Acquired immunodeficiency syndrome (AIDS) is a life-threating disease caused by human immuno-deficiency virus type 1 (HIV). The advent of highly active antiretroviral therapy (HAART) has resulted in profound suppression of HIV replication, substantial increase of CD4+ T cells and partial reconstitution of the immune system.1 US and European guidelines released in December 2007 recommend treating patients who have CD4+ counts <350 cells/μl.2,3 The effect of baseline CD4+ cell counts on efficacy of HAART has been extensively studied.4–7 Initiating HAART in patients with higher CD4+ cell counts may result in higher total CD4+ cell counts and more-durable virological suppression.4–9 Several cohort studies have observed that patients who started antiretroviral therapy (ART) with CD4+ cell counts <350 cells/μl were less likely to achieve normal levels.10,11 To our knowledge, there were few data about the impact of CD4+ T cell strata on HAART in the treatment-naive Chinese patients. In China, the most popular regimen of HAART is nevirapine-containing regimens. Therefore we conducted a prospective analysis of a large clinical cohort of Chinese HIV-infected treatment-naive patients grouped by the baseline CD4+ T cell counts.
This study was a prospective study conducted at thirteen research centers of China. This study was approved by the Ethics Committee of Peking Union Medical College Hospital. Enrollment began in January 2005. Patient Information and Consent to Medical Treatment was signed in a written consent form. Patients were found to be HIV-seropositive by standard serum enzyme-linked immunosorbent assay (ELISA) and also by Western blotting analysis. Patients were considered for inclusion in this study if they were antiretroviral-naive. The eligibility criteria for participants were CD4+ T-cell count from 100 cells/μl to 350 cells/μl and plasma viral load over 500 copies/ml. Main exclusive criteria were pregnancy or breastfeeding, anticipated nonadherence, AIDS-defining illness within 2 weeks of entry, white blood cell count less than 2.0×109/L, absolute neutrophil count less than 1.0×109/L, hemoglobin level less than 90 g/L, platelet count less than 0.75×1012/L, transaminase and alkaline phosphatase level more than 3 times the upper limit of the normal range, bilirubin level more than 2.5 times the upper limit of the normal range, serum creatinine level more than 1.5 times the upper limit of the normal range. Patients were randomized into 3 initial HAART strategies: stavudine (d4T, Desano, China) + lamivudine (3TC, GlaxoSmithKline, UK), or zidovudine (AZT, Northeast General Pharmaceutical Factory, China) + 3TC, or AZT + didanosine (ddI, Northeast General Pharmaceutical Factory, China) combined with nevirapine (NVP, Desano, China). They are all generic drugs except for 3TC. Patients were grouped by the baseline CD4+ cell count strata defined as 100-200 cells/μl, 201-350 cells/μl respectively. Patients were also stratified by baseline CD4+ T cell ≤250 cells/μl or CD4+ T cell >250 cells/μl in the analysis of adverse effects. The compliance of antiviral medicine utilization was good in all the patients.
Viral load measurements
Plasma was separated from whole blood by centrifugation within 4 hours of collection and was stored frozen at −80°C until tested. The QUANTIPLEX HIV-1 RNA assay, version 3.0 (bDNA 3.0 assay), was performed according to the manufacturer's instructions that were provided with the assay kit. The limits of detection of the assay, indicated by the manufacturer, were 50-500 000 HIV-1 RNA copies/ml.
Immunofluorescent surface staining and flow cytometric analysis
Peripheral blood mononuclear cells were obtained by separation from the centrifugation gradient. Subpopulations of CD3+, CD4+, and CD8+ cells were determined by three-color flow cytometry (Beckman-Coulter, USA) at baseline and at the end of weeks 4, 12, 24, 36, 52, 68, 84, and 100. The following groups of monoclonal antibodies were used: PEcy5-CD4/PE-CD8/FITC-CD3 (CD4+/CD8+ T cell counts). All monoclonal antibodies were purchased from Beckman-Coulter and Immunotech, USA.
Blood routines, liver functions, and serum lipids were administered by the clinical laboratory department of each research centers.
