Nicola Gianotti

Lamivudine monotherapy in HIV-1-infected patients harbouring a lamivudine-resistant virus: a randomized pilot study (E-184V study).

Objective:We compared the immunological and clinical outcomes of lamivudine monotherapy and complete therapy interruption in the treatment of HIV-1-infected patients harbouring lamivudine-resistant virus. Methods:This 48-week, open-label pilot study randomly assigned HIV-infected patients receiving lamivudine-containing HAART and harbouring the M184V mutation to monotherapy with lamivudine 300 mg once daily (lamivudine group) or the discontinuation of all antiretroviral drugs (TI group). The primary endpoint was the occurrence of immunological or clinical failure; immunological failure was defined as the first report of a CD4 T-cell count less than 350 cells/μl, and clinical failure as the occurrence of a Centers for Disease Control and Prevention grade B or C event. The data were analysed on the basis of the intention-to-treat principle. Results:By week 48, 20 of 29 patients in the TI group (69%; 95% CI 51–83%) and 12 of 29 in the lamivudine group (41%; 95% CI 26–59%) had discontinued the study because of immunological or clinical failure, which was significantly delayed in the lamivudine group (P = 0.018). Only patients in the TI group (6/29, 20.7%) experienced grade 3–4 clinical adverse events at least possibly related to HIV-1 (P = 0.02). The mean decline in CD4 cell percentage, viral rebound and recovery of HIV-1 replication capacity were significantly lower in the lamivudine group. The 24-week virological and immunological response after therapy resumption in patients who prematurely discontinued the study was similar in the two groups. Conclusion:In HIV-1-infected patients harbouring a lamivudine-resistant virus, lamivudine monotherapy may lead to a better immunological and clinical outcome than complete therapy interruption.

Nested polymerase chain reaction for diagnosis and monitoring treatment response in AIDS patients with tuberculous meningitis.

ObjectiveTo investigate the usefulness of polymerase chain reaction (PCR) from cerebrospinal fluid (CSF) for rapid diagnosis, and assessing treatment response of tuberculous meningitis (IBM) in AIDS patients. PatientsForty-four CSF samples from 10 patients with TBM confirmed by autopsy or by a culture of CSF (41 samples), and from two patients with highly probable TBM (three samples) were analysed. CSF specimens were collected before, and during standard antituberculous treatment. CSF samples from 24 AIDS patients with autopsy evidence of other neurologic diseases were studied as controls. MethodsA nested PCR amplifying a 123 base-pair fragment of the IS6770 sequence was developed. Heating to 95±C for 15 min was used for pre-PCR treatment of samples. ResultsDetection limit was 102 colony-forming units per ml or 10fg purified Mycobacterium tuberculosis DNA. M. tuberculosis DNA was detected in CSF from all the 12 confirmed or highly probable TBM cases. CSF was positive by nested PCR in 17 of 17 (100%), and 18 of 27 (67%) samples collected before, and during therapy, respectively. Clinical, and microbiological follow-up ≥2 weeks was available for seven patients. PCR-positive CSF converted to M. tuberculosis DNA negative in four patients that showed improvement during treatment, but it remained positive in three patients who died of disseminated tuberculosis. All the CSF samples from the non-TBM controls were negative by nested PCR. ConclusionsNested PCR for detection of M. tuberculosis DNA is specific for diagnosis of TBM, and more sensitive than conventional bacteriology. Moreover, nested PCR could be a useful method for assessing treatment response in AIDS patients with TBM.

Dynamic patterns of human immunodeficiency virus type 1 integrase gene evolution in patients failing raltegravir-based salvage therapies.

Objective:Evaluate HIV-1 subtype B integrase gene evolution in patients failing raltegravir (RAL)-based savage regimens by clonal analysis of the replicating viral quasispecies. Design:Seven triple class failure HIV-1 (subtype B)-infected patients, followed at San Raffaele Hospital and enrolled in the RAL Expanded Access Program (MK0518-023), were evaluated. Patients were followed up for 24–48 weeks and due to the absence of other active drugs, RAL was maintained in their regimens even if resistance mutations were detected. Methods:Immunologic and virologic parameters were recorded every 4 weeks, and amplification and clonal analysis of viral populations were performed at baseline and every 4–12 weeks in all patients. Results:Resistance to RAL appeared initially associated with selection of single variants (Y143R, Q148R N155H) in the majority of patients; however, in three patients, complex patterns of viral mutations were observed. The clonal analysis of viral quasispecies allowed to describe the evolution of each viral population and the progressive accumulation of RAL resistance-associated mutations and polymorphisms associated with therapy failure. Conclusion:The complex patterns of resistance mutations observed, including novel variants evolved under continuous RAL pressure, suggesting that they are the result of the equilibrium between drug resistance and enzyme function. Despite the efficacy of this compound, our data discourage its use in a functional monotherapy and maintaining RAL even in presence of RAL resistance-associated mutations may lead to the progressive formation of viral reservoirs with multiple integrase inhibitor-resistant variants that may limit the future efficacy of other integrase inhibitors due to cross-resistance.

