Constant mitochondrial DNA levels in blood leukocytes of patients enrolled in a NRTI-free therapeutic trial (BIKS-2 study).

OBJECTIVES: Determine if a nucleoside reverse transcriptase inhibitors (NRTI)-free regimen affected mitochondrial DNA (mtDNA) levels in peripheral blood mononuclear cells (PBMCs) of patients enrolled in BIKS-2 trial. METHODS: Antiretroviral (ARV) naïve (N=13) and NRTI experienced (N=7) patients, received lopinavir/ritonavir, a boosted protease inhibitor, and efavirenz, a non-nucleoside reverse transcriptase inhibitor from Month (M) 0 to M12 (1-year BIKS trial) and from M12 to M36 (2-year BIKS-2 trial). MtDNA was quantified at M12, M24 and M36 via real-time PCR assay. RESULTS: From M12 to M36, the 20 patients have maintained undetectable plasma HIV-1 RNA, gained CD4 cells and had no side effects attributable to these drugs. Median mtDNA contents were constant: 478.6 at M12, 478.6 at M24 and 324.4 copies/cell at M36 (pM12-M36=0.5). Because M0 data is missing, these results were compared to those of two groups of age matched individuals: healthy donors and HIV-infected patients before and after exposure to NRTIs. Healthy donors have higher contents (871), followed by patients never treated (602), than by BIKS patients where 7 had toxic NRTIs (478.6) and at last by patients exposed for six months to the most toxic combination (ddI-d4T) (85 copies/cell). CONCLUSION: Lopinavir/ritonavir+efavirenz did not affect mtDNA contents in PBMCs.

Predictors of plasma human immunodeficiency virus type 1 RNA control after discontinuation of highly active antiretroviral therapy initiated at acute infection combined with structured treatment interruptions and immune-based therapies.

Thirty patients with acute human immunodeficiency virus (HIV) type 1 infection received a combination of 3 antiretroviral drugs (n=15) or 4 antiretroviral drugs plus hydroxyurea and interleukin-2 (n=15) for 24 months, followed by 1-3 structured therapeutic interruptions (STIs). Viral control, defined as maintaining plasma viremia <5000 copies/mL without therapy, was achieved in 14 cases. Lymphocyte subsets, plasma HIV-1 RNA loads, proviral DNA loads in peripheral blood mononuclear cells (PBMCs), residual HIV-1 RNA loads in PBMCs and in lymph node cells, and anti-p24 lymphoproliferative response were measured. In the multivariate analysis, proviral DNA loads in PBMCs and anti-p24 lymphoproliferative response assessed at 24 months were independently correlated with viral control after STI. These results enabled us to define a subgroup of patients for whom safe discontinuation of therapy initiated at acute infection was suitable and contributed to ascertaining priority for biological parameter assessment in future clinical trials.

TRIZAL study: switching from successful HAART to Trizivir (abacavir-lamivudine-zidovudine combination tablet): 48 weeks efficacy, safety and adherence results.

OBJECTIVE: To assess the antiviral efficacy, safety, and adherence in subjects who switched to Trizivir following long-term HIV-1 RNA suppression. STUDY DESIGN: A randomized, open-label, multicentre, 48-week comparative study in subjects who have received two nucleoside reverse transcriptase inhibitors plus a protease inhibitor or an nonnucleoside reverse transcriptase inhibitor or three nucleoside reverse transcriptase inhibitors for at least 6 months, with a history of undetectable plasma HIV-1 RNA since initiation of therapy and plasma viral load of < 50 HIV-1 RNA copies/mL at screening. METHODS: Subjects were randomized 1:1 to continue their current treatment or to switch to a simplified treatment with Trizivir administered twice daily. Assessments included plasma HIV-1 RNA, lymphocyte counts, clinical laboratory evaluations, adverse events, and adherence to treatment (obtained via subject self-report). Treatment failure was defined as a plasma viral load of >/= 400 HIV-1 RNA copies/mL on two consecutive occasions or premature discontinuation of randomized treatment. RESULTS: At week 48, the proportion of treatment failures in Trizivir arm (23/106, 22%) was noninferior to that observed in continued arm (23/103, 22%) with a treatment difference stratified by prior ART of 1.2%[-10.1; 12.5]. Incidence of adverse events was similar in both treatment groups. The incidence of possible hypersensitivity reaction in the Trizivir trade mark arm was 10%. Significant reductions in cholesterol and triglyceride plasma levels were observed in the Trizivir arm (P < 0.001 and P = 0.006, respectively). CONCLUSION: Switching to Trizivir offers a potent and simplified regimen with equivalent efficacy and significant improvement in lipid abnormalities compared to continued triple therapy.

