Immunodeficiency Virus Type 1-infected Patients Research Articles

Association of TREX1 polymorphism with disease progression in human immunodeficiency virus type-1 (HIV-1) infected patients.

The time interval between HIV-1 infection and AIDS development is not the same in all patients and depends largely on the genetic background of the individual. Polymorphisms in the TREX1 gene, the main enzyme in the clearance of cytosolic DNA, affect type 1 interferon-mediated inflammatory response in HIV-1 infection. We aimed to study the role of a single nucleotide polymorphism (rs3135941) of the TREX1 gene and the rate of disease progression in patients infected with HIV-1. A total of 190 HIV-1 infected patients were recruited. Patients' demographic and laboratory data including CD4 counts, viral load, and antiretroviral therapy (ART) were collected. The genotype of rs3135941 was determined by a PCR-SSP method. The rate of progression to AIDS was calculated with Kaplan-Meier survival analysis using Stata software. The patients were divided into rapid and slow progressors based on time interval of CD4 drop below 350/µl. Kaplan-Meier analysis revealed an accelerated disease progression in patients with TC and CC genotypes (HR = 1.49, 95% CI = 1.01-2.17). The mean values of the first 5-year CD4 counts were significantly different in patients who had CC and TC genotypes compared to the TT group (p = 0.036). The result of this study emphasizes the importance of TREX1 polymorphism in HIV-1 progression. These data warrant further investigation into the role of other polymorphisms of TREX1.

Comparative performances of serologic and molecular assays for detecting human T lymphotropic virus type 1 and type 2 (HTLV-1 and HTLV-2) in patients infected with human immunodeficiency virus type 1 (HIV-1)

The present study evaluated several techniques currently available (commercial kits and in-house assays) for diagnosing human T lymphotropic viruses types 1 and 2 in two groups of patients enrolled at HIV/AIDS specialized care services in São Paulo: Group 1 (G1), n=1608, 1237 male/371 female, median age 44.3 years old, majority using highly active antiretroviral therapy (HAART); G2, n=1383, 930 male/453 female, median age of 35.6 years old, majority HAART naïve. Enzyme immunoassays [(EIA) Murex and Gold ELISA] were employed for human T lymphotropic viruses types 1 and 2 screening; Western blotting (WB), INNO-LIA (LIA), real-time PCR pol (qPCR), and nested-PCR-RFLP (tax) were used to confirm infection. Samples were considered human T lymphotropic viruses types 1 and 2 positive when there was reactivity using at least one of the four confirmatory assays. By serological screening, 127/2991 samples were positive or borderline, and human T lymphotropic virus infection was confirmed in 108 samples (three EIA-borderline): 56 human T lymphotropic virus type 1 [G1 (27)+G2 (29)]; 45 human T lymphotropic virus type 2 [G1 (21)+G2 (24)]; one human T lymphotropic virus type 1+human T lymphotropic virus type 2 (G2); six human T lymphotropic virus [G1 (2)+G2 (4)]. Although there were differences in group characteristics, human T lymphotropic viruses types 1 and 2 prevalence was similar [3.1% (G1) and 4.2% (G2), p=0.113]. The overall sensitivities of LIA, WB, qPCR, and PCR-RFLP were 97.2%, 82.4%, 68.9%, and 68.4%, respectively, with some differences among groups, likely due to the stage of human T lymphotropic virus infection and/or HAART duration. Indeterminate immunoblotting results were detected in G2, possibly due to the seroconversion period. Negative results in molecular assays could be explained by the use of HAART, the occurrence of defective provirus and/or the low circulating proviral load. In conclusion, when determining the human T lymphotropic virus infection, the findings highlight that there is a need to consider the blood samples with borderline results in screening assays. Of all the tested assays, LIA was the assay of choice for detecting human T lymphotropic virus type 1 and human T lymphotropic virus type 2 in human immunodeficiency virus type 1-infected patients.

