Indinavir Monotherapy Research Articles

Short Communication: Profiling Resistance-Related Mutations in the Protease Region of the pol Gene: Single Genome Sequencing of HIV in Plasma and Peripheral Blood Mononuclear Cells

Genotypic resistance is currently assessed through direct sequencing, which cannot detect resistant strains below 20%. We compared the genotypic resistance profile of virions and proviruses using population-based analysis and single genome sequencing of the protease region of the pol gene in samples collected from five individuals in whom indinavir monotherapy resulted in treatment failure. Single genome sequencing showed that not all strains present the same resistance mutations, which can be dispersed across different HIV genomes. The resistance profile found in plasma was very similar to that found in peripheral blood mononuclear cells (PBMCs), confirming the utility of assessing proviral DNA as for a means of determining antiretroviral resistance.

Analysis of the protease sequences of HIV-1 infected individuals after Indinavir monotherapy.

Protease inhibitors (PI) are an important HIV-1 treatment tool. The HIV-1 genetic diversity as a result of antiretroviral exposure is a potential barrier to successful antiretroviral therapy. To describe the impact of the selective pressure of the PI Indinavir in the protease region of the pol gene of HIV-1. We have examined the extent of protease sequence heterogeneity in previously antiretroviral drug naive HIV-1 infected individuals receiving Indinavir as monotherapy for at least 48 weeks. Analysis based on the consensus of this group of sequences showed regions with higher and lower polymorphism. The degree of genetic variation was greater in regions less critical for the structure and function of the enzyme. To investigate the selective pressure imposed by drug therapy, we have analyzed the rate of synonymous (ds) and nonsynonymous (dn) substitutions. The three critical regions for enzyme activity showed ds/dn ratio >1. whereas other regions had ds/dn ratio <1. The detected amino acid mutations had a trend to be conservative, thus maintaining the physical chemical amino acid characteristics. Phylogenetic analysis established the presence of subtype B (n=38), subtype F (n=9), and B/F recombinants within the protease region of pol gene (n=3). More prevalent detected mutations, thought to contribute to antiretroviral resistance, were L63P (42%), L10I (35%), M36I (30%), V82A/T/F (26%). A great deal of predicted cross-resistance between PIs was observed. Out of the 50 individuals, 34% were considered to have major mutations to Indinavir, and 66% had minor or no mutations to Indinavir. Viral loads were significantly higher among patients with major mutations, compared with patients minor/no mutations, although no differences in the CD4 counts were found. The viral load at baseline and nadir (week 4) was able to predict the group of individuals with greater chances of selecting drug resistance related mutations.

AIDS, activism, and the regulation of clinical trials in Brazil: Protocol 028

This paper examines the politics and practices of drug evaluation in Brazil. It traces the history of AIDS activists' influence on the organization of modern clinical trials and their scientific rationale. Using the Merck indinavir trial as a case study, the authors discuss how organized civil society has developed strategies to intervene in the course of drug evaluation trials, shaping them according to its own interests. Adopting translation sociology as the theoretical framework, the paper describes and analyzes the strategies used by activists from "Grupo PelaVidda/SP" (an AIDS NGO) to build a consensus concerning indinavir monotherapy's lack of efficacy. The study considers the several regulatory forums involved in dealing with the controversy during the trial period.

The Effect of Nevirapine in Combination With Nelfinavir in Heavily Pretreated HIV-1–Infected Patients

To determine whether initiation of antiretroviral therapy that includes the protease inhibitor indinavir causes insulin resistance or abnormal B-cell function in study subjects with HIV infection. Methods: Nonwasted, HIV-infected study subjects who did not have concurrent diabetes were prospectively evaluated by oral and intravenous glucose tolerance testing at baseline, at 2 weeks after starting indinavir monotherapy, and at another 6 weeks after initiating indinavir-based triple-therapy. Results: Mean CD4 count at entry was 282 cells/μl and median HIV RNA was 33,000 copies/ml; all experienced a virologic response. Fasting glucose increased from 83.2 ± 3.7 mg/dl at baseline to 86.8 ± 3.2 at week 2 and 91.7 ± 3.5 at week 8 (p = .003). Insulin sensitivity by minimal model analysis decreased by 30.5% over 8 weeks, from 3.83 ± 0.63 min-1 per μU/ml × 10-4 to 3.09 ± 0.53 at week 2 and 2.66 ± 0.35 at week 8 (p = .01). Insulin secretion by the acute insulin response to intravenous glucose did not change (baseline 822 ± 283 μU/ml × min, week 8 880 ± 289; p = 0.4), and the insulin response to oral glucose (30 minute insulin:glucose ratio) fell from 1.69 ± 0.54 μU/ml per mg/dl at baseline to 1.18 ± 0.34 at week 8 (p = .05). Conclusion: During 8 weeks of indinavir-based therapy, fasting glucose increased and insulin sensitivity decreased, without a compensatory increase in insulin release. This combination of insulin resistance without augmented B-cell response may explain the hyperglycemia and other metabolic abnormalities seen in some protease inhibitortreated patients.