Peripheral neuropathy during stavudine-didanosine antiretroviral therapy.

Peripheral neuropathy (PN) is among the most frequent side effects described with nucleoside reverse transcriptase inhibitors (NRTIs). We investigated the incidence, evolution and predictive factors of PN during stavudine (d4T)-didanosine (ddI) combination therapy in 65 HIV infected patients, previously treated with zidovudine and/or zalcitabine (ddC) for at least 3 months. A subset of 16 patients was referred for systematic electromyographic examination at weeks 0 and 24: six among the 16 exhibited nerve conduction abnormalities at day 0, probably related to previous ddC treatment in four of those and to HIV infection in the other two, with worsening of abnormalities in one patient at week 24. In total, seven of the 59 assessable patients (11.8%) exhibited grade 2-3 neuropathy, with a median time of occurrence of 17 weeks. Distal, symmetrical paraesthesias of the extremities were the first symptoms in all the patients; none had motor symptoms. In all the patients, PN resolved rapidly after stopping d4T. There were no statistically different parameters between the seven cases and the other 52 patients according to CD4 T cells, HIV RNA, Centers for Disease Control and Prevention (CDC) stage C or d4T daily dose. In our study, the d4T-ddI combination did not seem to increase the incidence of PN; risk factors for PN could not be identified, probably in part because of the low number of patients with PN.

Impact of immune interventions on proviral HIV-1 DNA decay in patients receiving highly active antiretroviral therapy.

OBJECTIVE: To measure the evolution of proviral HIV-1 DNA levels in patients receiving highly active antiretroviral therapy (HAART) compared to those treated with HAART plus interleukin-2 (IL-2) and hydroxyurea. DESIGN: Prospective randomised trial. METHODS: Twenty-two HIV-1 infected patients were randomly assigned to a five-drug antiretroviral regimen for 72 weeks, with or without IL-2, followed by a three-drug regimen up to week 120 with additional hydroxyurea in patients having received IL-2. HIV-1 DNA levels in peripheral blood mononuclear cells (PBMC) were measured regularly using the Amplicor Monitor kit from Roche Diagnostics (Meylan, France). Potentially infectious HIV-1 was cultured in enhanced conditions from circulating CD4 T cells at week 120. RESULTS: During the study period of 120 weeks, HIV-1 DNA levels in PBMC decreased by -1.1 log in patients treated with HAART only compared with -1.8 log in patients with additional IL-2 and hydroxyurea. A two-phase decay rate was observed, with an inflexion point at 12 weeks. The second decay was slow, with mean half-lives of 130.1 +/- 21.3 weeks and 95.1 +/- 26.3 weeks for patients on HAART and those receiving additional IL-2 and hydroxyurea, respectively. At week 120, one out of 11 patients with HAART alone compared to six out of 11 in the group with IL-2 and hydroxyurea had undetectable proviral DNA levels and three of them had unsuccessful recovery of replication-competent HIV-1 from blood CD4 T cells. CONCLUSION: Therapeutic strategies combining HAART and immune interventions have higher potency to decrease the number of infected cells than HAART alone.

Phenotypic and genotypic resistance to nucleoside reverse transcriptase inhibitors in HIV-1 clinical isolates.