Frequent gross deletions in pol gene in 10 HIV-1 infected patients treated with Korean red ginseng for 3 years: dosage dependency

Background To my knowledge, there is only one report on gross deletions in the pol gene (gΔpol) [1] although there are many reports on gross deletions in the nef gene (gΔnef). It is known that Korean red ginseng (KRG) slows depletion of CD4 T cells in human immunodeficiency virus type 1-infected patients [2]. We reported an association between KRG intake and gΔnef [3] in 10 HIV-1 infected patients treated with KRG. Here, we investigated whether KRG intake also induces gΔpol.

Effect of Cytomegalovirus-Induced Immune Response, Self Antigen–Induced Immune Response, and Microbial Translocation on Chronic Immune Activation in Successfully Treated HIV Type 1–Infected Patients: The ANRS CO3 Aquitaine Cohort

We evaluated the impact of cytomegalovirus (CMV)-induced immune responses, autoimmune-induced immune responses, and microbial translocation on immune activation in 191 human immunodeficiency virus type 1-infected patients from the ANRS CO3 Aquitaine Cohort. All enrolled subjects had achieved long-term virological suppression during receipt of combination antiretroviral therapy (cART). HLA-DR(+)/CD38(+) expression was 16.8% among CD8(+) T cells. Independent of age, CD4(+) T-cell count, 16S ribosomal DNA load, and regulatory T-cell count, positive results of Quantiferon CMV analysis (P = .02), positive results of CMV-pp65 enzyme-linked immunosorbent spot analysis (P = .01), positive results of CMV-pp65-specific CD8(+) T-cell analysis (P = .05), and CMV seropositivity (P = .01) were associated with a higher percentage of CD8+ T cells that expressed HLA-DR+/CD38+. Autoimmune response and microbial translocation were not associated with immune activation. Therefore, the CMV-induced immune response seems to be associated with chronic immune activation in cART recipients with sustained virological suppression.

Epstein-Barr Virus load and immune activation in Human Immunodeficiency Virus type 1-infected patients

BackgroundPatients infected with HIV-1 are at high risk of developing Epstein-Barr Virus (EBV)-related diseases. Chronic immune activation is a hallmark of HIV-1 pathogenesis and may play a role in B-cell stimulation and expansion of EBV-infected cells. ObjectivesThe aim of the study was to define the relationship between parameters of immune activation and EBV load in HIV-1-infected subjects. Study designA total of 156 HIV-1-infected patients were studied. EBV types 1 and 2 were quantified on peripheral blood mononuclear cells by multiplex real-time PCR. Plasma levels of cytokines and lipopolysaccharide (LPS) were determined by immunoenzymatic assays. B-cell activation was analyzed by flow cytometry. ResultsEBV-DNA was detected in 114 patients, and in all but 3 was EBV type 1. The median [interquartile] EBV-DNA load was 43[1–151] copies/105 PBMC. EBV-DNA load was higher in patients with detectable HIV-1 plasma viremia, despite good immunological status (CD4>500 cells/μl), than in patients with undetectable HIV-1 plasma viremia regardless of immunological status (46[5–136] copies/105 cells vs 17[1–56] copies/105 cells, p=0.008). Patients with high EBV-DNA load (>median value) had higher levels of LPS and proinflammatory cytokines (IL-6, IL-10 and TNF-α) than patients with low EBV load. Furthermore, percentages of activated B-cells correlated with EBV-DNA load (rs=0.754; p<0.001). ConclusionsOverall, these findings indicate a strong association between HIV-1 viremia, markers of immune activation and EBV load and suggest that persistence of HIV-1 viremia and immune activation, regardless of peripheral CD4 cell depletion/repopulation, may favor expansion of EBV-infected cells and onset of EBV-related malignancies.

Liver injury in HIV-1-infected patients receiving non-nucleosides reverse transcriptase inhibitors-based antiretroviral therapy.

