Richard T. D'Aquila

Immune control of HIV-1 after early treatment of acute infection

Virus-specific T-helper cells are considered critical for the control of chronic viral infections. Successful treatment of acute HIV-1 infection leads to augmentation of these responses, but whether this enhances immune control has not been determined. We administered one or two supervised treatment interruptions to eight subjects with treated acute infection, with the plan to restart therapy if viral load exceeded 5,000 copies of HIV-1 RNA per millilitre of plasma (the level at which therapy has been typically recommended) for three consecutive weeks, or 50,000 RNA copies per ml at one time. Here we show that, despite rebound in viraemia, all subjects were able to achieve at least a transient steady state off therapy with viral load below 5,000 RNA copies per ml. At present, five out of eight subjects remain off therapy with viral loads of less than 500 RNA copies per ml plasma after a median 6.5 months (range 5–8.7 months). We observed increased virus-specific cytotoxic T lymphocytes and maintained T-helper-cell responses in all. Our data indicate that functional immune responses can be augmented in a chronic viral infection, and provide rationale for immunotherapy in HIV-1 infection.

Antiretroviral Drug Resistance Testing in Adults Infected with Human Immunodeficiency Virus Type 1: 2003 Recommendations of an International AIDS Society-USA Panel

New information about the benefits and limitations of testing for resistance to human immunodeficiency virus (HIV) type 1 (HIV-1) drugs has emerged. The International AIDS Society-USA convened a panel of physicians and scientists with expertise in antiretroviral drug management, HIV-1 drug resistance, and patient care to provide updated recommendations for HIV-1 resistance testing. Published data and presentations at scientific conferences, as well as strength of the evidence, were considered. Properly used resistance testing can improve virological outcome among HIV-infected individuals. Resistance testing is recommended in cases of acute or recent HIV infection, for certain patients who have been infected as long as 2 years or more prior to initiating therapy, in cases of antiretroviral failure, and during pregnancy. Limitations of resistance testing remain, and more study is needed to refine optimal use and interpretation.

Standardized peripheral blood mononuclear cell culture assay for determination of drug susceptibilities of clinical human immunodeficiency virus type 1 isolates. The RV-43 Study Group, the AIDS Clinical Trials Group Virology Committee Resistance Working Group.

A standardized antiviral drug susceptibility assay for clinical human immunodeficiency virus type 1 (HIV-1) isolates has been developed for use in clinical trials. The protocol is a two-step procedure that first involves cocultivation of patient infected peripheral blood mononuclear cells (PBMC) with seronegative phytohemagglutinin-stimulated donor PBMC to obtain an HIV-1 stock. The virus stock is titrated for viral infectivity (50% tissue culture infective dose) by use of serial fourfold virus dilutions in donor PBMC. A standardized inoculum of 1,000 50% tissue culture infective doses per 10(6) cells is used in the second step of the procedure to acutely infect seronegative donor PBMC in a 7-day microtiter plate assay with triplicate wells containing zidovudine (ZDV) concentrations ranging from 0 to 5.0 microM. The ZDV 50% inhibitory concentrations (IC50) for reference ZDV-susceptible and ZDV-resistant HIV-1 isolates ranged from 0.002 to 0.113 microM and from 0.15 to > 5.0 microM, respectively. Use of this consensus protocol reduced interlaboratory variability for ZDV IC50 determinations with reference HIV-1 isolates. Among eight laboratories, the coefficient of variation ranged from 0.85 to 1.25 with different PBMC protocols and was reduced to 0.39 to 0.98 with the standardized assay. Among the clinical HIV-1 isolates assayed by the standardized drug susceptibility assay, the median ZDV IC50 increased gradually with more ZDV therapy. This protocol provides an efficient and reproducible means to assess the in vitro susceptibility to antiretroviral agents of virtually all clinical HIV-1 isolates.