OBJECTIVE: To assess phenotypic and genotypic cross-resistance to nucleoside reverse transcriptase inhibitors in patients treated with a combination including zidovudine, who were switched to a combination including stavudine. METHODS: We analysed 24 clinical HIV-1 isolates from 12 patients before and several months after therapeutic switching. Plasma HIV-1 RNA was measured using quantitative polymerase chain reaction (Roche). Genotypic resistance was measured by sequencing the reverse transcriptase gene from plasma HIV-1 RNA. Phenotypic resistance was measured using a recombinant assay (Virco). RESULTS: Patients were treated with a combination including zidovudine for a mean (+/- SEM) period of 21.8 +/- 3.5 months and had a plasma viral load of 4.1 +/- 0.2 log HIV-1 RNA copies/mL (time 1). After a mean period of 19.3 +/- 1.6 months following the therapeutic change, the plasma viral load was 3.6 +/- 0.1 log copies/mL (time 2). At time 1, genotypic resistance to zidovudine was found in all cases (41L: four cases; 41L, 215Y: five cases; 41L, 210W, 215Y: two cases; K70R: one case) with a mean 6.6 +/- 1.6-fold increase in the median inhibitory concentration (IC50) to zidovudine and 1.7 +/- 0.4-fold to stavudine. At time 2, genotypic resistance to zidovudine was found in 11 out of 12 cases (41L: two cases; 41L, 215Y: six cases; 41L, 210W, 215Y: two cases; M41L, D67N, L210W, T215Y: one case) with a mean 18.9 +/- 8.8-fold increase in the IC50 to zidovudine and 1.4 +/- 0.4-fold to stavudine CONCLUSIONS: In this clinical series of patients with suboptimal control of plasma HIV-1 RNA using a combination including zidovudine, the presence of zidovudine-related mutations was associated with a decreased phenotypic sensitivity to this drug. Despite persistent HIV-1 replication, switching to stavudine was associated with a further decrease in phenotypic sensitivity to zidovudine but not to stavudine after 19 months. These data suggest that stavudine remains a possible option in zidovudine-experienced patients.

Prednisolone does not prevent hypersensitivity reactions in antiretroviral drug regimens containing abacavir with or without nevirapine.

OBJECTIVES: To determine the effect of adjuvant prednisolone use on the development of abacavir (ABC)- and nevirapine (NVP)-associated hypersensitivity reactions (HSR). METHODS: Randomized open-label study in antiretroviral-naive adult HIV-1 infected patients using a factorial design in which NVP and/or hydroxyurea (HU) and/or prednisolone are added to a regimen of ABC, zidovudine and lamivudine. Prednisolone (40 mg once daily) was added for the first 2 weeks of treatment. As it was difficult to distinguish ABC-associated HSR from NVP-associated HSR, these events were treated as a composite endpoint. The odds ratio (OR) of developing HSR for prednisolone-use was calculated with and without stratification by NVP and/or HU. Logistic regression was performed to identify risk factors for developing HSR. RESULTS: Of the 229 patients 115 were randomized to prednisolone and 114 to no-prednisolone; 19 (17%) and 11 (10%) patients, respectively, developed HSR. The expected prevention of HSR by prednisolone use was not observed. In fact use of prednisolone showed an increased risk for HSR although this did not reach statistical significance [OR, 1.82; 95% confidence interval (CI), 0.82-4.03]. There was a higher incidence of HSR in the NVP group than in the non-NVP group (20% versus 6%; P = 0.002). An additional risk factor identified in a multivariate logistic model was a high baseline CD4 cell count (OR, 1.26 per 100 x 10(6) cells/l increase; 95% CI, 1.06-1.51). CONCLUSIONS: The simultaneous start of ABC and NVP in first-line antiretroviral regimens should be avoided because of a high (20%) incidence of HSR. Short-term therapy with prednisolone did not prevent HSR in patients using ABC with or without NVP.

Persistence of HIV-1 resistance in lymph node mononuclear cell RNA despite effective HAART.