Liver injury is one of the most important adverse effects of antiretroviral therapy, leading to therapy changing or discontinuation. Data on liver injury in human immunodeficiency virus-1-infected patients receiving antiretroviral therapy are limited in China. The purpose of this study was to investigate the features of liver injury in human immunodeficiency virus type 1-infected patients receiving non-nucleosides reverse transcriptase inhibitors-based antiretroviral therapy in China. Seventy-five patients on antiretroviral therapy containing non-nucleosides reverse transcriptase inhibitors were retrospectively studied. The patients were divided into 2 groups: group 1 (with liver injury, n = 45) and group 2 (without liver injury, n = 30). The features of liver injury were analyzed. The sex, age, baseline CD4 counts, hepatitis B virus (HBV) and/or hepatitis C virus (HCV) co-infection, hepatotoxic drug use and nevirapine or efavirenz use were compared between two groups. Forty-five patients (60.0%), 31 (68.9%) males and 14 (31.1%) females, aged 12 to 52 years (averaged (39 ± 9) years), experienced at least one episode of liver injury. Forty (53.3%) patients were co-infected with HBV and/or HCV, 42 (56%) patients had concomitant use of antituberculosis drugs or cotrimoxazole, 46 (61.3%) and 29 (38.7%) patients received regimen containing nevirapine and efavirenz, respectively. Grade 1 liver injuries were observed in 26 (57.8%) patients, grade 2 in 16 (35.6%), grade 3 in 2 (4.0%) and grade 4 in 1 (2.2%). Three (6.7%) patients discontinued highly active antiretroviral therapy (HAART) due to liver injury. In group 1, there were 29 (64.4%) patients co-infected with HBV and/or HCV, 32 (71.1%) patients received regimen containing nevirapine, and 30 (66.7%) patients had concomitant use of anti-tuberculosis drugs or cotrimoxazole, respectively, significantly higher than those in group 2 (11 (36.7%), 14 (46.7%) and 12 (40%), respectively; P = 0.018, 0.033, 0.023, respectively). The sex, age, baseline CD4 counts and disease stage were not factors associated with liver injury. Liver injury associated with HAART containing non-nucleosides reverse transcriptase inhibitors was mild to moderate and those who were co-infected with HBV and/or HCV, had concomitant use of antituberculosis drugs or cotrimoxazole and received a regimen containing nevirapine were prone to liver injury while receiving HAART.

Neutralization activity in a geographically diverse East London cohort of human immunodeficiency virus type 1-infected patients: clade C infection results in a stronger and broader humoral immune response than clade B infection

The array of human immunodeficiency virus (HIV) subtypes encountered in East London, an area long associated with migration, is unusually heterogeneous, reflecting the diverse geographical origins of the population. In this study it was shown that viral subtypes or clades infecting a sample of HIV type 1 (HIV-1)-positive individuals in East London reflect the global pandemic. The authors studied the humoral response in 210 treatment-naïve chronically HIV-1-infected (>1 year) adult subjects against a panel of 12 viruses from six different clades. Plasmas from individuals infected with clade C, but also plasmas from clade A, and to a lesser degree clade CRF02_AG and CRF01_AE, were significantly more potent at neutralizing the tested viruses compared with plasmas from individuals infected with clade B. The difference in humoral robustness between clade C- and B-infected patients was confirmed in titration studies with an extended panel of clade B and C viruses. These results support the approach to develop an HIV-1 vaccine that includes clade C or A envelope protein (Env) immunogens for the induction of a potent neutralizing humoral response.

Structural Basis for the Role of the K65R Mutation in HIV-1 Reverse Transcriptase Polymerization, Excision Antagonism, and Tenofovir Resistance

K65R is a primary reverse transcriptase (RT) mutation selected in human immunodeficiency virus type 1-infected patients taking antiretroviral regimens containing tenofovir disoproxil fumarate or other nucleoside analog RT drugs. We determined the crystal structures of K65R mutant RT cross-linked to double-stranded DNA and in complexes with tenofovir diphosphate (TFV-DP) or dATP. The crystals permit substitution of TFV-DP with dATP at the dNTP-binding site. The guanidinium planes of the arginines K65R and Arg72 were stacked to form a molecular platform that restricts the conformational adaptability of both of the residues, which explains the negative effects of the K65R mutation on nucleotide incorporation and on excision. Furthermore, the guanidinium planes of K65R and Arg72 were stacked in two different rotameric conformations in TFV-DP- and dATP-bound structures that may help explain how K65R RT discriminates the drug from substrates. These K65R-mediated effects on RT structure and function help us to visualize the complex interaction with other key nucleotide RT drug resistance mutations, such as M184V, L74V, and thymidine analog resistance mutations.

