HIV protease genotype and viral sensitivity to HIV protease inhibitors following saquinavir therapy

Craig, Charles; Race, Esther; Sheldon, Jonathan G.; Whittaker, Lynne; Gilbert, Sue; Moffatt, Alec; Rose, Jane S. L.; Dissanayeke, Shobana R.; Chirn, Gung-Wei; Duncan, I. B. R.; Cammack, Nick

doi:10.1097/00002030-199813000-00007

Cited by 56 publications

(34 citation statements)

References 24 publications

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“…Subsequently, isolates from patients began to show mutations in the protease gene that correlated directly to in vivo resistance not only to indinavir but also to a diverse panel of HIV-1 protease inhibitors (6). In similar fashions, cross-resistance was also observed for viral isolates from patients treated with other inhibitors (7)(8)(9)(10)(11).…”

Section: Resultsmentioning

confidence: 61%

Non-active Site Changes Elicit Broad-based Cross-resistance of the HIV-1 Protease to Inhibitors

Olsen

Stahlhut

Rutkowski

et al. 1999

Journal of Biological Chemistry

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Three high level, cross-resistant variants of the HIV-1 protease have been analyzed for their ability to bind four protease inhibitors approved by the Food and Drug Administration (saquinavir, ritonavir, indinavir, and nelfinavir) as AIDS therapeutics. The loss in binding energy (⌬⌬G b ) going from the wild-type enzyme to mutant enzymes ranges from 2.5 to 4.4 kcal/mol, 40 -65% of which is attributed to amino acid substitutions away from the active site of the protease and not in direct contact with the inhibitor. The data suggest that nonactive site changes are collectively a major contributor toward engendering resistance against the protease inhibitor and cannot be ignored when considering crossresistance issues of drugs against the HIV-1 protease.The objective of anti-retroviral therapy for human immunodeficiency virus type 1 (HIV-1) 1 infection is complete viral suppression below the limits of detection. This goal has not been routinely achieved until the development of potent inhibitors targeted against the HIV-1 protease, an enzyme essential for viral replication. One of the greatest barriers to achieving long term viral suppression is the emergence of drug-resistant strains of HIV. The appearance of drug-resistant viruses and the evaluation of drug cross-resistance for a given enzymatic target (reverse transcriptase or protease) have been the most serious issues in the treatment of HIV-infected individuals. The correlation of active site mutations in HIV-1 protease to decreases in inhibitor binding has been well documented, whereas the role of non-active site mutations has not been clearly defined (1). To date, several non-active site changes have been described as compensatory mutations that tend to offset the detrimental effects of active site mutations toward enzyme catalysis (2, 3), but the effect of non-active site mutations toward binding has hitherto not been analyzed quantitatively. In this report, our findings show that non-active site amino acid substitutions are major factors leading to the decreases in inhibitor binding to three clinically derived variants of the HIV-1 protease. MATERIALS AND METHODSEnzyme Preparation-Synthetic oligonucleotide cassettes of 444 base pairs were designed according to the wild-type sequence of pET-3b-HIVPR (4). Point mutations were incorporated into the DNA with a bias toward optimal codon usage in Escherichia coli to yield the amino acid mutations listed in Table I. The oligonucleotides were annealed and ligated into pUC18 or pUC19 by Midland Certified Reagent Co. The primary sequence was verified before subcloning into a pET-3b expression vector via NdeI and Bpu1102I sites and reconfirmed by automated double-stranded DNA sequencing. Clones carrying the mutant DNA were transformed and expressed as described previously (3,5). The cells were lysed in 50 mM Tris-HCl, pH 8.0, 1 mM EDTA, 0.1% Nonidet P-40, 10 mM MgCl 2 , and 100 g/ml DNase I using a microfluidizer processor (Microfluidics International Corp., Newton, MA). The mutant protease was extracted, refolded, a...

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Section: Resultsmentioning

confidence: 61%

Non-active Site Changes Elicit Broad-based Cross-resistance of the HIV-1 Protease to Inhibitors

Olsen

Stahlhut

Rutkowski

et al. 1999

Journal of Biological Chemistry

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“…In the United States and Europe, about 10% of new infections are with HIV-1 strains harboring resistance to at least one of three classes of anti-HIV drugs (16,23,30,93,144,233,234,317,357,372,384,416 [61][62]1999) (Table 3). …”

Section: Evolution Of Hiv-1 Drug Resistancementioning

confidence: 99%

“…The spectrum of mutations developing during therapy with indinavir, nelfinavir, saquinavir, and ritonavir have been well characterized (9,24,54,61,265,281,320,334), but fewer data are available for amprenavir (237a) and lopinavir (35). The dynamic susceptibility range for indinavir, ritonavir, saquinavir, nelfinavir, and lopinavir is about 100-fold in most drug susceptibility assays (148,149,290,393; Brun, S., D. Kempf, J. Isaacson, A. Molla, H. Mo, C. Benson, and E. Sun, abstract 452, 8th Conference on Retroviruses and Opportunistic Infections, Chicago, Ill., 2001).…”