OBJECTIVE: To analyse the presence of genotypic and phenotypic resistance in lymph node mononuclear cells from patients with sustained undetectable plasma HIV-1 RNA with highly active antiretroviral therapy. DESIGN: Cross-sectional study on 27 HIV-infected patients receiving triple therapy for a mean period of 232.8 +/- 22.1 weeks. METHODS: HIV-1 RNA was measured in plasma and lymph node cells using PCR. Reverse transcriptase and protease genes were sequenced from HIV-1 RNA obtained from lymph node cells and from peripheral blood mononuclear cell proviral DNA using a commercially available kit (TruGene). Phenotypic resistance was assessed by using a recombinant virus assay (AntiVirogram). RESULTS: Mutations were not found in lymph node mononuclear cell RNA in six out of nine patients on first-line regimens although they were detected in 15 out of 18 who received prior suboptimal combinations. Phenotypic resistance was confirmed in most of these cases. These patterns of resistance were closely related to patients' history of antiretroviral therapy and genotypic analysis of plasma HIV-1 RNA taken just before initiation of the current regimen. In half the patients analysed, resistance mutations found in lymph nodes were not always detected in archival proviral DNA from blood cells. Mean levels of HIV-1 RNA in lymph node cells were not different in patients exhibiting resistance compared with those harbouring wild-type viruses. CONCLUSION: These data demonstrate that resistant HIV-1 is produced in lymphoid tissues for prolonged periods despite effective therapy. The mechanism could represent a release from previously infected cells rather than new cycles of cellular infection.

HIV-1 induction-maintenance at the lymph node level: the "Apollo-97" Study.

OBJECTIVE: To assess the effects of five-drug combination therapy on HIV-1 load in lymph nodes and subsequent maintenance with four and three drugs. METHODS: Ten pharmacotherapeutically naive patients received a combination of zidovudine, lamivudine, didanosine, ritonavir, and saquinavir for 24 weeks, then zidovudine, lamivudine, didanosine, and saquinavir for the next 24 weeks, and finally zidovudine, lamivudine, and saquinavir for the last 24 weeks. HIV-1 RNA in lymph nodes was measured using quantitative polymerase chain reaction (PCR) at baseline, after 12, 24, 48, and 78 weeks. Plasma HIV-1 RNA, proviral DNA in peripheral blood mononuclear cells (PBMCs), circulating lymphocyte subsets, and protease inhibitor levels in blood were also regularly measured. Genotypic resistance was assessed in the different compartments in 2 patients who were failed by therapy. RESULTS: HIV-1 RNA decreased in lymph nodes in 9 patients and was stable in 1 despite initial control of plasma replication <20 copies/ml in each patient. Lymph node levels rebounded in 1 patient at week 72 as a result of lack of adherence and remained stable in the 8 others despite maintenance regimens. This represents a mean drop of -3.17 log in lymph nodes for the 8 patients maintaining undetectable viremia at 72 weeks. In the patient with stable lymph node viral RNA, selection of the M184V mutation was demonstrated at this level before detection in plasma and low blood saquinavir levels were found throughout the study. Continuous improvements in immune parameters were observed in all cases, although PBMC proviral DNA levels either showed a continuous decrease or stabilized to a plateau. CONCLUSIONS: More complex regimens do not perform better in lymph nodes than classic triple therapy. The persistence of HIV-1 RNA in lymph nodes could be related with cellular resistance mechanisms rather than an insufficient potency of the regimens.

Pilot study of a combination of highly active antiretroviral therapy and cytokines to induce HIV-1 remission.