Pharmacokinetic Assessment of Nevirapine and Metabolites in Human Immunodeficiency Virus Type 1-Infected Patients with Hepatic Fibrosis

Nevirapine is a nonnucleoside reverse transcriptase inhibitor used as part of combination therapy for human immunodeficiency virus (HIV) infection. Nevirapine may be prescribed for patients with hepatic fibrosis and cirrhosis. Significant autoinduction of cytochrome P450 3A4 and 2B6 following multiple dosing prompted an assessment of the metabolic profiles in patients with liver disease receiving chronic nevirapine therapy. HIV-infected patients with hepatic fibrosis who were receiving a stable antiretroviral regimen containing nevirapine for > or = 6 weeks had liver biopsy specimens assessed by Ishak histologic scoring and were grouped by severity (group 1, Ishak scores of 1 and 2; group 2, Ishak scores of 3 and 4; group 3, Ishak scores of 5 and 6). Steady-state trough nevirapine levels were determined for all patients, and additional measurements were obtained at 1, 2, and 4 h following nevirapine dosing for a subset of patients. The pharmacokinetics of nevirapine and its five metabolites were characterized, and a comparison of the results for the different Ishak groups was performed. Among 51 patients with hepatic fibrosis, the majority of whom were coinfected with hepatitis C virus or hepatitis B virus, differences between the maximum and the minimum observed plasma concentrations demonstrated a statistically significant flattening of the systemic exposure curves with progression from Ishak group 1 to Ishak group 2 or 3, suggesting a decrease in systemic clearance with the progression of liver disease. However, there were no significant differences in the trough and the maximum nevirapine concentrations between the Ishak groups. The metabolite profiles were also comparable across the Ishak groups. In HIV-infected patients who were chronically treated with nevirapine and who had various degrees of hepatic fibrosis, including cirrhosis, trough plasma nevirapine concentrations were not significantly increased, and thus, no dose adjustment is warranted.

Beneficial Effects of a Combination of Korean Red Ginseng and Highly Active Antiretroviral Therapy in Human Immunodeficiency Virus Type 1-Infected Patients

To determine whether Korean red ginseng (KRG) has beneficial effects on human immunodeficiency virus type 1 (HIV-1)-infected patients administered highly active antiretroviral therapy (HAART), we analyzed the CD4 T-cell count, viral load, and resistance mutations to HAART in 46 individuals. Thirteen patients harbored resistance mutations at baseline. The study population was divided into two groups: specifically, a group treated with a combination of HAART plus KRG (23 patients) and a group treated with HAART alone (23 patients). The annual increase in CD4 T-cell count in the combination group was significantly higher than that in the group treated with HAART alone (P < 0.05). Overall, 21 patients harbored resistance mutations after 3 years of therapy. Following exclusion of 13 patients displaying baseline resistance mutations, 7.1% of patients (1/14) in the combination group and 42.1% (8/19) in the HAART group were identified with resistance mutations. One patient with baseline resistance mutations in the combination group did not display resistance mutations 3 years after HAART therapy. High-level resistance mutations were significantly lower in the combination group than in the group treated with HAART alone. Five patients showed no improvement in viral copy number (26.3% [5/19]) in the combination group and 9 (45.0% [9/20]) showed no improvement in the HAART-only group. Our data support the clinical utility of KRG intake during HAART therapy.