Dynamics of HIV-1 viral load rebound among patients with previous suppression of viral replication.

To model the dynamics of HIV-1 rebound in patients receiving suboptimal therapy after suppression of plasma viremia to < 200 copies/ml by triple combination therapy. Mathematical modeling of data from 23 patients switched to indinavir maintenance therapy after viral replication was suppressed with a combination of indinavir, zidovudine and lamivudine. Modeling of HIV-1 rebound among 24 patients on zidovudine/lamivudine maintenance was also performed for comparison. Evaluation of slopes of rebound and of their heterogeneity; calculation of the basic reproductive number (Ro, the number of newly infected cells arising from each productively infected cell); regression analyses for predictors of the slope of rebound. Rebound of plasma HIV RNA followed a sigmoid curve with an initial exponential phase. There was significant heterogeneity in the slopes of rebound for individual patients (P < 0.001). In the indinavir maintenance rebounds, the average initial slope was estimated to be 0.587/day (doubling time 1.2 days). The slopes of rebound in patients on zidovudine/lamivudine maintenance tended to be less steep on average (P = 0.025). Among patients taking indinavir maintenance, the average Ro for the initial rebound of viremia was 4.3; in multivariate regressions, the slope of rebound was steeper during early rebound and in patients with higher viral load at the start of triple therapy or a higher CD4 cell count when indinavir monotherapy was initiated. The slope was less steep in patients with a greater increase in the number of CD4 cells during triple therapy. The rates of viral load increase among patients with viral rebound while receiving less than triple therapy are similar to those reported in patients interrupting therapy. Variability among patients may depend on viral fitness, target cell availability and extent of immune reconstitution.

Drug susceptibility in HIV infection after viral rebound in patients receiving indinavir-containing regimens.

Loss of viral suppression in patients infected with human immunodeficiency virus (HIV), who are receiving potent antiretroviral therapy, has been attributed to outgrowth of drug-resistant virus; however, resistance patterns are not well characterized in patients whose protease inhibitor combination therapy fails afterachieving viral suppression. To characterize drug susceptibility of virus from HIV-infected patients who are failing to sustain suppression while taking an indinavir-containing antiretroviral regimen. Substudy of the AIDS Clinical Trials Group 343, a multicenter clinical research trial conducted between February 1997 and October 1998. Twenty-six subjects who experienced rebound (HIV RNA level > or =200 copies/mL) during indinavir monotherapy (n = 9) or triple-drug therapy (indinavir, lamivudine, and zidovudine; n = 17) after initially achieving suppression while receiving all 3 drugs, and 10 control subjects who had viral suppression while receiving triple-drug therapy. Drug susceptibility, determined by a phenotypic assay and genotypic evidence of resistance assessed by nucleotide sequencing of protease and reverse transcriptase, compared among the 3 patient groups. Indinavir resistance was not detected in the 9 subjects with viral rebound during indinavir monotherapy or in the 17 subjects with rebound during triple-drug therapy, despite plasma HIV RNA levels ranging from 10(2) to 10(5) copies/mL. In contrast, lamivudine resistance was detected by phenotypic assay in rebound isolates from 14 of 17 subjects receiving triple-drug therapy, and genotypic analyses showed changes at codon 184 of reverse transcriptase in these 14 isolates. Mean random plasma indinavir concentrations in the 2 groups with rebound were similar to those of a control group with sustained viral suppression, although levels below 50 ng/mL were more frequent in the triple-drug group than in the control group (P = .03). Loss of viral suppression may be due to suboptimal antiviral potency, and selection of a predominantly indinavir-resistant virus population may be delayed for months even in the presence of ongoing indinavir therapy. The results suggest possible value in assessing strategies using drug components of failing regimens evaluated with resistance testing.