A pilot study of a combination of highly active antiretroviral therapy (HAART) and cytokines in early HIV-1 infection has been undertaken to test the hypothesis that HIV-1 remission can be reached with this strategy by flushing latently infected viral reservoirs. Ten previously antiretroviral naive patients have received a combination of zidovudine, lamivudine, didanosine, saquinavir, and ritonavir for 72 weeks. Between weeks 12 and 48, three courses of interleukin (IL)-2 (7.5 millions of international units [MUI] twice a day for 5 consecutive days) and 2 courses of gamma-interferon (IFN) (100 microg every other day during 2 weeks) were administered subcutaneously. All patients reached plasma HIV-1 RNA levels < 20 copies/ml within 12 +/- 4 weeks. Transient increases in plasma levels (< 120 copies/ml) were observed during administration of IL-2, but less frequently during gamma-IFN administration. HIV-1 RNA decreased in lymph node cells by approximately 4 log, then remained stable after week 24. A mean drop of -0.8 log in peripheral blood mononuclear cell (PBMC) proviral DNA was observed during the trial. Isolation of potentially infectious HIV-1 was successful in each case by coculture of CD4+ T cells taken at week 72. The 2 patients who stopped therapy at the end of the trial showed rebounding plasma HIV-1 RNA levels within a few weeks. No additional mutations were selected in comparison with those present at baseline in 8 patients. In addition, 2 patients developed new mutations in the reverse transcriptase or protease gene and in 1 case, resistance selection was found in lymphoid tissue HIV-1 RNA but not in latently infected cells. In all cases, a rapid increase in both naive and memory CD4+ T cells was observed, with a reduction in activation markers and preservation of the CD8+CD28+ subset. Consequently, an aggressive regimen of HAART and cytokines administered in early stage disease is associated with a positive effect in terms of proviral load reduction and immune reconstitution but is unable to induce HIV-1 remission, allowing low levels of viral replication to persist in lymphoid reservoirs.

Increased mitochondrial toxicity with ribavirin in HIV/HCV coinfection.

In two of 15 patients coinfected with HIV and hepatitis C virus who received interferon-alpha plus ribavirin in addition to HAART, we observed multiorgan dysfunction and lactic acidaemia. As ribavirin is a nucleoside analogue, an increased risk of mitochondrial toxicity can be induced in HIV-infected patients already treated with nucleoside analogues, leading to clinical deterioration in some cases.

Long-term evaluation of triple nucleoside therapy administered from primary HIV-1 infection.

OBJECTIVE: To study the long-term effect of triple-drug therapy initiated at the time of primary HIV-1 Infection and to evaluate the persistance of replication-competent virus in responding patients. METHODS: Prospective open-label pilot study. Patients received a combination of zidovudine, didanosine and lamivudine. Viral sequencing of the reverse transcriptase gene was performed before therapy and during follow-up. HIV-1 RNA and DNA as well as CD4 and CD8 T lymphocyte subsets were measured in blood and in lymph node biopsies during therapy. Isolated blood CD4 T cells were cultured in conditions that improved HIV isolation. Three patients received in vivo interleukin-2 and gamma-interferon in order to try to identify intracellular pools of replication-competent virus. SETTING: A tertiary care general hospital. PATIENTS: Fifteen patients observed within 28 days following the acute retroviral syndrome. RESULTS: After a mean follow-up of 27.5+/-2.9 months, plasma RNA remained < 20 copies/ml (four patients), fluctuated between 20 and 120 copies/ml (six patients) or rebounded (five patients). M184V and/or T215Y mutations were demonstrated in two of these last five patients. Proviral DNA in peripheral blood mononuclear cells (PBMC) decreased by an average of -1 log after 16+/-3 months, reaching undetectable levels in three patients. The culture of isolated CD4 T cells yielded virus in all but two patients. These last were characterized by a waning antibody reactivity on the Western blot, undetectable proviral DNA in PBMC and undetectable RNA in lymph nodes. Cytokine administration in vivo had no effect in one patient and unmasked plasma RNA in the other. Stopping therapy in the first patient led to a rebound in plasma RNA. CONCLUSION: Despite a lack of detectable plasma viral activity in some patients after 3 years of triple nucleoside therapy administered since the acute retroviral syndrome, replication-competent virus can still be demonstrated.

Efficacy and safety of stavudine and didanosine combination therapy in antiretroviral-experienced patients.