The study persisted two-year follow-up. The follow-up period was 8 visits at weeks 4, 12, 24, 36, 52, 68, 84, and 100. During all of the visits, the clinical assessment was recorded in the case report form (CRF) and the subjects had samples taken for laboratory assessment and further detection including T cell subset, plasma viral load (pVL). The adverse events were classified and graded according to division of AIDS table for grading the severity of adult and pediatric adverse events.12
All statistical analyses were performed using the SPSS 10.0 statistical package. To compare viral load and lymphocyte T cells during the follow-up, independent-sample t test were used. The viral load curve was denoted by mean and standard deviation. CD4+ T cell curve was expressed by median, 25th and 75th percentiles. Using a chi-square test, we compared the rates of response among the treatment groups in terms of the percentage of patients with suppression of plasma HIV-1 RNA levels less than 400 copies/ml as well as the percentage for whom it was less than 50 copies/ml. For all tests, P <0.05 was considered statistically significant.
Intention to treat (ITT) analysis (sometimes also called intent to treat) is an analysis based on the initial treatment intent, not on the treatment eventually administered. It is intended to avoid various misleading artifacts that can arise in intervention research. Per-protocol (PP) analysis is in contrast to the ITT analysis. It is a analysis strategy in which only patients who complete the entire clinical trial or other procedure analyzed, not like the ITT analysis which also includes those patients dropped out. Per-protocol analysis is also called “on treatment analysis”.
Of the 362 subjects screened, 198 HIV-positive antiretroviral-naive patients were recruited into this study. Demographic characteristics of the groups are summarized in Table 1. There were no significant differences between the two groups at baseline characteristics except for baseline CD4+ T cell count and HIV RNA viral load. The baseline HIV RNA viral loads were significantly lower in the 201-350 cells/μl group.
The changes of pVL during 100-week HAART were shown in Figure 1. In ITT analysis and PP analysis, there were no significant differences between the two groups in the plasma viral loads except for the baseline counts. There were significantly decreases in the first 12 weeks in the two groups. And there were no differences in the viral response rate in terms of the percentage of pVL less than 50 copies/ml or less than 400 copies/ml between the two groups, which were ITT analysis and showed in Figure 2. The results of viral response rate in PP analysis which were not shown were similar to that in ITT analysis. At the endpoint of study, the percentage of patients who achieved pVL <50 copies/ml was 60.34% in CD4+ 100-200 cells/μl group, 49.23% in CD4+ 201-350 cells/μl group (P=0.603). The percentage of patients who achieved pVL <400 copies/ml was 82.46% in 100-200 cells/μl group, 86.15% in 201-350 cells/μl group (P=0.217).
In ITT analysis and PP analysis, the CD4+ T cell counts were statistically higher in the 201-350 cells/μl group during the follow-ups, although CD4+ cell count increases were similar in different CD4+ cell strata. By 100 weeks, the median CD4+ cell count was 331 cells/μl in the CD4+ 100-200 cells/μl group, 462 cells/μl in the CD4+ 201-350 cells/μl group, respectively. The significant increases of CD4+ T cell were seen in the first 12 weeks in each group. The increases of CD4+ cell count were respectively 79 cells/μl and 55 cells/μl at the week 4, 108 cells/μl and 100 cells/μl respectively at the week 12, 202 cells/μl and 205 cells/μl respectively at the week 100.
Outcomes and adverse events
The incidence rate of nausea was 15/86 (17.4%), 33/112 (30.4%) in the lower CD4 group and higher CD4 group, respectively. The prevalence of nausea in the CD4+ 201-350 cells/μl group was significantly higher than the other group (P=0.05).
There were not significant differences in other adverse reactions such as rash, hair loss, neutropenia, thrombocytopenia, hyperlipidemia, anorexia and abnormal liver function test. Liver function abnormalities were further analyzed stratified by baseline CD4+ T cell ≤250 cells/μl or >250 cells/μl and sex. The prevalence of liver function abnormalities had no significant difference in female or male in the CD4 T cell ≤250 cells/μl group and >250 cells/μl group (the results were not shown).
