Control of viremia and prevention of clinical AIDS in rhesus monkeys by cytokine-augmented DNA vaccination.

With accumulating evidence indicating the importance of cytotoxic T lymphocytes (CTLs) in containing human immunodeficiency virus-1 (HIV-1) replication in infected individuals, strategies are being pursued to elicit virus-specific CTLs with prototype HIV-1 vaccines. Here, we report the protective efficacy of vaccine-elicited immune responses against a pathogenic SHIV-89.6P challenge in rhesus monkeys. Immune responses were elicited by DNA vaccines expressing SIVmac239 Gag and HIV-1 89.6P Env, augmented by the administration of the purified fusion protein IL-2/Ig, consisting of interleukin-2 (IL-2) and the Fc portion of immunoglobulin G (IgG), or a plasmid encoding IL-2/Ig. After SHIV-89.6P infection, sham-vaccinated monkeys developed weak CTL responses, rapid loss of CD4+ T cells, no virus-specific CD4+ T cell responses, high setpoint viral loads, significant clinical disease progression, and death in half of the animals by day 140 after challenge. In contrast, all monkeys that received the DNA vaccines augmented with IL-2/Ig were infected, but demonstrated potent secondary CTL responses, stable CD4+ T cell counts, preserved virus-specific CD4+ T cell responses, low to undetectable setpoint viral loads, and no evidence of clinical disease or mortality by day 140 after challenge.

Repetitive weekly cycles of 4-day continuous infusion of recombinant interleukin-2: a phase I study.

A phase I trial was performed with a new interleukin-2 (IL-2) given as a continuous intravenous infusion in patients with solid tumors. The objectives of the study were to examine the feasibility of administering IL-2 in 4-day cycles for 4 consecutive weeks, and to investigate the response pattern of peripheral blood lymphocytes. Tumor necrosis factor (TNF) and IL-2 serum concentrations were also measured. Prior to this study, IL-2 had been tested at increasing dosages during one 4-day cycle, and it appeared that a dose of 1300 mcg/m2/day was tolerated. However, when this treatment schedule was maintained for 4 consecutive weeks, the maximum tolerated dose was 430 mcg/m2/day. In this schedule, a dose-dependent progressive increase in rebound lymphocyte count occurred after each weekly cycle, resulting in a 5-70-fold increase after the 4th cycle. Serum TNF peak concentrations also showed a tendency to increase during each subsequent cycle, while serum IL-2 peak concentrations showed a paradoxical decrease. Clinical toxicity comprised several events, which, possibly, could be ascribed to autoimmune phenomena. Myocardial infarction as a late toxicity of IL-2 is suggested. One complete response (renal carcinoma) and two partial responses (renal and breast carcinoma) were documented, one of these occurring in a patient who previously had shown a transient response on interferon therapy.

Factors related to high levels of ostrich chick mortality from hatching to 90 days of age in an intensive rearing system.

Ostrich chick mortality was studied in 2522 chicks that were hatched artificially during the 1999/2000 breeding season. High levels of mortality were observed, with 1978 (78.4 %) of these chicks dying before 90 days after hatching. A total of 46.7 % (1,177) of these chicks died before 28 days of age, and a further 30.7% (801) died between 28 and 90 days post-hatching. Chick mortality to 28 days of age could not be conclusively related to sex, day of external pipping or breeder diet. Mortality rates were higher (P< 0.05) at the beginning and end of the breeding season than in the middle months. Differences in mortality levels of chicks incubated in different incubators could be related to the time of the breeding season during which the incubator was mostly used. The regression of chick mortality to 28 days of age on day-old chick mass followed a 2nd-degree polynomial. Chicks with day-old masses below 762.5 g were particularly at risk of dying before 28 days after hatching. Chicks hatching from eggs where excessive water loss to 35 days of incubation (>18%) was recorded were also at risk of succumbing before 28 days of age. Chick mortality percentages for the period from 28 to 90 days of age exceeded 80 % in chicks weighing an average of 1.050 g at 28 days. Mortality percentages declined sharply at higher live masses, to between 20 and 30 % in chicks weighing > or = 1,950 g. This 'core' level of mortality remained throughout, even in the heaviest chicks. It was concluded that the high levels of chick mortality could be related to stress in chicks, resulting from an inability to adapt to the rearing environment. The high subsequent mortality percentages of low live mass chicks that survived to 28 days after hatching could probably be attributed to residual setbacks suffered earlier. Abetter understanding of the underlying principles involved in ostrich chick mortality in intensive rearing environments is required for progress in this field, resulting in more predictable survival rates under these conditions.