Nevirapine, Zidovudine, and Didanosine Compared with Zidovudine and Didanosine in Patients with HIV-1 Infection: A Randomized, Double-Blind, Placebo-Controlled Trial

Strategies to improve the clinical benefit of therapy for human immunodeficiency virus type 1 (HIV-1) infection have included combining antiretroviral drugs to increase antiviral activity. Some combinations of two HIV-1 reverse transcriptase inhibitors synergistically inhibit HIV-1 replication in vitro; these combinations include that of the nucleoside analogues zidovudine and didanosine [1, 2] and that of zidovudine and the non-nucleoside reverse transcriptase inhibitor nevirapine [3-5]. The combination of zidovudine and didanosine also enhances antiviral activity in vivo [6-8] and delays disease progression [9-11] more effectively than zidovudine monotherapy. One way to further suppress HIV-1 replication may be to add a third antiretroviral agent to a combination regimen. Certain three-drug regimens, including the combination of zidovudine, didanosine, and a non-nucleoside reverse transcriptase inhibitor such as nevirapine, inhibit wild-type HIV-1 in vitro more effectively than one or two of these drugs [12-15]. Nevirapine (400 mg/d), as monotherapy or in combination with zidovudine or zidovudine and didanosine, has resulted in sustained antiviral effect in vivo, although nevirapine-resistant virus was isolated from patients in open-label trials [16-18] and from in vitro selection experiments [19, 20]. No adverse drugdrug interactions were noted in patients receiving the triple combination of nevirapine, zidovudine, and didanosine over a short period [18]. Therefore, we tested whether this triple combination would improve immunologic and virologic effects in vivo by comparing it with a combination of zidovudine and didanosine in a 48-week phase II, randomized clinical trial in adults with HIV-1 disease who had previously received prolonged nucleoside therapy. Methods Study Design and Treatment Regimens This multicenter, randomized trial was AIDS (acquired immunodeficiency syndrome) Clinical Trials Group Protocol 241. Patients received either nevirapine with open-label zidovudine and didanosine (the triple combination) or placebo with open-label zidovudine and didanosine (the double combination). The placebo tablets were identical in appearance to the active nevirapine tablets. Patients and investigators were blinded to treatment assignments; a permuted-blocks design within each institution was used for randomization. Patients were stratified according to their screening CD4 cell counts: 50 or fewer cells/mm3, 51 to 200 cells/mm3, and 201 to 350 cells/mm3. Zidovudine (Retrovir, provided by Glaxo Wellcome, Research Triangle Park, North Carolina) was given orally as two 100-mg tablets three times a day. Didanosine (Videx, provided by Bristol-Myers Squibb, Wallingford, Connecticut) was given orally as two 100-mg chewable and dispersable tablets twice daily for patients weighing at least 60 kg and as one 100-mg tablet and one 25-mg tablet twice daily for patients weighing less than 60 kg. Nevirapine (Viramune, provided by Boehringer-Ingelheim Pharmaceuticals, Ridgefield, Connecticut) was given orally as a 200-mg tablet once daily for the first 2 weeks and thereafter as one 200-mg tablet twice daily. This schedule was used because a lead-in period with a lower daily dose of nevirapine had decreased the frequency of nevirapine-related rashes in earlier studies [17]. Data were gathered, analyzed, and interpreted independently of the pharmaceutical companies that provided the drugs, according to the standard operating procedures of the AIDS Clinical Trials Group. Study Sample Adults who had documented HIV-1 infection and CD4 cell counts of 350 cells/mm3 or less within 30 days of randomization and who had had previous nucleoside therapy with zidovudine, didanosine, or zalcitabine for at least 6 months were eligible. Eligibility criteria also included a Karnofsky performance status score of at least 70% within 30 days of randomization, a hemoglobin concentration of at least 91 g/L or more for men and 88 g/L for women, a neutrophil count of 1000 cells/mm3 or more, a platelet count of 75 000 109/L or more, a serum creatinine concentration no more than 1.5 times the upper limit of normal, serum concentrations of alanine aminotransferase and aspartate aminotransferase no more than 3 times the upper limit of normal, and a serum amylase concentration no more than 1.5 times the upper limit of normal (unless the serum lipase concentration was 1.5 times the upper limit of normal or less). Patients were excluded if they were intolerant of zidovudine (at 500 or 600 mg/d) or didanosine (at 400 mg/d for tablets and 500 mg/d for sachets), had moderate peripheral neuropathy ( grade 2 according to the National Institute of Allergy and Infectious Diseases [NIAID] Division of AIDS Table for Grading Adult Adverse Experiences), had pancreatitis, or had previously used non-nucleoside reverse transcriptase inhibitors. The study was approved by the institutional