Ma Johnson

Comparison of first-line antiretroviral therapy with regimens including nevirapine, efavirenz, or both drugs, plus stavudine and lamivudine: a randomised open-label trial, the 2NN Study

BACKGROUND The 2NN Study was a randomised comparison of the non-nucleoside reverse-transcriptase inhibitors (NNRTI) nevirapine and efavirenz. METHODS In this multicentre, open-label, randomised trial, 1216 antiretroviral-therapy-naive patients were assigned nevirapine 400 mg once daily, nevirapine 200 mg twice daily, efavirenz 600 mg once daily, or nevirapine (400 mg) and efavirenz (800 mg) once daily, plus stavudine and lamivudine, for 48 weeks. The primary endpoint was the proportion of patients with treatment failure (less than 1 log(10) decline in plasma HIV-1 RNA in the first 12 weeks or two consecutive measurements of more than 50 copies per mL from week 24 onwards, disease progression [new Centers for Disease Control and Prevention grade C event or death], or change of allocated treatment). Analyses were by intention to treat. FINDINGS Treatment failure occurred in 96 (43.6%) of 220 patients assigned nevirapine once daily, 169 (43.7%) of 387 assigned nevirapine twice daily, 151 (37.8%) of 400 assigned efavirenz, and 111 (53.1%) of 209 assigned nevirapine plus efavirenz. The difference between nevirapine twice daily and efavirenz was 5.9% (95% CI -0.9 to 12.8). There were no significant differences among the study groups in the proportions with plasma HIV-1 RNA concentrations below 50 copies per mL at week 48 (p=0.193) or the increases in CD4-positive cells (p=0.800). Nevirapine plus efavirenz was associated with the highest frequency of clinical adverse events, and nevirapine once daily with significantly more hepatobiliary laboratory toxicities than efavirenz. Of 25 observed deaths, two were attributed to nevirapine. INTERPRETATION Antiretroviral therapy with nevirapine or efavirenz showed similar efficacy, so triple-drug regimens with either NNRTI are valid for first-line treatment. There are, however, differences in safety profiles. Combination of nevirapine and efavirenz did not improve efficacy but caused more adverse events.

Cytomegalovirus retinitis in AIDS patients : influence of cytomegaloviral load on response to ganciclovir, time to recurrence and survival

Objectives: Despite life‐long maintenance therapy, cytomegalovirus (CMV) retinitis frequently progresses in patients with AIDS. Virological markers that could clarify pathogenesis and identify risk factors for progression are required. Design and methods: We prospectively recruited 45 patients with CMV retinitis. Blood and urine samples were collected before and after induction therapy, and on a monthly basis thereafter during routine medical and ophthalmological assessment, and at any time retinitis progressed. CMV load was measured by quantitative‐ competitive polymerase chain reaction (PCR). Results: The median time to first progression of retinitis was 78 days and to death was 8.7 months. Eighty‐five per cent of patients who were PCR‐positive at diagnosis of retinitis became PCR‐negative after 21 days of ganciclovir induction therapy. Six patients who remained PCR‐positive after 21 days of treatment had a significantly higher CMV load at presentation (P = 0.005), and a shorter time to first progression of retinitis of 40 days. High CMV loads in blood at presentation were associated with a shorter time to progression (P = 0.16; relative hazard, 1.57) and a significantly shorter time to death (P = 0.004; relative hazard, 1.76). This significant relationship with survival remained after adjustment for potential confounding variables (CD4 count, age, method of drug administration). Conclusions: We conclude that CMV load in the blood of AIDS patients is an important factor in the pathogenesis of retinitis, and quantification of CMV could be used to both select patients for controlled clinical trials and to optimize individual anti‐CMV induction therapy.

Quantification of human herpesvirus 6 in immunocompetent persons and post-mortem tissues from AIDS patients by PCR.

A quantitative competitive PCR assay for human herpesvirus 6 (HHV-6) was developed. Firstly, viral burden was determined in the blood of 25 healthy persons. Using 1 microgram of DNA, the prevalence of HHV-6 was 36% (9/25). Eight persons had viral loads of < or = 32 HHV-6 genomes/microgram DNA. The viral burden in the ninth individual was 1.2 x 10(6) HHV-6 genome copies/microgram DNA, which remained constant over a period of 10 months. This demonstrates the persistence of a high HHV-6 load in the absence of apparent disease. Secondly, HHV-6 burden was determined in 100 post-mortem tissues from seven AIDS patients and three controls. For all tissues combined, there was a statistically significant higher median viral load in AIDS patients (56 copies/microgram DNA, range 0-43321) compared to controls (10 copies/microgram DNA, range 0-423) (P = 0.04). The precision and reproducibility of this assay will allow hypotheses concerning the pathogenic potential of HHV-6 to be tested quantitatively.