The use of intermediate dose cytosine arabinoside (ID Ara-C) in the treatment of acute non-lymphocytic leukaemia in relapse.

Cytosine arabinoside in a high dose of 3 g/m2 (HD Ara-C), alone or in combination with doxorubicin, has been advocated for the treatment of patients with acute non-lymphocytic leukaemia (ANLL) in relapse. Although a remission rate of 65% has been reported, the toxicity was severe and was possibly related to the high plasma concentrations of Ara-C (about 100 microM) reached during 1 h infusion. It has been postulated, however, that the intracellular enzymes which convert Ara-C into the active metabolite cytosine arabinoside triphosphate (Ara-CTP), are saturated at plasma concentrations of about 10 microM. We calculated that this level could be reached with an intermediate dose of 0.5 g/m2 Ara-C, given in a 1 h infusion (ID Ara-C). Subsequently 15 patients with ANLL (12 in relapse and three refractory to conventional therapy) were treated with ID Ara-C every 12 h for 6 d in combination with doxorubicin and vincristine. The overall remission rate was 80%. The median duration of bone marrow depression was 20 d (range 14-29 d) and side effects were comparable to conventional treatment. These preliminary data suggest that the therapeutic results of this ID Ara-C regimen are not inferior to comparable schedules with HD Ara-C as reported by others while toxicity is less severe.

Does the metabolite uracil arabinoside inhibit cytosine arabinoside (Ara-C) penetration into the cerebrospinal fluid during high-dose Ara-C therapy?

A patient with acute leukaemia was treated with i.v. 2-h infusions of Ara-C at a dose of 3.0 g/m2 every 12 h. During 6 d of therapy the concentrations of the metabolite Ara-U in the CSF reached rather high levels of between 60 and 70 mumol/l from d 2-6 due to high levels of Ara-U in the plasma. The concentration of Ara-C in the CSF after the first infusion was 10.8 mumol/l. After repetitive doses on d 2-6 the drug concentrations increased from about 3 mumol/l just before infusion to about 8 mumol/l at the end of infusion, indicating inhibition of Ara-C influx into the CSF during prolonged treatment. We suggest that the high levels of Ara-U in the plasma interfere with Ara-C transport across the blood-brain barrier.

Age dependent behaviour of red cell glycolytic enzymes in haematological disorders.

The age dependent behaviour of 11, mainly glycolytic, red blood cell enzymes in 26 patients with various haematological disorders has been investigated after separation of red blood cells by discontinuous density gradient centrifugation. The frequency of enzyme deficiencies in the old cells of these patients was significantly increased in comparison with the unseparated cells, 29 and 13 deficiencies, respectively. Particularly hexokinase activity, although normal or even increased in unseparated cells, was found deficient in old cells in seven cases. In addition, an increased number of phosphofructokinase deficiencies was observed in the patients' old cells (eight cases) as compared to the unseparated cells (three cases). However, the red blood cells of the majority of these patients were found to contain increased enzyme activities, irrespective of cell age. Enzyme activities in the youngest cell population did not correlate with the reticulocyte count. Cases of high pyruvate kinase and hexokinase activities were studied for kinetical, electrophoretical and immunological properties of the respective enzymes, but no abnormalities could be demonstrated, indicating an increased synthesis of these enzymes.

Meningeal involvement in patients with acute nonlymphocytic leukemia. Incidence, management, and predictive factors.