review board at each institution, and all participants gave written informed consent. Concomitant therapy with other antiretroviral agents, foscarnet, biological response modifiers, erythromycin, clavulanate-containing antibacterial agents, warfarin, phenytoin, or phenobarbital was not permitted. Prophylaxis for Pneumocystis carinii pneumonia was required for all patients with a CD4 cell count of 200 cells/mm3 or less or a history of P. carinii pneumonia. Maintenance therapies for other opportunistic infections were permitted. Patients could continue their prestudy antiretroviral nucleoside therapy until the day on which therapy with the study medications was started. Management of Toxicities Toxicity was graded according to the NIAID Division of AIDS Table for Grading Adult Adverse Experiences. Modification, interruption, or discontinuation of therapy with the study medications was done for many adverse effects one drug at a time, starting with the drug most likely to have caused the toxicity. Zidovudine was the first drug to be modified for severe anemia, myositis, neutropenia, fatigue, and headache; didanosine was the first to be modified for severe nausea, vomiting, diarrhea, constipation, hyperamylasemia, fasting hypertriglyceridemia, hyperuricemia, and peripheral neuropathy; and nevirapine or placebo was the first to be modified for rash, severe thrombocytopenia, and adverse neuropsychological effects. Therapy with all study medications was interrupted for other severe adverse experiences, including hepatotoxicity. Therapy with a medication or medications was interrupted until the toxicity resolved and was then resumed at half the original dose, except that didanosine therapy was permanently discontinued for pancreatitis and nevirapine or placebo use was permanently discontinued for severe rash. Recurrent or persistent adverse effects led to the permanent discontinuation of either zidovudine or didanosine therapy. All study treatment was stopped if therapy with nevirapine or placebo or any two study medications was permanently discontinued. Patient Evaluation The schedule of patient evaluations is shown in Table 1. Standardized assays were used for CD4 cell counts [21, 22] and serum p24 antigen levels (AIDS Clinical Trials Group Virus Quality Assurance Laboratory reference standards were used with enzyme-linked immunosorbent assay kits from either Coulter [Hialeah, Florida], Abbott Laboratories [North Chicago, Illinois], or DuPont [Wilmington, Delaware]). Plasma nevirapine concentrations were measured at Boehringer-Ingelheim Pharmaceuticals by a validated high-performance liquid chromatography assay [23]. Additional virologic and pharmacologic assessments were done on specimens from all patients enrolled at 8 of the 16 study sites (n = 198). The HIV-1 infectivity titers in peripheral blood mononuclear cells were calculated by using maximum likelihood estimation from a standardized HIV-1 quantitative microculture assay in quality-assured AIDS Clinical Trials Group virology laboratories [24]. Plasma HIV-1 RNA levels were measured by quantitative reverse transcriptase polymerase chain reaction (Roche Molecular Systems, Alameda, California, and Branchburg, New Jersey) done in batch at Roche Biomedical Laboratories (Research Triangle Park, North Carolina) [25]. Plasma zidovudine and didanosine concentrations were measured at the University of Alabama at Birmingham by validated radioimmunoassays [26, 27]. Viruses isolated at entry were classified as either syncytium-inducing or non-syncytium-inducing according to the results of an MT-2 cell assay [28]. Table 1. Schedule of Patient Evaluations Primary outcome measures were differences between treatment groups at the end of the study in mean absolute CD4 cell count, HIV-1 infectivity titer in peripheral blood mononuclear cells, serum p24 antigen level, and time to HIV-1 disease progression or death. Secondary outcome measures were differences between treatment groups at the end of the study in the mean percentage of T lymphocytes that were CD4 cells and in plasma HIV-1 RNA levels. These measurements at the end of the study were the geometric means of measurements obtained between weeks 40 and 48, expressed as the change from the geometric mean of the two measurements made before treatment. Standardized area-under-the-curve analyses of immunologic and virologic measures over time were additional secondary outcome measures used to assess differences between treatments that included short-term effects. Progression of HIV-1 disease was defined as the development of a new AIDS-defining clinical event according to the Centers for Disease Control and Prevention case definition (except that we excluded CD4 cell count as a disease progression criterion) [29], a newly diagnosed deep-seated bacterial infection or bacteremia unrelated to the use of injection drugs or an intravascular catheter, 1 month or more of symptomatic microsporidiosis, or the recurrence of either P. carinii pneumonia or central nervous system toxoplasmosis. All disease prog