Cytomegalovirus polymerase chain reaction viraemia in patients receiving ganciclovir maintenance therapy for retinitis

Objectives:To determine whether recurrence of polymerase chain reaction (PCR) viraemia during maintenance ganciclovir for cytomegalovirus (CMV) retinitis correlates with (i) CMV disease at a new anatomical site, (ii) progression of the presenting retinitis, or (iii) acquisition of genetic changes in gene UL97 associated with resistance to ganciclovir. Design:A previously described cohort of 45 patients presenting with first episode retinitis was followed clinically using ophthalmoscopy and serial tests for PCR viraemia for a median of 7 months. CMV viral load and genetic markers of ganciclovir resistance were measured in PCR-positive samples. Methods:PCR amplification of the glycoprotein B region of CMV and quantitative competitive PCR assays were employed. Genetic changes in UL97 were identified by sequencing/point mutation assay. Results:PCR viraemia correlated significantly with new episodes of CMV disease (P = 0.011) and a trend was seen for the association with progression of retinitis (P = 0.07). Amongst the 14 patients PCR-positive during maintenance ganciclovir, 10 (71%) had genetic markers of resistance. None of these patients became PCR-negative in blood after reinduction ganciclovir therapy compared with three out of four without markers of resistance (P = 0.022). Conclusions:CMV PCR viraemia correlated strongly with the development of new episodes of CMV disease. Most patients with progression of retinitis remained PCR-negative in blood, consistent with therapeutic failure due to poor intraocular penetration of ganciclovir. However, the minority who were PCR-positive in blood may have reinfected their eye, and frequently had markers of ganciclovir resistance. The implications of these findings for the management of patients with CMV disease are discussed.

Natural history of untreated cytomegalovirus retinitis

Cytomegalovirus infection is common in patients with AIDS, and often causes retinitis. Treatment is rarely curative, but the progression of retinitis is delayed. The untreated course of cytomegalovirus retinitis in AIDS is unknown. We report a 35-year-old man with retinitis who refused treatment. Retinitis resulted in blindness within 6 months. Measurement of cytomegalovirus genomes showed an increasing viral load in blood and urine.

Rapid reconstitution of humoral immunity against cytomegalovirus but not HIV following highly active antiretroviral therapy.

Objective: To determine the kinetics of reduction in human cytomegalovirus (HCMV) load and specific anti-glycoprotein B (gB) immune responses in patients with concurrent HCMV DNAaemia following the initiation of highly active antiretroviral therapy (HAART). Design: Sequential analysis of eleven patients with HCMV DNAaemia who received HAART and eleven control patients with HCMV DNAaemia. Methods: HCMV load was measured by quantitative competitive polymerase chain reaction and anti-gB, anti-HIV Env and Gag responses by an end-point dilution immunofluorescence assay using recombinant antigens expressed in insect cells. Estimates of the efficacy of the reconstituting immune system at controlling HCMV replication were based on previous dynamic models. Results: In patients initiating HAART, HCMV DNA levels in blood declined rapidly, with a median half-life of 5.2 days, consistent with an efficacy of the reconstituting immune system at inhibiting HCMV replication of 52.8–85% (median, 61%). Commensurate with this decrease, a significant increase in anti-gB titres was observed in the post-HAART period (corresponding to an average fourfold increase in titre by 1 month rising to an eightfold increase at month 3; P = 0.01). No changes in titre were observed in the control group or for anti-HIV Gag antibody levels, while anti-HIV Env antibody levels decreased after HAART. Conclusions: In patients with HCMV DNAaemia, reconstitution of humoral immunity to HCMV gB occurs rapidly following the initiation of HAART. These changes contrast with the patterns observed for anti-HIV humoral immune responses.

