Identification of dynamically distinct subpopulations of T lymphocytes that are differentially affected by HIV.

We examined the effects of human immunodeficiency virus infection on the turnover of CD4 and CD8 T lymphocytes in 17 HIV-infected patients by 30 min in vivo pulse labeling with bromodeoxyuridine (BrdU). The percentage of labeled CD4 and CD8 T lymphocytes was initially higher in lymph nodes than in blood. Labeled cells equilibrated between the two compartments within 24 h. Based on mathematical modeling of the dynamics of BrdU-labeled cells in the blood, we identified rapidly and slowly proliferating subpopulations of CD4 and CD8 T lymphocytes. The percentage, but not the decay rate, of labeled CD4 or CD8 cells in the rapidly proliferating pool correlated significantly with plasma HIV RNA levels for both CD4 (r = 0.77, P < 0.001) and CD8 (r = 0.81, P < 0.001) T cells. In six patients there was a geometric mean decrease of greater than 2 logs in HIV levels within 2 to 6 mo after the initiation of highly active antiretroviral therapy; this was associated with a significant decrease in the percentage (but not the decay rate) of labeled cells in the rapidly proliferating pool for both CD4 (P = 0.03) and CD8 (P < 0.001) T lymphocytes. Neither plasma viral levels nor therapy had an effect on the decay rate constants or the percentage of labeled cells in the slowly proliferating pool. Monocyte production was inversely related to viral load (r = -0.56, P = 0.003) and increased with therapy (P = 0.01). These findings demonstrate that HIV does not impair CD4 T cell production but does increase CD4 and CD8 lymphocyte proliferation and death by inducing entry into a rapidly proliferating subpopulation of cells.

CD4 T cell expansions are associated with increased apoptosis rates of T lymphocytes during IL-2 cycles in HIV infected patients.

OBJECTIVE AND DESIGN: In an attempt to determine the mechanisms underlying the CD4 T cell expansions in patients receiving intermittent interleukin (IL)-2, a cohort of 10 HIV infected patients were studied during a 5-day cycle of IL-2 to measure rates of apoptosis, the expression of activation markers in CD4 and CD8 T cell subsets and the serum levels of proinflammatory cytokines. All patients were receiving highly active antiretroviral therapy. METHODS: Peripheral blood mononuclear cells were tested pre- and at the completion of IL-2 treatment with annexin V/7-AAD for the measurement of apoptosis. Phenotypic analyses of T lymphocytes were performed in parallel. Serum levels of interferon (IFN)gamma, granulocyte-macrophage colony stimulating factor, IL-6 and tumor necrosis factor (TNF)alpha were tested by enzyme-linked immunosorbent assay. RESULTS: IL-2 increased the spontaneous apoptosis rates of CD4 and CD8 T lymphocytes (P = 0.003). Expression of HLA-DR, CD38 and CD95 increased on both CD4 and CD8 T lymphocytes whereas CD25 induction was observed exclusively on CD4 T cells. Significant increases of serum IL-6 and TNFalpha levels were noted in all patients whereas viral loads remained unchanged. CONCLUSION: Administration of IL-2 for 5 days in HIV infected patients leads to enhanced apoptosis of both CD4 and CD8 T cells despite an eventual increase of the CD4 T cell count. A profound activation state with induction of activation markers on T cells and high levels of TNFalpha and IL-6 accompanies the increased apoptosis during the IL-2 cycle. These data suggest that the CD4 expansions seen in the context of intermittent IL-2 therapy are the net result of increases in both cell proliferation and cell death.

Immunotherapy of HIV-infected patients with intermittent interleukin-2: effects of cycle frequency and cycle duration on degree of CD4(+) T-lymphocyte expansion.