Monitoring plasma HIV-1 RNA levels in addition to CD4+ lymphocyte count improves assessment of antiretroviral therapeutic response

The duration of disease-free survival after infection with human immunodeficiency virus type 1 (HIV-1) varies considerably during antiretroviral therapy. Patients with similar CD4+ lymphocyte counts progress at different rates when they are given the same antiretroviral therapy. Better prediction of risk for progression and its association with viral suppression may help improve antiretroviral management for individual patients and speed the development of new drugs. Higher plasma HIV-1 RNA levels are associated with poorer clinical status and lower CD4+ lymphocyte counts [1-3] and predict subsequent outcome [4-11]. The biological variability of plasma HIV-1 RNA levels in patients receiving stable therapeutic regimens must be quantified to define the magnitude of an antiviral effect that can be reliably detected after antiretroviral treatment is initiated. Determination of infectious HIV-1 titers in mononuclear cells of peripheral blood by quantitative microculture [12, 13] or syncytium-inducing phenotype of an HIV-1 isolate may provide information that is different from or complementary to the information gleaned from measuring plasma HIV-1 RNA levels [14-16]. However, studies have not yet conclusively determined whether measurements of CD4+ lymphocytes in conjunction with any or all of these virological variables should be recommended to optimize prediction or guide antiretroviral treatment more effectively. In this report, we quantify the relative roles of CD4+ lymphocyte counts, plasma HIV-1 RNA levels, infectious HIV-1 titers in mononuclear cells of peripheral blood, and the syncytium-inducing viral phenotype as predictors of disease progression during a clinical trial of combination therapy [17]. Our approach was to assess the value of plasma HIV-1 RNA levels and CD4+ lymphocyte count, both of which are readily available to clinicians, and then to assess the additional value of the infectious HIV-1 titer in mononuclear cells of peripheral blood and the syncytium-inducing viral phenotype. We also quantify the variability of plasma HIV-1 RNA levels. Our results suggest guidelines for using these measures in clinical practice for predicting the effectiveness of antiretroviral therapy over 1 year. Methods Study Design We prospectively evaluated virological, immunologic, and clinical data from patients who participated in the intensive virology substudy of ACTG (AIDS Clinical Trials Group) Protocol 241; ACTG Protocol 241 was a multicenter, randomized, double-blind, placebo-controlled trial of 398 patients receiving nevirapine, zidovudine, and didanosine compared with zidovudine and didanosine [17]. All patients at 8 of the 16 AIDS Clinical Trials Units who participated in the main study were enrolled in the substudy (n = 198). For 48 weeks, all 198 patients received open-label zidovudine (600 mg/d) and didanosine (400 mg/d for patients weighing 60 kg and 250 mg/d for patients weighing <60 kg). One hundred of the substudy patients were randomly assigned to receive nevirapine (200 mg/d for the first 2 weeks and 400 mg/d thereafter), and 98 were assigned to receive matching placebo. Participants gave written informed consent, and the protocol was approved by the institutional review board at each participating AIDS Clinical Trials Unit. The study was funded by the ACTG of the National Institute of Allergy and Infectious Diseases; supplemental funding for virology was provided by Boehringer Ingelheim Pharmaceuticals (Ridgefield, Connecticut). Study drugs were provided by Glaxo Wellcome (Research Triangle Park, North Carolina), Bristol-Myers Squibb (Princeton, New Jersey), and Boehringer Ingelheim Pharmaceuticals. However, all data were gathered by members of the ACTG and were analyzed and interpreted by the authors, who had sole responsibility for the decision to submit the manuscript for publication. Evaluation of Patients Stable therapy at baseline was defined as the absence of reported change in antiretroviral therapy from 30 days before the preentry visit until the entry visit. All patients were followed prospectively for progression of HIV-related disease. Progression was defined as the development of a new acquired immunodeficiency syndrome (AIDS)-defining event [18]; a newly diagnosed, deep-seated bacterial infection or bacteremia that was not related to injection drug use or an intravascular catheter; pulmonary or extrapulmonary tuberculosis; recurrent Pneumocystis carinii pneumonia; recurrent toxoplasmosis of the central nervous system; or death. Reports of disease progression were reviewed by the study chair; only events that could be confirmed were used in the analysis. We measured CD4+ lymphocyte counts, plasma HIV-1 RNA levels, and infectious HIV-1 titers in mononuclear cells of peripheral blood at the preentry visit (within 14 days of starting study treatment), at the entry visit (before starting study treatment and at least 72 hours