy and Resistance: Genetic and Structural in Silico Analyses

The extensive polymorphisms among HIV-1 subtypes have been implicated in drug resistance development. Integrase inhibitors represent the latest addition to the treatment of HIV-1, and their efficacy and resistance patterns among M group strains are currently under investigation. This study analyzed the intersubtype variation within 108 integrase sequences from seven subtypes. The residues associated with catalytic activity and primary resistance to raltegravir were highly conserved among all strains. Variations were observed in residues associated with secondary resistance. Molecular modeling studies indicated a two-way binding mode of raltegravir that explains the resistance pathways and the implication of nonconservative mutations in integrase-raltegravir interactions.

Selection of the Q148R integrase inhibitor resistance mutation in a failing raltegravir containing regimen

Selection of the Q148R integrase inhibitor resistance mutation in a failing raltegravir containing regimen

HIV-2 integrase gene polymorphism and phenotypic susceptibility of HIV-2 clinical isolates to the integrase inhibitors raltegravir and elvitegravir in vitro

We investigated the in vitro phenotypic susceptibility of HIV-2 isolates from integrase inhibitor (INI)-naive patients to INIs and its relation to HIV-2 integrase gene polymorphism. We determined the phenotypic susceptibility to raltegravir and elvitegravir of co-cultured isolates obtained from the HIV-2 ROD reference strain and from 14 clinical isolates. IC(50) values were compared with those for HIV-1 reference strains. HIV-2 integrase gene polymorphism was assessed in isolates from 52 INI-naive patients enrolled in the French HIV-2 cohort. Median raltegravir and elvitegravir IC(50) values for the 14 clinical HIV-2 isolates were 2.4 and 0.7 nM, respectively, and were similar to those observed for HIV-2 ROD and HIV-1 reference strains. Overall, 38% of HIV-2 integrase amino acids were polymorphic. The catalytic triad DDE and the HHCC and RKK motifs were fully conserved, at the same genomic positions as described in HIV-1. In subtype B isolates, the total length of the integrase gene varied, owing to the presence of stop codons at positions 288, 294, 297 and 302. Fourteen of the positions associated with substitutions conferring INI resistance in HIV-1 were polymorphic in HIV-2. Despite 40% heterogeneity between the HIV-1 and HIV-2 integrase genes, the phenotypic susceptibility of clinical HIV-2 isolates to INIs was similar to that of HIV-1. This new class of antiretroviral drugs thus represents a novel therapeutic possibility for HIV-2-infected patients who otherwise have few treatment options.

Analysis of Natural Sequence Variation and Covariation in Human Immunodeficiency Virus Type 1 Integrase

Human immunodeficiency virus type 1 (HIV-1) integrase inhibitors are in clinical trials, and raltegravir and elvitegravir are likely to be the first licensed drugs of this novel class of HIV antivirals. Understanding resistance to these inhibitors is important to maximize their efficacy. It has been shown that natural variation and covariation provide valuable insights into the development of resistance for established HIV inhibitors. Therefore, we have undertaken a study to fully characterize natural polymorphisms and amino acid covariation within an inhibitor-naïve sequence set spanning all defined HIV-1 subtypes. Inter- and intrasubtype variation was greatest in a 50-amino-acid segment of HIV-1 integrase incorporating the catalytic aspartic acid codon 116, suggesting that polymorphisms affect inhibitor binding and pathways to resistance. The critical mutations that determine the resistance pathways to raltegravir and elvitegravir (N155H, Q148K/R/H, and E92Q) were either rare or absent from the 1,165-sequence data set. However, 25 out of 41 mutations associated with integrase inhibitor resistance were present. These mutations were not subtype associated and were more prevalent in the subtypes that had been sampled frequently within the database. A novel modification of the Jaccard index was used to analyze amino acid covariation within HIV-1 integrase. A network of 10 covarying resistance-associated mutations was elucidated, along with a further 15 previously undescribed mutations that covaried with at least two of the resistance positions. The validation of covariation as a predictive tool will be dependent on monitoring the evolution of HIV-1 integrase under drug selection pressure.

