Biochemical Analysis of HIV-1 Integrase Variants Resistant to Strand Transfer Inhibitors

Dicker, Ira B.; Terry, Brian; Lin, Zheng‐Zhe; Li, Zhufang; Bollini, Sagarika; Samanta, Himadri; Gali, Volodymyr; Walker, Michael; Krystal, Mark

doi:10.1074/jbc.m804213200

Cited by 30 publications

(27 citation statements)

References 43 publications

(59 reference statements)

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“…InSTI mutations affected IC 50 but not m. Increases in IC 50 can be attributed to decreased affinity, resulting in the need for higher drug concentrations for inhibition. Biochemical analyses of integrase mutants indicate that the off-rate is remarkably increased relative to WT (53). This finding, coupled with our observation of a constant slope close to 1, indicates that integrase inhibitors affect WT and mutant viruses by blocking a noncooperative reaction.…”

Section: Analysis Of Resistance Mutations Using Primary Cells Vs Trasupporting

confidence: 61%

Dose–response curve slope is a missing dimension in the analysis of HIV-1 drug resistance

Sampah¹,

Shen

Jilek³

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

126

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HIV-1 drug resistance is a major clinical problem. Resistance is evaluated using in vitro assays measuring the fold change in IC 50 caused by resistance mutations. Antiretroviral drugs are used at concentrations above IC 50 , however, and inhibition at clinical concentrations can only be predicted from IC 50 if the shape of the dose-response curve is also known. Curve shape is influenced by cooperative interactions and is described mathematically by the slope parameter or Hill coefficient (m). Implicit in current analysis of resistance is the assumption that mutations shift dose-response curves to the right without affecting the slope. We show here that m is altered by resistance mutations. For reverse transcriptase and fusion inhibitors, single resistance mutations affect both slope and IC 50 . For protease inhibitors, single mutations primarily affect slope. For integrase inhibitors, only IC 50 is affected. Thus, there are fundamental pharmacodynamic differences in resistance to different drug classes. Instantaneous inhibitory potential (IIP), the log inhibition of single-round infectivity at clinical concentrations, takes into account both slope and IC 50 , and thus provides a direct measure of the reduction in susceptibility produced by mutations and the residual activity of drugs against resistant viruses. The standard measure, fold change in IC 50 , does not correlate well with changes in IIP when mutations alter slope. These results challenge a fundamental assumption underlying current analysis of HIV-1 drug resistance and suggest that a more complete understanding of how resistance mutations reduce antiviral activity requires consideration of a previously ignored parameter, the dose-response curve slope.HIV/AIDS | pharmacology | virology | highly active antiretroviral therapy | evolution

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Section: Analysis Of Resistance Mutations Using Primary Cells Vs Trasupporting

confidence: 61%

Dose–response curve slope is a missing dimension in the analysis of HIV-1 drug resistance

Sampah¹,

Shen

Jilek³

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

126

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“…Michaelis-Menten enzyme analysis has previously been used to determine enzyme parameters of HIV integrase (24). While 3= processing has been conclusively shown to follow nonMichaelis-Menten kinetics (25,26), there has been no similar study of HIV integrase strand transfer.…”

Section: Methodsmentioning

confidence: 99%

“…The data within the linear phase of the 3= processing reaction were used to calculate kinetic parameters by fitting the data to the Michaelis-Menten equation (24). As such, relative enzyme efficiency calculations of 3= processing activity were calculated as described above for strand transfer.…”

Section: Methodsmentioning

confidence: 99%

Differential Effects of the G118R, H51Y, and E138K Resistance Substitutions in Different Subtypes of HIV Integrase

Quashie

Oliviera

Veres

et al. 2015

J Virol

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Dolutegravir (DTG) is the latest antiretroviral (ARV) approved for the treatment of human immunodeficiency virus (HIV) infec-tion. The G118R substitution, previously identified with MK-2048 and raltegravir, may represent the initial substitution in a dolutegravir resistance pathway. We have found that subtype C integrase proteins have a low enzymatic cost associated with the G118R substitution, mostly at the strand transfer step of integration, compared to either subtype B or recombinant CRF02_AG proteins. Subtype B and circulating recombinant form AG (CRF02_AG) clonal viruses encoding G118R-bearing integrases were severely restricted in their viral replication capacity, and G118R/E138K-bearing viruses had various levels of resistance to dolutegravir, raltegravir, and elvitegravir. In cell-free experiments, the impacts of the H51Y and E138K substitutions on resistance and enzyme efficiency, when present with G118R, were highly dependent on viral subtype. Sequence alignment and homology modeling showed that the subtype-specific effects of these mutations were likely due to differential amino acid residue networks in the different integrase proteins, caused by polymorphic residues, which significantly affect native protein activity, structure, or function and are important for drug-mediated inhibition of enzyme activity. This preemptive study will aid in the interpretation of resistance patterns in dolutegravir-treated patients. IMPORTANCERecognized drug resistance mutations have never been reported for naive patients treated with dolutegravir. Additionally, in integrase inhibitor-experienced patients, only R263K and other previously known integrase resistance substitutions have been reported. Here we suggest that alternate resistance pathways may develop in non-B HIV-1 subtypes and explain how "minor" polymorphisms and substitutions in HIV integrase that are associated with these subtypes can influence resistance against dolutegravir. This work also highlights the importance of phenotyping versus genotyping when a strong inhibitor such as dolutegravir is being used. By characterizing the G118R substitution, this work also preemptively defines parameters for a potentially important pathway in some non-B HIV subtype viruses treated with dolutegravir and will aid in the inhibition of such a virus, if detected. The general inability of strand transfer-related substitutions to diminish 3= processing indicates the importance of the 3= processing step and highlights a therapeutic angle that needs to be better exploited.

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“…Of note, biochemical studies have suggested that binding of INIs to HIV-1 IN proceeds by a two-step mechanism with a slow second step, and mutations that increase resistance may alter this mechanism (19,32). In addition, fast INI dissociation has been proposed to contribute to INI resistance with the N155H, T66I, and Q148R IN substitutions (14,23).…”

mentioning

confidence: 99%

Dolutegravir (S/GSK1349572) Exhibits Significantly Slower Dissociation than Raltegravir and Elvitegravir from Wild-Type and Integrase Inhibitor-Resistant HIV-1 Integrase-DNA Complexes

Hightower

Wang

Deanda

et al. 2011

Antimicrob Agents Chemother

203

192

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Biochemical Analysis of HIV-1 Integrase Variants Resistant to Strand Transfer Inhibitors

Cited by 30 publications

References 43 publications

Dose–response curve slope is a missing dimension in the analysis of HIV-1 drug resistance

Dose–response curve slope is a missing dimension in the analysis of HIV-1 drug resistance

Differential Effects of the G118R, H51Y, and E138K Resistance Substitutions in Different Subtypes of HIV Integrase

Dolutegravir (S/GSK1349572) Exhibits Significantly Slower Dissociation than Raltegravir and Elvitegravir from Wild-Type and Integrase Inhibitor-Resistant HIV-1 Integrase-DNA Complexes

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