after the preentry visit), and at the visits 8 and 48 weeks after the start of study treatment. Specimens could be obtained at any time of day. We used the geometric mean of preentry and entry measurements as the baseline value for each variable. The presence of the syncytium-inducing viral phenotype was determined at the entry visit. Standardized assays were used to determine CD4+ lymphocyte counts [19, 20], infectious HIV-1 titer in mononuclear cells of peripheral blood (in infectious units per million cells) using real-time testing [13, 21], and syncytium-inducing viral phenotype of a virus isolated from mononuclear cells of peripheral blood using MT-2 cells [22]. Plasma samples were frozen at 70C; HIV-1 RNA levels were measured by quantitative reverse transcription polymerase chain reaction assay (Roche Molecular Systems, Alameda, California, and Branchburg, New Jersey) [23]. The lower limit of detection for this assay was 200 HIV-1 RNA copies/mL. Levels of HIV-1 RNA in plasma samples collected from the same patient at the preentry, entry, week 8, and week 48 visits were determined in a single laboratory assay. Statistical Analysis Analysis of plasma HIV-1 RNA levels and infectious HIV-1 titers in mononuclear cells of peripheral blood was done after log10 transformation. Plasma levels of HIV-1 RNA that were below the detectable limit were assigned the value of 200 copies/mL. Infectious HIV-1 titers in mononuclear cells of peripheral blood outside the measurable range (0.22 to 7493 infectious units per million cells) were assigned the value of 0.22 infectious units per million cells if they were below the range and 7493 infectious units per million cells if they were above the range. Linear regression analysis [24] was used to compare the mean plasma HIV-1 RNA levels, infectious HIV-1 titers in mononuclear cells of peripheral blood, and CD4+ lymphocyte counts according to patient characteristics at baseline and to assess factors associated with the long-term change (from baseline to week 48) in CD4+ lymphocyte counts. Logistic regression analysis [25] was used to assess the association at baseline of the percentage of patients who had AIDS with virological measures and CD4+ lymphocyte counts. The intrapatient SD of plasma HIV-1 RNA levels was estimated using the method of moments for variance components [26]. Spearman correlation coefficients were used to assess the association between preentry and entry measurements. Proportional hazards models [27] were used to assess the association between the risk for disease progression or death and baseline levels and early changes (from baseline to week 8) in plasma HIV-1 RNA levels, infectious HIV-1 titers in mononuclear cells of peripheral blood, and log-transformed CD4+ lymphocyte counts as well as baseline syncytium-inducing viral phenotype. These models were stratified by study treatment to control for any differential effects of the two study regimens. Results Patient Characteristics at Study Entry The mean CD4+ lymphocyte count of the 198 patients before treatment was 145 cells/mm3 (range, 1 to 443 cells/mm3). Patients were a median of 39 years of age, predominantly male (81%), predominantly white (76%), and predominantly free of a previous AIDS-defining diagnosis (86%). All but 3 patients had taken zidovudine before study entry, 44% had taken didanosine, and 35% had taken zalcitabine. The median duration of cumulative previous nucleoside therapy was 25 months, and 34% of patients had received therapy for longer than 36 months. Virological Measures at Baseline by Patient Characteristics Table 1 shows the mean plasma HIV-1 RNA levels, infectious HIV-1 titers in mononuclear cells of peripheral blood, and CD4+ lymphocyte counts at baseline for patients stratified by characteristics that were significantly associated with viral load. We also assessed the associations with age, sex, racial or ethnic group, self-reported homosexuality, and duration of previous nucleoside therapy, but these associations were not significant. Table 1. Plasma HIV-1 RNA Level, Infectious HIV-1 Titer in Mononuclear Cells of Peripheral Blood, and CD4+ Lymphocyte Count at Baseline* Patients with a history of AIDS had a significantly higher mean baseline level of HIV-1 RNA in plasma and a significantly lower mean CD4+ lymphocyte count than did those without such a history (Table 1). More patients with a history of AIDS than those without had baseline HIV-1 isolates with the syncytium-inducing viral phenotype (58% compared with 36%; P = 0.015). However, in a multivariate analysis, only the CD4+ lymphocyte count at baseline was significantly associated with a history of AIDS. Thus, disease status at baseline was explained by CD4+ lymphocyte counts and not by any of the virological measures that were considered. Variability of Virological Measures in Patients Receiving Stable Treatment Variation in plasma HIV-1 RNA levels was evaluated by comparing the preentry and entry measures from the 167 patients who reported no changes in treatment from 30 days