OBJECTIVES: To assess the efficacy, tolerance, and safety of combination antiretroviral therapy with didanosine and stavudine in HIV-infected patients with CD4+ cell counts > 100 x 10(6)/l and HIV plasma RNA > 10(4) copies/ml previously treated with other antiretroviral agents for at least 3 months. DESIGN: In this open, multicentre, non-randomized, Phase II pilot study, adult patients were administered didanosine (200 mg twice daily) plus stavudine (40 mg twice daily) for 6 months. Patients for whom the first regimen had led to undetectable HIV RNA levels were offered a second 6-month course of treatment; those who had achieved insufficient immunological and virological gains in the first 6 months were given a new combination. METHODS: Primary evaluation of efficacy was based on viral load measured by branched DNA second-generation testing (lower limit of detection, 500 copies/ml) and CD4+ cell counts; secondary evaluations included AIDS-defining events and clinical side-effects. RESULTS: Sixty-five patients with median prior antiretroviral therapy of 24 months (65 with zidovudine, 29 with zalcitabine) were included in the study. At baseline, median CD4+ cell count was 198 x 10(6)/l and median plasma HIV RNA was 80000 copies/ml (4.9 log10 copies/ml). In this heavily pretreated population, an increase in the mean CD4+ cell count was observed (+70 x 10(6)/l at 24 weeks). In addition, rapid and prolonged antiviral activity was seen, with a mean maximal decrease of 1.1 log10 copies/ml at week 4, a mean decrease of 0.89 log10 copies/ml at week 24, and a plasma RNA viraemia < 500 copies/ml achieved in 14% of patients at week 24. CONCLUSIONS: Combination therapy with stavudine and didanosine is safe and leads to a sustained antiviral effect, even in patients with prolonged prior antiretroviral exposure and low CD4+ cell counts.

Correlation between plasma levels of cytokines and HIV-1 RNA copy number in HIV-infected patients.

The plasma levels of HIV-1 RNA, tumor necrosis factor alpha (TNF-alpha), soluble receptors type II of TNF-alpha (sTNF-alpha RII), soluble receptors of interleukin-4 (sR IL-4), interleukin-6 (IL-6), soluble receptors of interleukin-6 (sR IL-6), granulocyte macrophage colony stimulating factor (GM-CSF), soluble receptors of GM-CSF (sR GM-CSF), RANTES, MIP-1 alpha and MIP-1 beta were measured in 80 HIV-infected patients. All patients had not been treated previously with antiretroviral drugs and did not present a recent history of opportunistic infection. A statistically significant correlation was found between HIV-1 RNA and TNF-alpha (p = 0.005) or sTNF-alpha RII levels (p < 0.001). RANTES and MIP-1 alpha levels did not correlate with HIV-1 RNA. MIP-1 beta levels were correlated with plasma RNA titers in patients with CD4+ T cells < 200 x 10(6)/l (p = 0.03). MIP-1 alpha and sR IL-4 levels were significantly different according to the CD4+ T cell range (p = 0.003 and p = 0.0002, respectively). GM-CSF and sR GM-CSF were undetectable in each case. These data confirm that HIV-1 replication in the plasma is correlated with TNF-alpha levels, but do not show a clear correlation with levels of the chemokines studied.

Residual human immunodeficiency virus type 1 RNA in lymphoid tissue of patients with sustained plasma RNA of <200 copies/mL.

Human immunodeficiency virus type 1 (HIV-1) RNA was measured in lymph node (LN) mononuclear cells of 50 patients with sustained plasma RNA of <200 copies/mL with therapy. Six patients had received a combination of three reverse transcriptase inhibitors (RTIs) since primary infection, 11 received this same combination during chronic disease, 21 received a combination of two RTIs plus a protease inhibitor (PI), and 12 received three RTIs plus a PI. The mean overall duration of therapy was 8.9 +/- 0.5 months (range, 5-24), with no significant difference between groups. LN HIV-1 RNA levels varied from undetectable to 1.7 million copies/10(6) cells according to cases. The mean LN HIV-1 RNA level was 2.99 +/- 0.42 log10 copies/10(6) cells in the 17 patients receiving three RTIs compared with 1.93 +/- 0.25 log10 copies/10(6) cells in the 33 patients receiving a PI (t test, P = .02). These data demonstrate that highly active antiretroviral regimens have unequivalent effects on LNs and invite redefinition of suboptimal therapy at this level.