The ability of IL-2 to induce expansion of the CD4(+) T lymphocyte pool has made it the most studied cytokine in the treatment of HIV infection. The majority of trials have used an empirical regimen of 5-day IL-2 cycles given every 8 weeks--a regimen based upon early pharmacodynamic studies and patient preference. To better define optimal duration and frequency of cycles, a randomized trial was conducted in which patients who received this "standard" regimen were compared to patients who received cycles of variable duration (based on individual patterns of cell cycle progression) and to patients who received cycles of variable frequency (based on individual CD4(+) T lymphocyte responses to previous cycles). Twenty-two patients with HIV-1 infection and CD4(+) T lymphocyte counts > 200 cells/mm(3) were randomized to one of three treatment groups for 32 weeks of study. Eight participants received four 5-day IL-2 cycles (controls) every 8 weeks; 7 participants received four cycles of longer duration (mean 7.7-days); and 7 participants received an increased frequency of 5-day cycles (every 4.1 weeks on average). All three groups experienced significant increases in mean CD4(+) T lymphocytes. However, there were no statistically significant differences in CD4(+) T lymphocyte increases between the group that received longer cycles (median increase 239 cells/mm(3), P = 0.78) or between the group that received more frequent cycles (median increase 511 cells/mm(3), P = 0.54) and the control group (median 284 cells/mm(3)). HIV-1 viral loads decreased during the study period in all three groups. Our inability to demonstrate a significant advantage of increased frequency or duration of IL-2 administration provides corroborating experimental evidence for the use of an IL-2 regimen consisting of 5-day cycles administered no more frequently than every 8 weeks in future clinical trials aimed at expanding the CD4(+) T lymphocyte pool.

CD40 ligand trimer and IL-12 enhance peripheral blood mononuclear cells and CD4+ T cell proliferation and production of IFN-gamma in response to p24 antigen in HIV-infected individuals: potential contribution of anergy to HIV-specific unresponsiveness.

It has been suggested that CD4+ T cell proliferative responses to HIV p24 Ag may be important in the control of HIV infection. However, these responses are minimal or absent in many HIV-infected individuals. Furthermore, while in vitro and in vivo responses to non-HIV recall Ags improve upon administration of highly active antiretroviral therapy, there does not appear to be a commensurate enhancement of HIV-specific immune responses. It is possible that CD4+ p24-specific T cells are deleted early in the course of infection. However, it is also possible that a discrete unresponsiveness, or anergy, contributes to the lack of proliferation to p24. To evaluate the possible contribution of unresponsiveness to the lack of CD4+ T cell proliferation to p24 in HIV-infected individuals, we attempted to overcome unresponsiveness. CD40 ligand trimer (CD40LT) and IL-12 significantly increased PBMC and CD4+ T cell proliferative responses to p24 Ag in HIV-infected, but not uninfected, individuals. No increase in proliferative response to CMV Ag was observed. CD40LT exerted its effect through B7-CD28-dependent and IL-12- and IL-15-independent mechanisms. Finally, the increase in proliferation with CD40LT and IL-12 was associated with an augmented production of IFN-gamma in most, but not all, individuals. These data suggest the possible contribution of HIV-specific unresponsiveness to the lack of CD4+ T cell proliferation to p24 Ag in HIV-infected individuals and that clonal deletion alone does not explain this phenomenon. They also indicate the potential for CD40LT and IL-12 as immune-based therapies for HIV infection.

Hepatitis G virus and human immunodeficiency virus coinfection: response to interferon-alpha therapy.

The prevalence and consequences of hepatitis G virus (HGV) infection were determined in 180 patients with human immunodeficiency virus (HIV) infection (predominantly male homosexuals) who participated in a trial that compared treatment with zidovudine versus interferon (IFN)-alpha versus the combination. HGV RNA levels were measured by branched DNA signal amplification assay. Initially, 66 (37%) had HGV RNA. Sexual transmission was the sole risk factor for infection in all but 4 subjects. Pretreatment clinical features were similar between HGV RNA-positive and -negative patients. After 6 months, only 5% treated with zidovudine became HGV RNA negative, compared with 95% who received IFN-alpha alone and 66% on combination therapy with low-dose IFN-alpha. After therapy, HGV RNA levels returned to baseline in most subjects. Thus, HGV infection is common among HIV-infected homosexual males but does not appear to influence clinical features in early HIV infection. HGV RNA levels are suppressed by IFN but not by zidovudine.