Genotypic Characterization of CRF01_AEenvGenes Derived from Human Immunodeficiency Virus Type 1-Infected Patients Residing in Central Thailand

CRF01_AE is a major subtype of human immunodeficiency virus type 1 (HIV-1) circulating in Southeast Asia, including Thailand. HIV-1 env genes were amplified by polymerase chain reaction from blood samples of HIV-1-infected patients residing in Thailand in 2006, and cloned into the pNL4-3-derived reporter viral construct. Generated envelope protein (Env)-recombinant virus was examined for its infectivity, and then 35 infectious CRF01_AE Env-recombinant viruses were selected. Sequencing analysis revealed that the interclone variation of the deduced amino acid sequences was higher in CRF01_AE env genes isolated in 2006 than in those isolated in the early 1990s, suggesting that env gene variation has been increasing gradually among CRF01_AE viruses prevalent in Thailand. We also examined the characteristics of the deduced amino acid sequences of 35 CRF01_AE env genes. Our results may provide useful information to help in better understanding the genotype of env genes of CRF01_AE viruses currently circulating in Thailand.

Clinical significance of "anti-HBc alone" in human immunodeficiency virus-positive patients

To determine the prevalence and clinical relevance of isolated antibodies to hepatitis B core antigen as the only marker of infection ("anti-HBc alone") among human immunodeficiency virus (HIV) type-1 infected patients. Occult hepatitis B infection frequency was also evaluated. Three hundred and forty eight histories from 2388 HIV-positive patients were randomly reviewed. Patients with serological markers of hepatitis B virus (HBV) infection were classified into three groups: past hepatitis, "anti-HBc alone" and chronic hepatitis. Determination of DNA from HBV, and RNA and genotype from hepatitis C virus (HCV) were performed on "anti-HBc alone" patients. One hundred and eighty seven (53.7%) HIV-positive patients had markers of HBV infection: 118 past infection (63.1%), 14 chronic hepatitis (7.5%) and 55 "anti-HBc alone" (29.4%). Younger age [2.3-fold higher per every 10 years younger; 95% confidence intervals (CI) 1.33-4.00] and antibodies to HCV infection [odds ratio (OR) 2.87; 95% CI 1.10-7.48] were factors independently associated with the "anti-HBc alone" pattern. No differences in liver disease frequency were detected between both groups. Serum levels of anti-HBs were not associated with HCV infection (nor viral replication or HCV genotype), or with HIV replication or CD4 level. No "anti-HBc alone" patient tested positive for HBV DNA. "Anti-HBc alone" prevalence in HIV-positive patients was similar to previously reported data and was associated with a younger age and with antibodies to HCV infection. In clinical practice, HBV DNA determination should be performed only in those patients with clinical or analytical signs of liver injury.

Blepharoptosis and External Ophthalmoplegia Associated with Long‐Term Antiretroviral Therapy

Long-term antiretroviral therapy (ART) is associated with lipodystrophy, peripheral neuropathy, lactic acidosis, and myopathy. Blepharoptosis, without prior ART association, is usually caused by age-associated involutional ptosis, but it is also seen in mitochondrial myopathies with external ophthalmoplegia, cardiac conduction disturbances, and neurological impairments. Patients presented over a 2-year period. Four patients underwent surgical blepharoptosis repair. Five human immunodeficiency virus type 1-infected patients (median age, 50 years; range, 46-53 years) who were receiving ART presented with severe blepharoptosis; 2 of these 5 also presented with external ophthalmoplegia. Findings included decreased palpebral fissure height (median, 6.5 mm; normal height, 9 mm), mildly impaired levator function (median, 10 mm; normal, >13 mm), and markedly decreased marginal reflex distance (median, 0.5 mm; normal, 4 mm). A greater advancement of the levator aponeurosis was required during surgical repair, a finding consistent more with myogenic than with involutional blepharoptosis. All patients had severe lipodystrophy, which preceded blepharoptosis by a median interval of 4.7 years (range, 2.8-5.7 years). Four patients also presented with peripheral neuropathy and metabolic abnormalities before the onset of blepharoptosis, and 3 had cardiac conduction disturbances. Patients received ART for a median of 7.8 years (range, 4.9-11.2 years), thymidine analogue-containing ART for a median of 7.1 years (range, 1.2-7.9 years), and protease inhibitor-containing ART for a median of 7.1 years (range, 4.9-8.9 years). We report the novel findings of blepharoptosis and external ophthalmoplegia in patients who are receiving ART. Ptosis was preceded by lipodystrophy with long-term use of both thymidine-analogue- and protease inhibitor-containing ART. The findings are most consistent with myogenic ptosis in a generalized mitochondrial myopathy syndrome. Clinicians should also be watchful for other potential myopathic ptosis-associated complications, including proximal weakness, dysphagia, deafness, and cardiac conduction disturbances.