Section: Pismentioning

confidence: 99%

Genotypic Testing for Human Immunodeficiency Virus Type 1 Drug Resistance

2002

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There are 16 approved human immunodeficiency virus type 1 (HIV-1) drugs belonging to three mechanistic classes: protease inhibitors, nucleoside and nucleotide reverse transcriptase (RT) inhibitors, and nonnucleoside RT inhibitors. HIV-1 resistance to these drugs is caused by mutations in the protease and RT enzymes, the molecular targets of these drugs. Drug resistance mutations arise most often in treated individuals, resulting from selective drug pressure in the presence of incompletely suppressed virus replication. HIV-1 isolates with drug resistance mutations, however, may also be transmitted to newly infected individuals. Three expert panels have recommended that HIV-1 protease and RT susceptibility testing should be used to help select HIV drug therapy. Although genotypic testing is more complex than typical antimicrobial susceptibility tests, there is a rich literature supporting the prognostic value of HIV-1 protease and RT mutations. This review describes the genetic mechanisms of HIV-1 drug resistance and summarizes published data linking individual RT and protease mutations to in vitro and in vivo resistance to the currently available HIV drugs

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“…Assay results along with the mutational makeup of each isolate tested are summarized in Table 1. Sequence analysis of the HIV-1 RF variants selected in cell culture by IDV, NFV, and RTV revealed that the Prt substitutions present were mostly representative of those identified in clinical trials (1,4,5,6,26,33,38). However, the resistant RF strain selected with APV contained only the mutation V82I Table 1 showed that both the clinical and laboratory IDV-resistant isolates (15-and 24-fold decreases in sensitivity to IDV, respectively) displayed 6-to 9-fold resistance to BMS-232632, NFV, and SQV.…”

Section: Bms-232632 Susceptibilities Of Hiv-1 Strains Resistant To Cumentioning

confidence: 99%

“…The current group of Prt inhibitors select for distinct but overlapping sets of amino acid substitutions within the Prt molecule. The key signature substitutions for IDV and RTV resistance reside at amino acid residues V82, I84, or L90, those for SQV resistance reside at G48, I84, or L90, those for NFV resistance reside at D30 or L90, and those for APV resistance reside at I50 or I84 (1,4,5,6,7,10,11,15,16,22,23,26,30,32,33,36,38; M. Tisdale, R. E. Myers, M. Al T-Khaled, and W. Snowden, 6th Conf. Retrovir.…”

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confidence: 99%

In Vitro Resistance Profile of the Human Immunodeficiency Virus Type 1 Protease Inhibitor BMS-232632

Gong¹,

Robinson²,

Rose³

et al. 2000

Antimicrob Agents Chemother

123

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BMS-232632 is an azapeptide human immunodeficiency virus (HIV) type 1 (HIV-1) protease inhibitor that displays potent anti-HIV-1 activity (50% effective concentration [EC 50 ], 2.6 to 5.3 nM; EC 90 , 9 to 15 nM). In vitro passage of HIV-1 RF in the presence of inhibitors showed that BMS-232632 selected for resistant variants more slowly than nelfinavir or ritonavir did. Genotypic and phenotypic analysis of three different HIV strains resistant to BMS-232632 indicated that an N88S substitution in the viral protease appeared first during the selection process in two of the three strains. An I84V change appeared to be an important substitution in the third strain used. Mutations were also observed at the protease cleavage sites following drug selection. The evolution to resistance seemed distinct for each of the three strains used, suggesting multiple pathways to resistance and the importance of the viral genetic background. A cross-resistance study involving five other protease inhibitors indicated that BMS-232632-resistant virus remained sensitive to saquinavir, while it showed various levels (0.1-to 71-fold decrease in sensitivity)-of cross-resistance to nelfinavir, indinavir, ritonavir, and amprenavir. In reciprocal experiments, the BMS-232632 susceptibility of HIV-1 variants selected in the presence of each of the other HIV-1 protease inhibitors showed that the nelfinavir-, saquinavir-, and amprenavir-resistant strains of HIV-1 remained sensitive to BMS-232632, while indinavir-and ritonavirresistant viruses displayed six-to ninefold changes in BMS-232632 sensitivity. Taken together, our data suggest that BMS-232632 may be a valuable protease inhibitor for use in combination therapy. , integrase, and Prt) (18). The Prt functions at the late stages of viral replication during virion maturation and has proved to be an effective target for antiviral intervention. Currently, five peptidic Prt inhibitors, saquinavir (SQV), indinavir (IDV), ritonavir (RTV), nelfinavir (NFV), and amprenavir (APV), are approved for clinical use (7,19,30,32,41). This class of drugs suppresses viral replication to a greater extent than the RT inhibitors in HIV-1-infected patients (12,13,24,25,27,28,42). Today, the standard care for AIDS patients involves the use of two RT inhibitors and one Prt inhibitor to reduce viremia to unquantifiable levels for an extended period of time (2, 13, 14, 27, 29; M. Markowitz, Y. Cao, A. Hurley, R. Schluger, S. Monard, R. Kost, B. Kerr, R. Anderson, S. Eastman, and D. D. Ho, 5th Conf. Retrovir. Opportunistic Infections, abstr. 371, 1998). Despite such a remarkable result, 30 to 50% of patients ultimately fail therapy, presumably due to patient nonadherence to drug schedules (as a consequence of inconvenient dosing and side effects) (43), insufficient drug exposure, and resistance development. Therefore, additional Prt inhibitors that display greater potency, improved bioavailability, fewer side effects, and distinct resistance profiles are needed.The emergence of resistant variants results from the larg...

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HIV protease genotype and viral sensitivity to HIV protease inhibitors following saquinavir therapy

Cited by 56 publications

References 24 publications

Non-active Site Changes Elicit Broad-based Cross-resistance of the HIV-1 Protease to Inhibitors

Non-active Site Changes Elicit Broad-based Cross-resistance of the HIV-1 Protease to Inhibitors

Genotypic Testing for Human Immunodeficiency Virus Type 1 Drug Resistance

In Vitro Resistance Profile of the Human Immunodeficiency Virus Type 1 Protease Inhibitor BMS-232632

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