Raltegravir, maraviroc, etravirine: an effective protease inhibitor and nucleoside reverse transcriptase inhibitor-sparing regimen for salvage therapy in HIV-infected patients with triple-class experience.

We prospectively evaluated 28 triple-class experienced HIV-1-infected patients harbouring R5 virus, who received maraviroc, raltegravir and etravirine. By on-treatment analysis, 26 (92%) had less than 50 copies HIV-RNA/ml at week 48. The median (interquartile range) 48-week increase in CD4+ cell counts was 267 (136–355) cells/μl. Three serious adverse events occurred: one recurrence of mycobacterial spondylodiscitis, one anal cancer, one Hodgkin lymphoma. Although long-term safety needs further study, this protease inhibitor and nucleoside analogue-sparing regimen showed sustained efficacy.

Efficacy of Low-Dose Intermittent Subcutaneous Interleukin (IL)–2 in Antiviral Drug–Experienced Human Immunodeficiency Virus–Infected Persons with Detectable Virus Load: A Controlled Study of 3 IL-2 Regimens with Antiviral Drug Therapy

To evaluate the safety and efficacy of 3 regimens of intermittent subcutaneous (sc) interleukin (IL)--2 in a phase 2 study, 61 antiviral drug-experienced human immunodeficiency virus (HIV)--positive patients were randomly assigned to one of the following study arms: antiretroviral therapy (ART) plus IL-2 (12 million IU [MIU] by continuous intravenous infusion, followed by 7.5 MIU twice a day, sc, every 8 weeks); ART plus IL-2 (7.5 MIU twice a day, sc, every 8 weeks); ART plus IL-2 (3 MIU twice a day, sc, every 4 weeks); or ART alone. A significant increase of circulating CD4 cells was observed in IL-2--treated subjects, compared with those given ART alone. Low doses of IL-2 were better tolerated. Despite the incomplete suppression of viral replication, IL-2 with ART did not increase either plasma viremia or cell-associated HIV DNA levels. Low doses of intermittent sc IL-2 induced a stable increase of peripheral CD4 cells that was indistinguishable from those associated with higher, less well-tolerated doses of IL-2.

Prevalence of transmitted HIV-1 drug resistance in HIV-1 infected patients in Italy: evolution over 12 years and predictors

OBJECTIVES Transmitted HIV-1 drug resistance (TDR) can reduce the efficacy of first-line antiretroviral therapy. PATIENTS AND METHODS A retrospective analysis was performed to assess the prevalence and correlates of TDR in Italy over time. TDR was defined as the presence of at least one of the mutations present in the surveillance drug resistance mutation (SDRM) list. RESULTS Among 1690 antiretroviral therapy-naive patients, the most frequent HIV subtypes were B (78.8%), CRF02_AG (5.6%) and C (3.6%). Overall, TDR was 15%. TDR was 17.3% in subtype B and 7.0% in non-B carriers (P < 0.001). TDR showed a slight, although not significant, decline (from 16.3% in 1996-2001 to 13.4% in 2006-07, P = 0.15); TDR declined for nucleoside reverse transcriptase inhibitors (from 13.1% to 8.2%, P = 0.003) but remained stable for protease inhibitors (from 3.7% to 2.5%, P = 0.12) and non-nucleoside reverse transcriptase inhibitors (from 3.7% to 5.8%). TDR to any drug was stable in B subtype and showed a decline trend in non-B. In multivariable analysis, F1 subtype or any non-B subtype, compared with B subtype, and higher HIV RNA were independent predictors of reduced odds of TDR. CONCLUSIONS Prevalence of TDR to nucleoside reverse transcriptase inhibitors seems to have declined in Italy over time. Increased prevalence of non-B subtypes partially justifies this phenomenon.

In vitro phenotypes to elvitegravir and dolutegravir in primary macrophages and lymphocytes of clonal recombinant viral variants selected in patients failing raltegravir