Imported Malaria in HIV-Infected Patients Enrolled in the ANRS CO4 FHDH Study

To describe episodes of imported malaria in human immunodeficiency virus type 1-infected patients and to study the risk factors for severe Plasmodium falciparum malaria. Patients enrolled in the French Hospital Database on HIV who were diagnosed with a first episode of malaria between 1996 and 2003 were included. The severity of P. falciparum imported malaria was graded with World Health Organization criteria. Geographic areas were classified according to P. falciparum chemoresistance. Risk factors for severe malaria were identified with logistic regression. We studied 190 patients infected by P. falciparum in 178 cases. All but four of the patients were infected in sub-Saharan Africa, and half were returning from a country with a high P. falciparum chloroquine resistance. Their median age was 37.5 years, and 57% came from a country endemic with malaria. The median CD4 cell count was 299/mm, and the median plasma human immunodeficiency virus type 1 RNA load was 4.5 log10 copies/mL. Sixty-five (36.5%) episodes of P. falciparum malaria were severe. Severe imported malaria was associated with CD4 cells/mm <350 (odds ratio = 2.58; 95% confidence interval: 1.19 to 5.57). The risk of severe malaria was lower in patients returning from a country with a high prevalence of chemoresistance (odds ratio = 0.50; 95% confidence interval: 0.25 to 0.99). Severe imported malaria in human immunodeficiency virus type 1-infected patients is associated with decreased CD4 cell count. The risk seems lower when P. falciparum infection was acquired in areas of high prevalence of chemoresistance.

Development of an Immunofluorescence Assay Using Recombinant Proteins Expressed in Insect Cells To Screen and Confirm Presence of Human Herpesvirus 8-Specific Antibodies

Human herpesvirus 8 (HHV-8), or Kaposi's sarcoma (KS)-associated herpesvirus, has been linked to all forms of KS. The results of most current serological assays for the detection of HHV-8-specific antibodies have low levels of concordance among themselves. To establish a sensitive and specific testing strategy that can be used to screen for HHV-8-specific antibodies, three HHV-8 proteins, ORF65, ORF73, and K8.1A, were expressed by using baculoviral vectors in insect cells and incorporated into a monoclonal antibody-enhanced immunofluorescence assay (mIFA) termed the Sf9 three-antigen mIFA. The results obtained by this mIFA were compared to those obtained by a standard mIFA with an HHV-8-infected B-cell line (BC3 mIFA). Test sera were obtained from patients diagnosed with KS, human immunodeficiency virus type 1-infected patients at high risk for HHV-8 infection, and healthy controls from a local blood bank. The combined use of both assays had a sensitivity of 94% and a specificity of 96%. The performance of these two assays when they were used together indicates that they may be useful for the reliable detection of HHV-8-specific immunoglobulin G antibodies in a population.

Maintaining Antiretroviral Therapy Reduces the Risk of AIDS-Defining Events in Patients with Uncontrolled Viral Replication and Profound Immunodeficiency