Genotypic Correlates of Phenotypic Resistance to Efavirenz in Virus Isolates from Patients Failing Nonnucleoside Reverse Transcriptase Inhibitor Therapy

ABSTRACT Efavirenz (also known as DMP 266 or SUSTIVA) is a potent nonnucleoside inhibitor of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) activity and of HIV-1 replication in vitro and in vivo. Most patients on efavirenz-containing regimens have sustained antiviral responses; however, rebounds in plasma viral load have been observed in some patients in association with the emergence of mutant strains of HIV-1. Virus isolates from the peripheral blood mononuclear cells (PBMCs) of patients with such treatment failures, as well as recombinant viruses incorporating viral sequences derived from patient plasma, show reduced in vitro susceptibility to efavirenz in association with mutations in the RT gene encoding K103N, Y188L, or G190S/E substitutions. Patterns of RT gene mutations and in vitro susceptibility were similar in plasma virus and in viruses isolated from PBMCs. Variant strains of HIV-1 constructed by site-directed mutagenesis confirmed the role of K103N, G190S, and Y188L substitutions in reduced susceptibility to efavirenz. Further, certain secondary mutations (V106I, V108I, Y181C, Y188H, P225H, and F227L) conferred little resistance to efavirenz as single mutations but enhanced the level of resistance of viruses carrying these mutations in combination with K103N or Y188L. Viruses with K103N or Y188L mutations, regardless of the initial selecting nonnucleoside RT inhibitor (NNRTI), exhibited cross-resistance to all of the presently available NNRTIs (efavirenz, nevirapine, and delavirdine). Some virus isolates from nevirapine or delavirdine treatment failures that lacked K103N or Y188L mutations remained susceptible to efavirenz in vitro, although the clinical significance of this finding is presently unclear.

Use of Changes in Plasma Levels of Human Immunodeficiency Virus Type 1 RNA to Assess the Clinical Benefit of Antiretroviral Therapy

Data from 1330 human immunodeficiency virus type 1 (HIV-1)-infected patients enrolled in seven antiretroviral treatment trials were analyzed to characterize the clinical benefit of treatment-mediated reductions in plasma HIV-1 RNA levels. The risk of a new AIDS-defining event or death was reduced proportionally to the magnitude of the reduction of the HIV-1 RNA level during the first 6 months of therapy. Pretherapy HIV-1 RNA levels were prognostic independently of on-therapy levels. In addition, the reduction in risk associated with any given reduction of the level of HIV-1 RNA did not vary by pretherapy level. Having either a reduction in HIV-1 RNA level or an increase in CD4+ lymphocyte count, or both, was associated with a delay in clinical disease progression. This indicates that patient prognosis should be assessed using both HIV-1 RNA and CD4+ lymphocyte responses to therapy.

Virological and immunological effects of treatment interruptions in HIV-1 infected patients with treatment failure.