Development of a point mutation assay for the detection of human cytomegalovirus UL97 mutations associated with ganciclovir resistance

A point mutation assay was developed to detect the quantitative prevalence of mutations at codons 460 (M to I; M to V), 520 (H to Q), 594 (A to V) and 595 (L to F; L to S) within the UL97 gene of human cytomegalovirus which segregate with ganciclovir resistance. Synthetic mixtures of wild-type and mutant plasmids containing the UL97 gene were amplified by nested polymerase chain reaction and the 700 base pair amplicon subsequently subjected to the point mutation assay. In plasmid reconstruction experiments, there was a high correlation between experimentally derived percentage mutant with the theoretical values. The assay was then used to assess the changes in the genetic composition of the UL97 gene in three patients on prolonged ganciclovir therapy. All three patients developed genotypic resistance against ganciclovir involving mutation at codon L595S, L595F and double mutation at codons L595F and M460I. In one patient, alteration of therapy to foscarnet did not affect the composition of UL97 and virus remained genotypically resistant to ganciclovir. In contrast, in two patients whose therapy was altered to cidofovir (HPMPC), repopulation with cytomegalovirus strains carrying the wild-type (ganciclovir-sensitive) codon at positions 595 and 460 occurred. The potential use of this assay for the rapid detection of cytomegalovirus resistance in patients on long-term ganciclovir therapy is discussed.

Flow cytometric analysis of the stimulatory response of T cell subsets from normal and HIV-1+ individuals to various mitogenic stimuli in vitro

A novel technique is described which allows the study of the responses of T cell subpopulations stimulated in bulk cultures without interfering with cell‐cell interactions. The number and phenotype of lymphoblasts developing following stimulation with phytohaemagglutinin (PHA), anti‐CD3, staphylococcal protein A (SPA) and pokeweed mitogen (PWM) was determined in HIV‐P and HIV‐1+ patients using a new five‐parameter flow cytometric method. We found that normal T ceils responded faster to PHA than lo any of the other mitogens tested. The peak of the PHA response occurred on day 3. followed by anti‐CD3 and SPA on day 4 and PWM mitogen on day 5. Although PHA and anti‐CD3 stimulated up to 95% and 80% of lymphocytes, respectively, SPA and PWM stimulated only 40% and 30% of cells, respectively. A defective T cell response was observed in lymphocytes cultured from asymptomatic HlV‐1+ patients compared with negative controls. This loss of response was related to a selective mortality of T cells following mitogenic stimulation, referred to as activation‐associated lymphocyte death (AALD). The results showed that stronger mitogens (PHA and anti‐CD3) induced AALD in a larger proportion (50‐60%) of T cells than weaker mitogens such as SPA and PWM (50‐40%). and that AALD affected different lymphocyte subsets to different extents. AALD occurred more frequently in total CD4+ and CD45RO+ T ceils compared with CD4+ and CD45RA+ T cells, but memory CD4+ T cells were the population most severely affected in samples from HIV‐I+ donors.

Antiretroviral treatment reverses HIV-induced reduction in the expression of surface antigens on alveolar macrophages in AIDS patients.

MoAbs and immunoperoxidasc methods were used to identify antige‐resenting and phagocytic cells and to assess expression of HL‐R molecules on cells obtained by bronchoalvcolar lavagc (BAL) from 33 AIDS patients and nine normal volunteers. In 17 patients, not receiving antiretroviral therapy, the expression of HL‐R molecules (MoAb RFDR1) as well as the percentages of cells expressing RFD1 marker for antige‐resenting cells and RFD7 marker for mature phagocytes were significantly reduced. However, in BAL obtained after commencing treatment with zidovudine (AZT)in 21 patients or with 2′,3′–dideoxyinosine(DD1)in five patients, the expression of the markers studied was found to have returned to levels of expression seen in normal lavages. The changes observed were clearly associated with antiretroviral treatment and did not correlate with applications of other drugs, blood CD4 counts or presence of infectious organisms in BAL fluid. As the alterations in the expression of HL‐R molecules and RFD1 marker on macrophages have been shown to be associated with functional capacities of these cells, the reversal of impaired expression of phenotypic markers on alveolar macrophages in AIDS patients by AZT and DDI signifies an important ability of these drugs to modify immune reactivity and emphasizes the need to monitor such functions in HIV disease.

Pooled week 48 safety and efficacy results from the ECHO and THRIVE phase III trials comparing TMC278 vs EFV in treatment-naïve, HIV-1-infected patients

7‐11 November 2010, Tenth International Congress on Drug Therapy in HIV Infection, Glasgow, UK