Peripheral expansion of pre-existing mature T cells is an important means of CD4+ T-cell regeneration HIV-infected adults.

The CD4+ T-cell pool in HIV-infected patients is in a constant state of flux as CD4+ T cells are infected and destroyed by HIV and new cells take their place. To study T-cell survival, we adoptively transferred peripheral blood lymphocytes transduced with the neomycin phosphotransferase gene between syngeneic twin pairs discordant for HIV infection. A stable fraction of marked CD4+ T cells persisted in the circulation for four to eighteen weeks after transfer in all patients. After this time there was a precipitous decline in marked cells in three of the patients. At approximately six months, marked cells were in lymphoid tissues in proportions comparable to those found in peripheral blood. In two patients, the proportion of total signal for the transgene (found by PCR analysis) in the CD4/CD45RA+ T-cell population relative to the CD4/CD45RO+ population increased in the weeks after cell infusion. These findings indicate that genetically-marked CD4+ T cells persist in vivo for weeks to months and that the CD4+ T-cell pool in adults is maintained mostly by the division of mature T cells rather than by differentiation of prethymic stem cells. Thus, after elements of the T-cell repertoire are lost through HIV infection, they may be difficult to replace.

Randomized, controlled phase I/II, trial of combination therapy with delavirdine (U-90152S) and conventional nucleosides in human immunodeficiency virus type 1-infected patients.

Delavirdine mesylate (DLV) is a potent nonnucleoside reverse transcriptase inhibitor with activity specific for human immunodeficiency virus type 1. In the present phase I/II study we evaluated the safety, toxicity, pharmacokinetics, and antiretroviral activities of two-drug and three-drug combinations of DLV and conventional doses of nucleoside analogs compared with those of both DLV monotherapy and two-drug nucleoside analog therapy. A total of 85 human immunodeficiency virus type 1 infected patients with CD4 counts of 100 to 300 cells per mm3 were enrolled in two periods: in the first period patients were randomized to receive either zidovudine (ZDV) plus didanosine (group 1) or ZDV plus didanosine plus escalating doses (400 to 1,200 mg/day) of DLV (group 2). In the second period, patients were randomized to receive either 1,200 mg of DLV alone per day (group 3) or ZDV plus 1,200 mg of DLV per day (group 4). DLV demonstrated good oral bioavailability at all five doses tested. The major toxicity was a transient mild rash which appeared in 44% of all DLV recipients. Overall, group 2 patients demonstrated more sustained improvements in CD4 counts, percent CD4 cells, branched DNA levels, p24 antigen levels, and virus titers in plasma than group 1, 3, or 4 patients. The magnitude of the response correlated with the intensity of prior nucleoside analog treatment, the non-syncytium-inducing or syncytium-inducing viral phenotype at baseline, and the presence of a wild-type codon at amino acid position 215 in the baseline reverse transcriptase genotype. Despite a transient rash, DLV therapy was well tolerated. Combination therapy with DLV and nucleoside analogs appears promising, with the three-drug combination appearing to be more potent that either two-drug combinations or monotherapy.

Use of recombinant soluble CD4 Pseudomonas exotoxin, a novel immunotoxin, for treatment of persons infected with human immunodeficiency virus.

