Phase I trial of subcutaneous recombinant human interleukin-12 in patients with advanced renal cell carcinoma.

Patients with advanced renal cell carcinoma were treated in a Phase I trial with escalating doses of recombinant human interleukin-12 (rHuIL-12) given on days 1, 8, and 15 of each 28-day cycle. Treatment in the initial dose scheme consisted of a fixed dose with dose levels of 0.1, 0.5, and 1.0 microg/kg given to cohorts composed of three or six patients. On the basis of the toxicity profile, a second scheme (up-titration) was undertaken wherein rHuIL-12 was escalated for each patient from week 1 to week 2, to a target dose given week 3 and thereafter; cohort target dose levels were 0.5, 0.75, 1.0, 1.25, and 1.5 microg/kg. Fifty-one patients were treated: 32 (63%) had prior cytokine therapy and 19 (37%) had received no prior systemic therapy. The maximum tolerated dose for the fixed dose scheme was 1.0 microg/kg. Dose-limiting toxicities included increase in transaminase concentration, pulmonary toxicity, and leukopenia. The most severe toxicities occurred with the first injection and were milder upon further treatment. With the up-titration dose scheme, the maximum tolerated dose was reached at 1.5 microg/kg, and dose-limiting toxicity consisted of an increase in serum transaminase levels. At the maximum tolerated dose of 1.5 microg/kg, serum IL-12 levels increased to a mean peak level of 706 pg/ml. Serum levels of IFN-gamma increased to a mean peak level of about 200 pg/ml at 24 h after the first maintenance dose of 1.5 microg/kg. The best responses were as follows: one patient had complete response, 34 patients were stable, 14 patients showed progression, and 1 patient was inevaluable. In conclusion, rHuIL-12 was relatively well tolerated when administered by s.c. injection. The recommended dose according to the up-titration schedule of rHuIL-12 (microg/kg) for Phase II trials was as follows: cycle 1, 0.1 (day 1), 0.5 (day 8), 1.25 (day 15); cycle 2 onwards, 1.25. Phase II trials of rHuIL-12 were initiated in previously untreated patients with renal cell carcinoma and in patients with melanoma.

Signal transduction abnormalities in T lymphocytes from patients with advanced renal carcinoma: clinical relevance and effects of cytokine therapy.

Studies have demonstrated abnormalities of the CD3/T-cell antigen receptor (TCR) and pathways of signal transduction in T lymphocytes from animals and patients with advanced malignancy. Diminished expression of TCRzeta and p56(lck) that are associated with the TCR and reduced nuclear localization of RelA containing nuclear factor kappaB (NFkappaB) complexes have been noted. These defects have been described in T cells from patients with malignant melanoma, renal cell carcinoma (RCC), ovarian cancer, and colorectal cancer. Preliminary observations also indicate possible correlation with clinical variables such as stage in selected instances. To further characterize altered expression of TCRzeta, p56(lck), and impaired activation of NFkappaB, T lymphocytes were obtained from 65 patients with RCC, the majority of whom were receiving combination cytokine therapy [interleukin (IL)-2, IFN alpha-containing regimens] and 37 control individuals. In 29 of these patients, levels of TCRzeta and p56(lck) were determined by Western blots of T-cell lysates and semiquantitated using densitometry. Relative levels were then correlated with a series of clinical variables including response to therapy, performance status, survival, disease sites, age, and others. In another group of 28 patients (three individuals from the first group), the frequency of abnormal NFkappaB activation was studied using electrophoretic mobility shift assays after activation of T cells with phorbol myristate acetate/ionomycin or anti-CD3 monoclonal antibody. Changes in these signaling molecules during cytokine treatment were also investigated. TCRzeta and p56(lck) were detected in the peripheral blood T cells in 27 of 29 patients, and overall, reduced levels were noted visually in 12 of 29 (41%) and 13 of 29 (45%) individuals, respectively. When levels were semiquantitated using densitometry, significant decreases of TCRzeta (P = 0.029) and p56(lck) (P = 0.029) but not CD3epsilon (P = 0.131), compared with control levels, were found. In patients treated with IL-2/IFN alpha-based therapy, relative levels of TCRzeta increased significantly (P = 0.002) on day 15 of cycle one compared with the baseline. Correlations of TCRzeta or p56(lck) levels with response or disease variables, except for lower TCRzeta levels (P < 0.001) in the presence of bone metastases, were not found. Abnormal NFkappaB activation after stimulation with phorbol myristate acetate/ionomycin and/or anti-CD3 monoclonal antibody was found in 59% of patients (17 of 28) and was not accounted for by the advanced age of the study cohort. Activation of NFkappaB in peripheral blood T cells was inducible during cytokine therapy in four of six individuals who displayed impaired NFkappaB activity prior to therapy. Moreover, impaired activation of NFkappaB does not appear linked to a reduction of TCRzeta expression, because in five patients, normal TCRzeta levels were present although kappaB binding was not inducible. In the majority of patients with advanced RCC, peripheral blood T cells express TCRzeta and p56(lck), and in a subset, reduced levels of these TCRzeta associated molecules are seen that may increase during cytokine-based therapy. Abnormal activation of NFkappaB is also present in >50% of patients and may also revert to normal during IL-2/IFN alpha-based treatment. This alteration in NFkappaB activation occurred in the presence of normal expression of TCRzeta-associated signaling elements. The clinical significance of these findings remains unclear.