The introduction of HAART has been effective in decreasing morbidity and mortality rates due to HIV infection in the developed countries; similar rate reductions have also been observed in HIV patients treated with HAART in developing countries.13,14 The CD4+ T cell count is the most significant predictor of HIV clinical disease progression and survival. To evaluate the influence of baseline CD4+ T cell count on HAART, we analyzed 198 treatment-naive Chinese HIV-infected patients and followed up for nearly two years.
In this 2-year follow-up study, there were no significant differences in the pVL except for the baseline counts. Significant decreases of viral loads happened in the first 12 weeks without relations with baseline CD4+ T cell counts. There were no differences in the viral response rate of the two groups. The CD4+ T cell counts were statistically higher in the 201-350 cells/μl group during the follow-ups, but the increase of CD4+ T cell count were similar in different CD4+ cell strata. The significant increase of CD4+ T cell was seen in the first 12 weeks for any group. The prevalence of nausea in the CD4+ 201-350 cells/μl group was slightly higher. The prevalence of liver function abnormalities had no significant difference in female or male in the CD4 T cell ≤250 cells/μl group and 250 cells/μl group.
The important finding of this study is that viral loads and viral response rates have no relationship with the baseline CD4+ counts for two years. The rapidly decrease of viral load at the beginning is irrespective of baseline CD4+ T cell count. The results were different form the former studies which demonstrated the viral response rate was lower in the low baseline CD4+ T cell counts.9 Maybe the response of viral inhibition of Chinese patients is different. Further study and large samples should be considered.
In this study, CD4+ cell counts for patients in the lower strata remained lower, as in prior reports,8,10,15 patients in the lower strata did not achieve levels of those who started with higher CD4+ cell counts, even after nearly 2-year HAART. Immunological reconstitution is typically measured by circulating CD4+ cell counts, which follow a biphasic pattern: an initial rapid increase during the first few months of HAART, followed by a slower increase.16–20 There was significantly increase during the first 12 weeks, which is consistent with the literature. However CD4+ cell count increases were similar regardless of baseline CD4+ stratum which is different from the studies before.21 In a study performed in the EuroSIDA cohort,21 both older age and a lower CD4+ cell count were associated with a lower increase in the CD4+ cell count at 12 months of HAART. Maybe Chinese patients have their specialities, so further large Chinese group are necessary to make more clear.
This study is difference from the former reports about the incidence of adverse events stratified by baseline CD4 cell count. In other studies, the prevalence of rash and liver function abnormalities is higher when started at a higher CD4 cell count.22,23 whereas increased prevalence of nausea happened in the CD4+ 201-350 cells/μl groups in our study, whereas there are no differences in rash or abnormal liver function in female stratified by CD4+ 250 cells/μl. The results should be evaluated cautiously. Nausea is tolerable and no patients are ended therapy because of nausea. The mechanism of higher prevalence of nausea in higher baseline CD4+ T cell is unclear, which need further study.
An important caveat to our results is that we do not have data on the specific T lymphocyte subsets in this cohort. And we did not grouped by different treatment regimens. The sample is small in our study and need further larger sample. There was higher missing rate (38.4%) during follow-up. Two-year follow-up maybe is shorter. The treatment efficacy, adverse drug reaction and drug resistence of HAART need longer time evaluation due to AIDS treatment all the life.
In summary, patients who initiated HAART with a lower CD4+ cell count never achieve CD4+ cell counts that are comparable to those of patients who initiated treatment at CD4+ cell counts 201-350 cells/μl. We recommend that consideration be given to initiation of HAART at a CD4+ cell count 201-350 cells/μl to achieve better immune recovery. The viral suppression has no correlation with the baseline CD4+ cell count. Maybe higher CD4+ can increase the risk of nausea in the adverse events of HAART.
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Keywords:© 2009 Chinese Medical Association
human immunodeficiency virus; acquired immunodeficiency syndrome; highly active antiretroviral therapy; CD4+ T cell counts; nevirapine