The benefits of continuing antiretroviral therapy are questionable in human immunodeficiency virus (HIV) type 1-infected patients with profound immunodeficiency and multiple treatment failure due to viral resistance. From the French Hospital Database on HIV, we selected 12,765 patients with a CD4(+) cell count <200 cells/mm(3) who received a combination antiretroviral therapy (cART) during 2000-2005. Three groups of patients were defined: patients who interrupted cART at least once, patients who had at least 2 consecutive detectable viral loads (VLs) while receiving cART, and patients who had undetectable VL during treatment with cART. Incidence rates and risks of new acquired immunodeficiency syndrome-defining events (ADEs) were assessed among the 3 groups of patients, overall and after CD4(+) cell count stratification (<50 and 50-200 cells/mm(3)). The estimated incidence rates +/- standard deviation of ADEs were 18.5+/-1.9, 14.5+/-0.7, and 4.9+/-0.5, respectively, for patients who interrupted cART, patients who had detectable VL during treatment with cART, and patients who had undetectable VL during treatment with cART. These differences were observed in both CD4(+) cell count strata. Overall, after adjustment, risks of a new ADE in patients who had detectable VL and in patients who had undetectable VL while receiving cART were 22% and 62% lower, respectively, than in patients who stopped cART. Among patients with CD4(+) cell count <50 cells/mm(3), the risk of a new ADE was 22% lower in patients who continued to receive a failing cART regimen than in patients who stopped treatment with cART. Likewise, among patients with a CD4(+) cell count of 50-200 cells/mm(3), the risk was 34% lower in patients who continued to receive a failing cART regimen than in those who stopped taking cART. Even when effective virological control is no longer achievable, cART still reduces the risk of ADEs in profoundly immunodeficient HIV-infected patients.

Evaluation of Human Immunodeficiency Virus Type 1-Infected Chinese Patients Treated with Highly Active Antiretroviral Therapy for Two Years

The purpose of this study was to evaluate the long-term efficacy and safety of nevirapine in combination with didanosine and stavudine in the treatment of human immunodeficiency virus (HIV)1-infected Chinese patients in routine clinical practice. The study, from April 2003 to May 2005, with follow-up through 24 mo, was conducted at the Department of Infectious Diseases, Second Xiangya Hospital, Central-South University in Changsha, Hunan Province, China. Twenty-seven HIV1-infected patients received didanosine, stavudine, and nevirapine. Information from case notes regarding age, sex, side effects, viral load, naive and memory T cells, and CD4(+) and CD8(+) T cell count at baseline, 3, 6, 12, 18, and 24 mo was collected and analyzed. Virologic suppression, defined as an HIV RNA concentration of less than 50 copies/mL at months 3, 6, 12, 18, and 24, was considered the main outcome measure. Of 27 patients, 17 were men with a mean age 33.5 yr. The mean baseline viral load was 5.15 log copies/mL and the mean CD4(+) cell count was 185 cells/dL. Of 27 patients, 3 patients discontinued study medication; treatment was changed, because of side effects, from didanosine (ddI), stavudine (d4T), and nevirapine (NVP) to zidovudine, lamivudine, and NVP for 24 patients who had completed 24 mo of treatment with ddI, d4T, and NVP; and viral load suppression was attained in 17 patients (70.8%) at 12 mo, in 14 patients (58.3%) at 18 mo, and in 13 patients (56.6%) at 24 mo. The CD4 T cell count increased by 114 cells/microL (mean, 299 cells/microL) after 12 mo of treatment and by 132 cells/microL (mean, 317 cells/microL) after 24 mo of treatment. Naive T cells and memory cells also increased in number, but at a slower rate. Activated (CD38(+)) CD8(+) T cells were elevated at baseline (67.7%) and declined by month 24 (49.7%), but did not reach normal levels. We conclude that a regimen of NVP with ddI and d4T provided durable suppression of plasma viral load in HIV-infected patients, with significant improvement in the CD4 cell count, and can be well tolerated by patients with HIV-1 infection.

Role of the Envelope Genetic Context in the Development of Enfuvirtide Resistance in Human Immunodeficiency Virus Type 1-Infected Patients