OBJECTIVES The cross-resistance profiles of elvitegravir and dolutegravir on raltegravir-resistant variants is still controversial or not available in macrophages and lack extensive evaluations on wide panels of clonal variants. Thus, a complete evaluation in parallel with all currently available integrase inhibitors (INIs) was performed. METHODS The integrase coding region was RT-PCR-amplified from patient-derived plasma samples and cloned into an HIV-1 molecular clone lacking the integrase region. Twenty recombinant viruses bearing mutations to all primary pathways of resistance to raltegravir were phenotypically evaluated with each integrase inhibitor in freshly purified CD4+ T cells or monocyte-derived macrophages. RESULTS Y143R single mutants conferred a higher level of raltegravir resistance in macrophages [fold change (FC) 47.7-60.24] compared with CD4+ T cells (FC 9.55-11.56). All other combinations had similar effects on viral susceptibility to raltegravir in both cell types. Elvitegravir displayed a similar behaviour both in lymphocytes and macrophages with all the tested patterns. When compared with raltegravir, none to modest increases in resistance were observed for the Y143R/C pathways. Dolutegravir maintained its activity and cross-resistance profile in macrophages. Only Q148H/R variants had a reduced level of susceptibility (FC 5.48-18.64). No variations were observed for the Y143R/C (+/-T97A) or N155H variants. CONCLUSIONS All INIs showed comparable antiretroviral activity in both cell types even if single mutations were associated with a different level of susceptibility in vitro to raltegravir and elvitegravir in macrophages. In particular, dolutegravir was capable of inhibiting with similar potency infection of raltegravir-resistant variants with Y143 or N155 pathways in both HIV-1 major cell reservoirs.

Expression levels of MDR1, MRP1, MRP4, and MRP5 in peripheral blood mononuclear cells from HIV infected patients failing antiretroviral therapy

The aim of the study was to evaluate the mRNA expression of four relevant ABC‐transporter genes [MDR1 (P‐glycoprotein; Pgp), MRP1, MRP4, and MRP5] in HIV‐positive individuals failing treatment and analyze the association between the levels of their expression and viral load, CD4 cell count, and therapeutic history. Ninety‐eight HIV‐positive samples and 20 samples from healthy donors were analyzed, retrospectively. Peripheral blood mononuclear cells (PBMCs) from HIV1‐positive individuals were collected at the time of virological failure. Expression of mRNA of Pgp, MRP1, MRP4, and MRP5 in PBMCs was evaluated by real‐time PCR. A high inter‐individual variability was observed in both HIV‐positive individuals and healthy donors but the expression levels of all mRNA analyzed were significantly higher in the HIV‐infected group (P < 0.05). A weak but significant inverse correlation was observed between CD4 cell counts and expression levels of MRP4 and MRP5. Comparison of mRNA expression between individuals with different therapeutic histories showed that expression of MRP4 and MRP5 genes in patients who were both protease inhibitor (PI) and non‐nucleoside reverse transcriptase inhibitor (NNRTI)‐experienced was significantly higher than in patients who were PI experienced but NNRTI‐naïve. In conclusion, the mRNA expression of Pgp, MRP1, MRP4, and MRP5 varies among HIV‐infected patients and healthy donors but is significantly higher in HIV‐positive patients than in donors. The expression of MRP4 and MRP5 seems to correlate with CD4 cell counts. The same protein seems to be overexpressed in patients receiving NNRTIs. J. Med. Virol. 80:766–771, 2008.

Residual viraemia does not influence 1 year virological rebound in HIV-infected patients with HIV RNA persistently below 50 copies/mL

OBJECTIVES It is currently debated whether patients with residual viraemia are at higher risk of virological failure than those attaining <1 HIV RNA copy/mL. We therefore investigated the effect of residual viraemia on virological rebound. METHODS We used a prospective, non-interventional, single-centre, study. This analysis was based on HIV-infected patients with two consecutive HIV RNA viral loads (VLs) of <50 copies/mL as tested by Versant bDNA, followed by two HIV RNA VLs of <50 copies/mL as tested using the Versant kinetic PCR molecular system (kPCR; limit of quantification = 1 copy/mL). Virological rebound was defined as two consecutive HIV RNA values of >50 copies/mL after baseline, and the time to virological rebound was calculated using the Kaplan-Meier method. RESULTS There were 739 eligible patients; 446 (60.4%) had HIV RNA <1 copy/mL (group A) and 293 (39.6%) had residual viraemia (1-49 HIV RNA copies/mL; group B). After a follow-up (median 48.9 weeks), virological rebound occurred in four patients in group A (0.9%) and six patients in group B (2%); the time to virological rebound was similar in the two groups (log-rank test P = 0.231). CD4+ cell recovery (slope) was significantly less in the patients with residual viraemia; +14.3 (-7.7, 43.9) cells/mm(3) per year versus +21.2 (-2.5, 53.2) cells/mm(3) per year; P = 0.036. CONCLUSIONS Residual viraemia assessed by kPCR was not associated with virological rebound during 1 year of follow-up. However, the patients attaining <1 HIV RNA copy/mL showed a small but statistically significant improvement in CD4+ cell recovery.

Number of daily pills, dosing schedule, self‐reported adherence and health status in 2010: a large cross‐sectional study of HIV‐infected patients on antiretroviral therapy

The aim of the study was to assess whether pill burden is associated with self‐reported adherence to current combination antiretroviral regimens and health status in a large sample of unselected and chronically treated HIV‐infected patients.