ObjectiveTo analyse the immunological and virological effects of treatment interruptions in HIV-1-infected patients with treatment failure and multidrug-resistant virus. MethodsDrug susceptibility was assessed using Antivirogram and genotypic analysis was based on population and clonal sequencing for 48 patients who had interrupted treatment (⩾ 2 months). ResultsTreatment interruption resulted in viral load increases (mean 0.7 log10 copies/ml;P = 0.0001) and CD4 cell count decreases (mean 89 × 106 cells/l;P = 0.0001). A complete shift to wild-type virus at the phenotypic, genotypic and clonal level was observed in 28/45 patients. These patients differed from those that did not show a shift to wild type in baseline CD4 cell counts (192 versus 59 × 106 cells/l;P = 0.007) and in the relationship between baseline viral load and CD4 cell count (no correlation versus a significant negative correlation;P = 0.008). Response to re-initiation of treatment fell with increasing viral load [relative hazard (RH) 0.33;P = 0.001] and with increasing total number of drugs with reduced susceptibility (RH 0.51;P = 0.0003); it improved with the number of new drugs received (RH 2.12;P = 0.0002) and a shift to wild type (RH 5.22, P = 0.006). ConclusionsChanges in surrogate markers suggest that treatment provided benefit in spite of virological failure and resistant virus. Although patients with a shift to wild-type virus responded better in the short term to treatment re-initiation, the long-term effects are not known and the risk of immune deterioration needs to be carefully considered.

Pre-existing Minority Drug-Resistant HIV-1 Variants, Adherence, and Risk of Antiretroviral Treatment Failure

BACKGROUND The clinical relevance of detecting minority drug-resistant human immunodeficiency virus type 1 (HIV-1) variants is uncertain. METHODS To determine the effect of pre-existing minority nonnucleoside reverse-transcriptase inhibitor (NNRTI)-resistant variants on the risk of virologic failure, we reanalyzed a case-cohort substudy of efavirenz recipients in AIDS Clinical Trials Group protocol A5095. Minority K103N or Y181C populations were determined by allele-specific polymerase chain reaction in subjects without NNRTI resistance by population sequencing. Weighted Cox proportional hazards models adjusted for recent treatment adherence estimated the relative risk of virologic failure in the presence of NNRTI-resistant minority variants. RESULTS The evaluable case-cohort sample included 195 subjects from the randomly selected subcohort (51 with virologic failure, 144 without virologic failure), plus 127 of the remaining subjects who experienced virologic failure. Presence of minority K103N or Y181C mutations, or both, was detected in 8 (4.4%), 54 (29.5%), and 11 (6%), respectively, of 183 evaluable subjects in the random subcohort. Detection of minority Y181C mutants was associated with an increased risk of virologic failure in the setting of recent treatment adherence (hazard ratio, 3.45 [95% confidence interval, 1.90-6.26]) but not in nonadherent subjects (hazard ratio, 1.39 [95% confidence interval, 0.58-3.29]). Of note, 70% of subjects with minority Y181C variants achieved long-term viral suppression. CONCLUSIONS In adherent patients, pre-existing minority Y181C mutants more than tripled the risk of virologic failure of first-line efavirenz-based antiretroviral therapy. CLINICAL TRIALS REGISTRATION NCT00013520.

Multiple sites in HIV-1 reverse transcriptase associated with virological response to combination therapy.

ObjectiveTo determine whether analysis of sequence variation in reverse transcriptase at baseline can explain differences in response to combination antiretroviral therapy. MethodsAmino acid sequences of reverse transcriptase obtained from baseline isolates from 55 patients included in a trial of zidovudine and didanosine versus zidovudine/didanosine/nevirapine (ACTG241) were analysed. Simple and multiple linear regression were used to determine the relationship between numbers and identity of mutations at baseline and virological response after 8 and 48 weeks. ResultsNumbers of baseline zidovudine resistance mutations were predictive of short-term response (week 8). Amino acid identity at position 215 explained > 20% of the variation in response at week 8, but less at week 48. Multiple regression identified the combinations: 215 + 44 and 41 + 202, each of which explained about 30% of the variation in week 8 response. A model incorporating amino acids 214 + 215 + 60 + 202 + baseline viral load explained > 40% of the variation in response at week 48. Unexpectedly, the mutant combination 60I + 215Y/F responded threefold better than 60V + 215Y/F over 48 weeks. ConclusionsUse of clinical data to analyse virological response to combination therapy has revealed effects of baseline amino acid mutations at sites not previously identified as being important in antiretroviral resistance. Predictors of long-term responses were different from those involved in the short term and may require more complex analysis.