Resistance, Fitness, Adherence, and Potency

ANEAR-UNIFORMLY FATAL CLINICAL SYNDROME, ACquired immunodeficiency syndrome (AIDS), has been transformed during the past 5 years into a treatable infectious disease. The availability of potent antiretroviral agents coincided with the ability to measure levels of circulating virus in vivo. When used in tandem, an understanding of human immunodeficiency virus (HIV) replication dynamics in vivo was made possible, forming the scientific basis for the use of combination antiretroviral therapy. However, the treatment of HIV infection remains far from perfect, and new issues arise with regularity. Critical to achieving optimal therapeutic outcomes is an understanding of treatment failure. Early clinical trials of protease inhibitor monotherapy suggested that the pathway to treatment failure was exclusively via drug resistance. Viral rebound was thought to reflect failure of all components of a regimen. Furthermore, it was assumed that the absence of resistance-conferring genotypic changes reflected patient nonadherence. In this issue of THE JOURNAL, articles by Descamps and colleagues and Havlir and colleagues question these assumptions in the context of 2 large clinical trials, Trilege and AIDS Clinical Trials Group 343. The inferior outcomes observed in patients randomly assigned to receive less intensive maintenance therapy have been recently published. In the articles in this issue, the authors seek to understand the findings. In the Trilege Study, a 3-month induction phase with zidovudine, lamivudine, and indinavir was followed by randomization to either zidovudine and lamivudine, zidovudine and indinavir, or continued triple-drug therapy if the level of HIV RNA in plasma was less than 500 copies/mL. The primary end point was virologic failure, defined by 2 consecutive plasma measurements above 500 copies/mL on 2 consecutive visits, 6 weeks apart. Fifty-eight (20.8%) of the 279 randomly assigned patients met this end point, 29 receiving zidovudine and lamivudine, 21 receiving zidovudine and indinavir, and 8 receiving triple therapy. Fifty-eight study patients with durable virologic suppression were carefully selected by investigators as case-controls. The results of genotypic studies revealed the presence of the lamivudine resistance-conferring M184V substitution in reverse transcriptase in nearly all patients treated with lamivudine. However, primary-resistance mutations associated with reduced susceptibility to indinavir did not emerge during combination therapy with zidovudine and indinavir or triple therapy. Similarly, zidovudine-associated resistanceconferring mutations were rare and when present were confined to changes at codons 41 and 70 of reverse transcriptase. Adherence as measured by pill counts revealed a statistically significantdifference inmedianadherenceratesbetween cases and controls for patients prescribed either zidovudine or indinavir during maintenance therapy. Furthermore, patients randomly assigned to receive zidovudine with indinavir only demonstrated statistically significant differences in adherence rates compared with controls. Plasma indinavir levelswere found tobe lower thanexpected in2groups, those failing triple therapy and those failing zidovudine and indinavir maintenance in association with a greater loss in antiviral efficacy. Indinavir levels tended to be in the expected range in those patients in the zidovudine and indinavir group in whom virologic failure was associated with a modest loss ofantiviralactivity.Ofnote,plasmaindinavir levelswereclearly higher in controls compared with cases in both the triple therapy and zidovudine and indinavir groups. In the AIDS Clinical Trials Group 343 study, after a 6-month induction with the same triple combination regimen as used in Trilege, patients with plasma HIV RNA levels below 200 copies/mL were randomly assigned to receive zidovudine and lamivudine, indinavir monotherapy, or continued triple therapy. Patients were followed up monthly and the study end point, virologic failure, was defined as a subsequent plasma HIV RNA level of 200 copies/mL or greater. Plasma indinavir levels and resistance testing by both genotypic assay and a novel recombinant phenotypic assay were performed retrospectively in 9 of 23

Human immunodeficiency virus type 1 population bottleneck during indinavir therapy causes a genetic drift in the env quasispecies.

The impact of emergence of genetic resistance, soon after the beginning of antiretroviral therapy, on the genotype of other viral loci not implicated in the development of resistance was studied in four human immunodeficiency type 1 (HIV-1)-infected patients subjected to indinavir monotherapy. Two patients were chosen because they showed no decrease in virus load during the study period and two were selected because they showed a rapid decline in plasma viraemia after the initiation of therapy and a virus rebound after 12 weeks of treatment. The evolution of virus sequences was analysed within the four infected patients by examining virus sequences spanning the protease and C2-V3 env genes by RT-PCR of plasma samples obtained at the beginning and after 12 weeks of therapy. PCR products from the two genomic regions from the two sample points per patient were cloned and 10-15 clones from each sample were sequenced. Genotypic indinavir resistance was present in the four patients after 12 weeks of therapy. The overall protease and C2-V3 env regions quasispecies diversity at time zero was higher than that after 12 weeks of therapy, but this difference was more significant in the two patients who showed a reduction in virus load soon after the initiation of treatment. C2-V3 env sequences indicated that changes during emergence of resistance to indinavir were only detected in the two patients who showed a drastic reduction in virus load. Thus, a temporal relationship was observed between the start of therapy, a drastic reduction in virus load and a drift in the HIV-1 env quasispecies.