Integrase polymorphism and HIV-1 group O diversity

Integrase polymorphism and HIV-1 group O diversity

Integrase Inhibitors: A New Treatment Option for Patients with Human Immunodeficiency Virus Infection

The emergence of antiretroviral drug resistance in patients infected by the human immunodeficiency virus (HIV) has prompted efforts to develop new antiretrovirals that differ from existing agents with regard to mechanism of action and resistance profiles. We evaluated the literature regarding a new class of antiretrovirals, the integrase inhibitors. A MEDLINE search (January 1996-May 2007) was performed to identify relevant clinical trials and review articles; abstracts from HIV conferences were also searched. Raltegravir (MK-0518) and elvitegravir (GS-9137) are the two integrase inhibitors in late-phase development. These agents prevent viral DNA integration into the CD4(+) cell chromosome. Both drugs showed potent antiviral activity in large clinical trials that were performed in treatment-experienced, multidrug-resistant patients. Promising results have also been seen in an initial dose-ranging study with raltegravir in treatment-naïve patients. Preliminary data describe integrase inhibitor resistance profiles, but more data are needed in this area. Both agents were well tolerated in clinical trials, with favorable pharmaco-kinetic profiles for once- or twice-daily dosing. Raltegravir and elvitegravir differ in their metabolism, resulting in distinct drug-interaction profiles for each agent. Based on available data, this new class of antiretrovirals will soon be widely used in antiretroviral-experienced patients infected with HIV. In the future, this class of drugs may become a reasonable treatment option for antiretroviral-naïve patients, but more data are needed in that patient population.

Natural Polymorphism of the HIV-1 Integrase Gene and Mutations associated with Integrase Inhibitor Resistance

Two inhibitors of the HIV-1 integrase enzyme (INIs) are in late stage clinical development. To date, approximately 42 mutations within the HIV-1 integrase (IN) gene have been associated with INI drug resistance. Naturally occurring IN gene polymorphisms may have important implications for INI development. In this study, we evaluated clinical HIV-1 strains from INI-naive patients to determine the prevalence of IN gene polymorphisms, and the frequency of naturally occurring amino acid (aa) substitutions at positions associated with INI resistance and at sites crucial for LEDGF/p75 binding and HIV-1 integration. The IN gene from 67 INI-naive, HIV-1 clade B-infected patients were sequenced using standard population-based DNA sequencing methods. In addition, 176 unique full-length HIV-1 clade B IN gene sequences from INI-naive patients obtained from the HIV Los Alamos database were analysed. Analysis of 243 IN genes from HIV-1 clade B, INI-naive clinical strains revealed that 64% of the aa positions were polymorphic. Of the 42 aa substitutions currently associated with INI resistance, 21 occurred as natural polymorphisms: V72I, L74I, T97A, T112I, A128T, E138K, Q148H, V151I, S153Y/A, M154I, N155H, K156N, E157Q, G163R, V165I, V201I, I203M, T206S, S230N and R263K. IN aa positions crucial to LEDGF/P75 binding and HIV-1 integration were well conserved. Major INI mutations within the catalytic domain and extended active sites associated with high level resistance to the compounds in late stage development, especially strand transfer inhibitors (STIs), were infrequent in our study, which may help explain the excellent virological responses demonstrated in clinical trials.

Human immunodeficiency virus type-1 integrase containing a glycine to serine mutation at position 140 is attenuated for catalysis and resistant to integrase inhibitors

L-chicoric acid (L-CA) is a potent inhibitor of HIV integrase (IN) in vitro. In this report, the effects of a glycine to serine mutation at position 140 (G140S) on HIV IN and its effects on IN inhibitor resistance are described. HIV containing the G140S mutation showed a delay in replication. Using real-time polymerase chain reaction, the delay was secondary to a failure in integration. The mutant protein (IN G140S) was attenuated approximately four-fold for catalysis under equilibrium conditions compared to wild-type IN (IN WT) and attenuated five-fold in steady-state kinetic analysis of disintegration. Fifty percent inhibitory concentration assays were performed with IN inhibitors against both IN proteins in disintegration and strand transfer reactions. IN G140S was resistant to both L-CA and L-731,988, a diketoacid. HIV containing the mutation was resistant to both inhibitors as well. The G140S mutation attenuates IN activity and confers resistance to IN inhibitors, suggesting that diketoacids and L-CA interact with a similar binding site on HIV IN.