Between 1977 and 1983, symptomatic meningeal leukemia was diagnosed in 9 of 93 adult patients with acute nonlymphocytic leukemia (10%). All cases occurred after complete remission had been achieved (46 patients), either as the only site of relapse (3 patients), or together with a first bone marrow relapse (3 patients), or after a bone marrow relapse (3 patients). Extramedullary involvement at initial diagnosis was the only independent predictive factor (P = 0.005), the number of patients with initial hyperleukocytosis being too small (three) for evaluation. Remission of the meningeal leukemia was obtained after treatment in four of eight patients. The presence of meningeal leukemia and the response to therapy had no influence on survival. However, the morbidity and therapy related toxicity of meningeal leukemia were impressive. Some recommendations for prophylactic treatment are suggested.

Human erythrocyte pyruvate kinase deficiency: the use of a kinetic study of mutant enzymes for the detection of heterozygotes.

Erythrocyte pyruvate kinase (PK) deficiency was detected in a boy of dutch origin. Immunologic, electrophoretic, and kinetic studies of the enzymes of propositus and the members of his family demonstrated that the boy was heterozygote for two different mutant PK alleles. The mutant enzyme, for which his mother was heterozygote, was characterized by a lower immunologic specific activity, a decreased affinity for the substrate phospho-enol-pyruvate, and a loss of homotropic interactions toward this substrate, an increased affinity toward the allosteric inhibitor MgATP2-, a decreased affinity for the activator fructose-1,6-diphosphate, and a lowered pH optimum. Electrophoresis, Km app. for MgADP, and the reactivity toward the purine nucleotide substrate analogues were normal. The mutant enzyme for which his father was heterozygote was characterized by a decreased affinity for the substrate phospho-enol-pyruvate and a loss of homotropic interactions toward this substrate. All other parameters mentioned above were normal. The use of a kinetic study of mutant enzymes for the detection of heterozygotes is discussed.

Low dose cytosine arabinoside (Ara C) in myelodysplastic syndromes.

Myelodysplastic syndromes (MDS) are acquired bone-marrow disorders, characterized by a decreased ability of the haemopoietic cell to differentiate, resulting in peripheral cytopenias. The majority of patients will die either from acute myeloid leukaemia or from infection and/or haemorrhage. Thirty-eight courses of low dose Ara C were administered in 26 MDS patients. Nineteen courses (50%) were associated with good (12) or partial (7) response. Three complete remissions were observed. The median duration of response overall was 19.5 weeks, 26 weeks for the good and 10 weeks for the partial responders. A high incidence of treatment failure was seen in patients treated after transition to AML. Major complications were observed during 14 courses and mortality was directly related to therapy in five patients. Platelet transfusions were required during 26 courses. Aggravation of peripheral-blood cytopenia during the first weeks and hypocellularity of bone-marrow aspirates at the end of therapy suggest that low-dose Ara C exerts its main activity by suppression of leukaemic growth, rather than by induction of differentiation in malignant cells. Our results in MDS patients demonstrate that low-dose Ara C can be of value in severe cytopenia and can decrease the proportion of leukaemic cells in the bone marrow, but the danger of treatment should not be underestimated.

Mean cardiac output by thermodilution with a single controlled injection.