Associations between amino acids in the evolution of HIV type 1 protease sequences under indinavir therapy.

Significant diversity exists in amino acid sequences encoding HIV-1 protease in individuals naive for protease inhibitors, which could influence the rate of evolution of resistance. High-level resistance to indinavir requires multiple substitutions among at least 11 amino acid sites, and no single substitution was observed in all of 29 resistant isolates obtained from patients on long-term indinavir monotherapy. We have analyzed the evolution of PR in these sequences. The divergence from the baseline amino acid sequence by week 24 was 4%, increasing more than 7% by week 60. The mean difference between sequences from different patients at baseline was 6% (3-9%), rising to 10% after 40 weeks (3-16%), although at all time points nonsynonymous substitutions were less frequent than synonymous nucleotide changes. Analysis of associations between variants at different amino acid sites using a mutual information statistic revealed four pairs of sites to be significantly associated. In three cases these associations included residue 82. Clusters of baseline and week 24 amino acid sequences identified by maximum parsimony did not correlate significantly with the IC95 to indinavir, although a weak correlation of baseline clusters with phenotype at the week 24 time point was suggested.

Indinavir: a review of its use in the management of HIV infection.

Indinavir is a protease inhibitor used in the treatment of patients with HIV infection. Combination antiretroviral therapy with indinavir plus 2 nucleoside reverse transcriptase inhibitors (NRTIs) is associated with greater reductions in viral load, greater increases in CD4+ cell counts, and reduced morbidity and mortality when compared with 2 NRTIs alone. In the landmark clinical trial ACTG 320, the rate of progression to AIDS or death (primary end-point) among zidovudine-experienced patients treated with indinavir, zidovudine and lamivudine was approximately half that of patients who received only zidovudine plus lamivudine (6 vs 11%; p < 0.001). The durability of an indinavir-containing regimen was demonstrated in Merck protocol 035, an ongoing trial in which a significant proportion of patients had sustained viral suppression for up to 3 years. Merck protocol 039, also an ongoing trial, showed a greater effect on surrogate markers of HIV disease progression with indinavir-based triple therapy than with zidovudine plus lamivudine or indinavir monotherapy in patients with advanced disease (median baseline CD4+ count 15 cells/microL). Numerous additional clinical trials have established the beneficial antiviral and immunological effects of indinavir in both antiretroviral-naive and -experienced patients with HIV infection. Indinavir is associated with various drug class-related adverse events, including gastrointestinal disturbances (e.g. nausea, diarrhoea), headache and asthenia/fatigue. A lipodystrophy syndrome has been commonly reported with indinavir and other protease inhibitors combined with NRTIs, but it has also been reported in many protease inhibitor-naive patients, and a definitive causal link has not been established between the syndrome and protease inhibitors. Nephrolithiasis may develop in about 9% of patients receiving indinavir but does not appear to be associated with other protease inhibitors; <0.5% of patients receiving indinavir discontinue the drug because of nephrolithiasis, which may be the extreme end of a continuum of crystal-related renal syndromes. Additional renal problems (e.g. nephropathy) have been reported in small numbers of patients receiving indinavir. In summary, indinavir is a protease inhibitor with well documented efficacy when used as part of combined therapy in patients with HIV infection. Both US and UK treatment guidelines continue to recommend protease inhibitor-based regimens including indinavir as a first-line option. Indinavir is being studied as a twice daily and once daily regimen with a low dosage of ritonavir as a way to alleviate tolerability, drug interaction and patient compliance/adherence issues. Indinavir-containing triple therapy has demonstrated positive effects not only on surrogate markers of disease progression, but also on clinical end-points of mortality and morbidity in patients with HIV disease. Protease inhibitors are a significant advance in the care of patients with HIV infection, and, in an era of evidence-based medicine, indinavir represents an important component of antiretroviral treatment strategies.