Single and multiple doses of sCD4-PE40, a soluble recombinant fusion toxin selectively toxic to gp120-expressing cells, were evaluated in persons infected with human immunodeficiency virus type 1 (HIV-1). Seventeen of 24 patients who completed a single-dose safety trial were given either 1, 5, 10, or 15 micrograms/kg of sCD4-PE40 by intravenous bolus once a month for 2 months, then weekly for 6 weeks. The weekly maximally tolerated dose was 10 micrograms/kg. The major toxicity was a transient dose-dependent elevation in hepatic aminotransferases peaking 48 h after infusion. Anti-Pseudomonas exotoxin antibody developed in 58% of recipients, and sera from 13 of 17 showed neutralizing activity against sCD4-PE40. No consistent changes in immunologic or virologic markers were observed. Weekly infusions of < or = 10 micrograms/kg of sCD4-PE40 are generally well tolerated, but additional studies correlating optimal dosing and frequency of administration with efficacy will be needed to define the role of this novel agent in the management of HIV-1-infected patients.

Interferon-alpha in patients with asymptomatic human immunodeficiency virus (HIV) infection. A randomized, placebo-controlled trial.

STUDY OBJECTIVE: To evaluate the toxicity and clinical efficacy of interferon-alpha 2b (IFN-alpha) in patients with asymptomatic human immunodeficiency virus (HIV) infection. DESIGN: Randomized, placebo-controlled, and double-blind study. SETTING: Outpatient clinic of a government referral-based research hospital. PATIENTS: Volunteer sample of 34 patients with asymptomatic HIV infection who had CD4 counts of 400 cells/mm3 or more, positive peripheral blood mononuclear cell cultures for HIV, or p24 antigenemia. INTERVENTIONS: Patients were randomly assigned to receive either IFN-alpha or placebo, 35 x 10(6) units per day subcutaneously. Doses of IFN-alpha or placebo were modified according to predefined laboratory and clinical criteria. Therapy lasted at least 12 weeks. MEASUREMENTS AND MAIN RESULTS: Seventeen patients were randomly assigned to each group. The two groups had similar mean CD4 counts at study entry. Thirty-five percent of patients assigned to receive IFN-alpha withdrew from the study because of toxicity. The average daily dose of IFN-alpha was 17.5 x 10(6) units. All patients receiving IFN-alpha reported flu-like symptoms; other toxicities included granulocytopenia (55%) and elevated liver enzyme levels (45%). While receiving IFN-alpha, 7 patients (41%) became HIV culture negative (three or more consecutive negative peripheral blood mononuclear cell cultures taken at least 2 weeks apart). In contrast, 2 patients in the placebo group (13%) became culture negative while on study (P = 0.05). During the treatment period, CD4 lymphocyte percentages were sustained at or above the baseline level in patients receiving IFN-alpha and declined slightly in patients receiving placebo. Of the 32 study patients followed after study (range, 5 to 33 months), no patients in the IFN-alpha group developed an acquired immunodeficiency syndrome (AIDS)-defining opportunistic infection, compared with 5 patients in the placebo group (P = 0.02). CONCLUSIONS: Treatment of early-stage HIV infection with IFN-alpha can result in a decrease in frequency of viral isolation. Although its use may be accompanied by dose-dependent toxicities, IFN-alpha may have a role in slowing progression of HIV disease.

Anti-retroviral effects of interferon-alpha in AIDS-associated Kaposi's sarcoma.

21 patients with AIDS and Kaposi's sarcoma were enrolled in an open therapeutic trial to determine the in vivo anti-retroviral activity of recombinant interferon-alpha (IFN-alpha). 8 (38%) showed a complete or partial anti-tumour response. The mean pretreatment CD4 count for the responders was 399 cells/microliter vs 154 cells/microliter for the non-responders. All 5 of the patients with more than 400 CD4 cells/microliter pretreatment showed a significant reduction in tumour, whereas none of the 7 patients with under 150 CD4 cells/microliter had any response. 5 of the 6 complete or partial responders with greater than 50 pg/ml of human immunodeficiency virus (HIV) p24 before IFN therapy showed a 75% or greater reduction by 12 weeks of therapy, with 3 patients having persistently negative HIV cultures. The anti-viral effects were also most pronounced in the patients with the highest CD4 counts. These data demonstrate the potential benefits, both anti-tumour and anti-retroviral, of treatment with IFN-alpha in the early stages of HIV infection and Kaposi's sarcoma.