OBJECTIVE: A new method to estimate mean cardiac output by thermodilution with a single duration-controlled injection was evaluated in patients. DESIGN: Prospective criterion standard study. SETTING: University hospital cardiac surgical intensive care unit and cardiac operation room. PATIENTS: Of 33 patients, 24 underwent coronary bypass graft surgery, four had a valve replacement, and five were treated in the intensive care unit. INTERVENTIONS: Interventions consisted of thermodilution cardiac output measurements. One single duration-controlled injection of cold fluid was used to calculate cardiac output. This controlled injection was performed with a duration equal to one whole ventilation cycle of the ventilator. An algorithm adapted to this duration-controlled injection calculated cardiac output. Moreover, this algorithm has properties to reduce errors caused by artificial ventilation and thermal noise. MEASUREMENTS AND MAIN RESULTS: In 33 patients, the averaged values of four measurements equally spread over the ventilatory cycle (phase-controlled) were compared with the values of two single duration-controlled measurements. The measurements were performed during periods of stable respiration and circulation. No significant difference was observed between the mean of four phase-controlled measurements and the mean of the two duration-controlled measurements. The cardiac output values in the intensive care patients were significantly higher compared with the two other patient groups (p <.05). The difference between the two methods could not be subdivided for the three patient groups (p >.05). The coefficient of variation of the single duration-controlled thermodilution measurements was significantly lower than the single phase-controlled measurements, 3% vs. 6% (p <.01). CONCLUSIONS: One single duration-controlled injection thermodilution measurement is as accurate and repeatable as the mean of four phase-controlled measurements and is clinically feasible.

Intensive cytotoxic therapy followed by autologous bone marrow transplantation for non-Hodgkin's lymphoma of high-grade malignancy.

Fourteen patients with non-Hodgkin's lymphoma (NHL) of high-grade malignancy were treated with cyclophosphamide and total body irradiation followed by autologous bone marrow transplantation (ABMT). All patients were pretreated with conventional chemotherapy. Three of four patients with drug-resistant disease achieved complete remission (CR), but relapse occurred within six months. Four patients in partial remission (PR) achieved CR; one died because of sepsis, two relapsed within six months, and one is still in CR 28+ months later. Six were treated in CR, five in first CR, and one in second CR. From these six patients (who received this treatment as consolidation therapy), five are in unmaintained CR seven to 31+ months after ABMT (one patient died of a secondary illness). There were two therapy-related deaths, both in patients with a poor clinical condition. Toxicity of this treatment was mild for those receiving transplants who were in better condition. These preliminary results suggest that intensive cytoreductive therapy followed by ABMT may improve disease-free survival in patients in NHL of high-grade malignancy in CR.

Cardiotoxicity as a dose-limiting factor in a schedule of high dose bolus therapy with interleukin-2 and alpha-interferon. An unexpectedly frequent complication.

BACKGROUND: In a group of patients with metastatic melanoma treated with high dose immunotherapy, there was an unexpectedly high incidence of severe cardiac adverse effects. METHODS: Sixteen patients with metastatic melanoma were treated with high dose interleukin-2 (IL-2) and alpha-interferon (alpha-IFN). Each treatment cycle consisted of IL-2 at a dose of 12 MIU/m2 and alpha-IFN at a dose of 3 MIU/m2, given as intravenous bolus injections every 8 hours on Days 1-5, every 3 weeks for a total of three cycles. Before treatment, careful cardiologic screening was performed, including electrocardiogram (ECG), stress test, cardiac multiple uptake-gated acquisition (MUGA) scan, and echocardiography. During therapy, patients were monitored with daily ECG and creatine phosphokinase measurements. Once cardiac damage was suspected, IL-2 and alpha-IFN were discontinued, and echocardiography, stress test and MUGA-scan were repeated. If indicated, cardiac catheterization with endomyocardial biopsies was performed. RESULTS: Despite pretreatment cardiac screening, seven patients (44%) exhibited myocardial injury. Acute myocardial infarction occurred in one patient, cardiomyopathy developed in four, asymptomatic ECG changes appeared in one, and 1 patient died of acute cardiac arrest. Echocardiography showed hypokinesis and decreased left ventricular ejection fraction. These abnormalities disappeared within 6 months. Cardiac catheterization in four affected patients revealed normal coronary arteries, but endomyocardial biopsies showed interstitial edema, vacuolation, and degeneration of myocytes. Electron-microscopic examination showed fragmentation of myofibrils, swelling of mitochondria and loss of mitochondrial cristae. CONCLUSIONS: This intensive treatment schedule of IL-2 and alpha-IFN is prohibited by severe and life-threatening cardiac toxicity.