Acquired human immunodeficiency virus type 1(HIV-1) resistance to the fusion inhibitor enfuvirtide (ENF) is primarily associated with mutations within the highly conserved first heptad repeat (HR1) region of gp41. Viral env sequences, however, are remarkably variable, and the envelope genetic background could have an important impact on optimal expression of HR1 mutations. We have examined the genetic evolution of env sequences, ENF susceptibility, and Env replicative capacity in patients failing ENF treatment. Sequential plasma-derived virus populations, obtained from six patients initiating ENF treatment as part of a salvage therapy, were studied using a recombinant phenotypic assay evaluating the entire gp120 and the gp41 ectodomains. Regardless of major differences in the baseline ENF susceptibilities, viral populations with similar phenotypic ENF resistance (50% inhibitory concentration, >3,000 ng/ml) were selected under treatment in four of six patients. As expected, in all patients ENF-resistant viruses harbored one or more HR1 mutations (positions 36, 38, and 43). Interestingly, in five patients the emergence of resistance mutations was not associated with reduced Env replicative capacity. Phylogenetic analysis of env sequences in sequential samples from two patients showed that the HR1 mutations had emerged in the context of env quasi-species that were different from those prevalent at baseline. Thus, the envelope genetic context appears to play a critical role in the selection of HR1 mutations and the expression of ENF resistance, thereby conditioning the evolution of HIV-1 under fusion inhibitor selective pressure.

Viral Suppression and Immune Restoration in the Gastrointestinal Mucosa of Human Immunodeficiency Virus Type 1-Infected Patients Initiating Therapy during Primary or Chronic Infection

Although the gut-associated lymphoid tissue (GALT) is an important early site for human immunodeficiency virus (HIV) replication and severe CD4+ T-cell depletion, our understanding is limited about the restoration of the gut mucosal immune system during highly active antiretroviral therapy (HAART). We evaluated the kinetics of viral suppression, CD4+ T-cell restoration, gene expression, and HIV-specific CD8+ T-cell responses in longitudinal gastrointestinal biopsy and peripheral blood samples from patients initiating HAART during primary HIV infection (PHI) or chronic HIV infection (CHI) using flow cytometry, real-time PCR, and DNA microarray analysis. Viral suppression was more effective in GALT of PHI patients than CHI patients during HAART. Mucosal CD4+ T-cell restoration was delayed compared to peripheral blood and independent of the time of HAART initiation. Immunophenotypic analysis showed that repopulating mucosal CD4+ T cells were predominantly of a memory phenotype and expressed CD11 alpha, alpha(E)beta 7, CCR5, and CXCR4. Incomplete suppression of viral replication in GALT during HAART correlated with increased HIV-specific CD8+ T-cell responses. DNA microarray analysis revealed that genes involved in inflammation and cell activation were up regulated in patients who did not replenish mucosal CD4+ T cells efficiently, while expression of genes involved in growth and repair was increased in patients with efficient mucosal CD4+ T-cell restoration. Our findings suggest that the discordance in CD4+ T-cell restoration between GALT and peripheral blood during therapy can be attributed to the incomplete viral suppression and increased immune activation and inflammation that may prevent restoration of CD4+ T cells and the gut microenvironment.

Development and validation of a quantitative assay for the measurement of two HIV-fusion inhibitors, enfuvirtide and tifuvirtide, and one metabolite of enfuvirtide (M-20) in human plasma by liquid chromatography–tandem mass spectrometry

A method for the quantification of two peptide HIV-1 fusion inhibitors (enfuvirtide, T-20 and tifuvirtide, T-1249) and one metabolite of enfuvirtide (M-20) in human plasma has been developed and validated, using liquid chromatography coupled with electrospray tandem mass spectrometry (LC-MS/MS). The analytes were extracted from plasma by solid-phase extraction (SPE) on vinyl-copolymer cartridges. Chromatographic separation of the peptides was performed on a Symmetry 300 C(18) column (50mmx2.1mm I.D., particle size 3.5 microm), using a water-acetonitrile gradient containing 0.25% (v/v) formic acid. The triple quadrupole mass spectrometer was operated in the positive ion-mode and multiple reaction monitoring (MRM) was used for peak detection. Deuterated (d60) enfuvirtide and (d50) tifuvirtide were used as internal standards. The assay was linear over a concentration range of 20-10,000 ng/ml for enfuvirtide and tifuvirtide and of 20-2000 ng/ml for M-20. Intra- and inter-assay precisions and deviations from the nominal concentrations were </=13%. Stability of the analytes was tested under all relevant conditions for sample handling. The method was capable to measure concentrations of enfuvirtide and its metabolite in plasma samples of human immunodeficiency virus type-1 (HIV-1) infected patients treated with the drug.