Phase I trial of subcutaneous interleukin 3 in patients with refractory malignancy: hematological, immunological, and pharmacodynamic findings.

We conducted a Phase I trial of s.c. recombinant human interleukin 3 (rhIL-3) to evaluate the toxicity, maximal tolerated dose, pharmacokinetics, and in vivo biological effects of this cytokine. Thirty-one patients with refractory cancer were entered into the study between November 1991 and June 1993. Therapy consisted of s.c. rhIL-3 daily for 15 days administered to cohorts of three to nine patients at dose levels of 60-4000 microgram/m2/day. Cycles were repeated at intervals of 28 days. Seventy-five cycles of rhIL-3 were administered (median, two per patient) and the maximal tolerated dose was 2000 microgram/m2/day. Toxicity was moderate, with most patients developing chills, fever, and myalgia. Dose-limiting toxicity consisted of diarrhea (two patients) and headache (one patient). Hematological effects of rhIL-3 included significant dose-related increases of WBC (P < 0.001), neutrophils (P < 0.001), and eosinophils (P < 0.001). Platelet counts and absolute lymphocyte numbers also increased. Various CD3(+) lymphocyte subsets increased; however, lytic activity (natural killer and lymphokine-activated killer) of peripheral blood lymphocytes was not enhanced. Serum levels of the soluble IL-2 receptor increased in a dose-related fashion, and IL-2-induced lymphocyte proliferation also was increased variably. Pharmacokinetic studies were performed in 13 patients, and area under the curve and maximal concentration values increased with increasing rhIL-3 dose levels (P < 0.001) and correlated with maximal changes from baseline in WBC, neutrophils, and eosinophils. rhIL-3 antibodies were detected in 8% of patients by day 29 of cycle 1 but were not neutralizing. rhIL-3 is well tolerated when administered s.c. and has reproducible hematological and immunological effects. The pleiotropic effects of this cytokine on various in vivo biological parameters were demonstrated clearly. Further studies of its immunoregulatory effects are warranted.

Sweet taste preference as a risk factor for alcohol dependence.

OBJECTIVE: Previous research has found that alcoholics have a greater preference for sweet solutions than comparison subjects. This study tested the hypothesis that preference for sweet solutions is a marker for alcoholism risk. METHOD: A total of 122 nonalcoholic subjects (59 men) participated. Fifty-eight subjects had a paternal history of alcoholism, and 64 did not. Each subject rated a series of sucrose solutions for intensity of sweetness and degree of preference. RESULTS: Subjects were able to rate accurately the relative intensity of sweetness in the sucrose solutions. Both subjects with and those without a paternal history of alcoholism preferred a 0.42-M sucrose solution, irrespective of gender. CONCLUSIONS: This study failed to support the hypothesis that sweet preference is a marker of alcoholism risk. The sweet preference observed previously among alcoholics may be a consequence of chronic alcohol consumption or other factors associated with heavy drinking.

A method to study apoptosis in eosinophils by flow cytometry.

The aim of this study was to develop a simple flow cytometric procedure to study eosinophil apoptosis. Eosinophils were isolated from the peripheral blood of healthy, non-allergic individuals and then cultured in basal culture medium. The cells were examined after 24, 48 and 72 h for forward- and side scatter (FS-SSC) pattern, staining with FDA, PI, and anti-CD95, and light microscopic appearance. After culture for >24 h, two populations with different FS-SSC-patterns appeared, referred to as A and B. Population A consisted of living, FDA-positive eosinophils. The eosinophils in population B showed a lower FS scatter than those in population A and a staining pattern with PI indicating the presence of hypodiploid DNA. Anti-CD95 demonstrated a significant staining of the eosinophils in population B, which increased after 2 days in culture. The cells were sorted using a FACS-Scan cell sorter and by Annexin V-coated magnetic beads to permit separate analyses of PI-staining pattern, DNA electrophoresis, and light microscopic examination of the cells in population B. The present study suggest that it is possible to discriminate between apoptotic and living eosinophils using the FS-SSC pattern and the PI-staining pattern obtained by flow cytometry.

GM-CSF safety and effects in the management of advanced/refractory multiple myeloma patients: a phase I trial.

PURPOSE: Some limitations of effective therapy in multiple myeloma include the low growth fraction of the malignant plasma cells, multi-drug resistance, and the presence of other concurrent diseases in this patient population. A phase I study was conducted to evaluate the toxicity of granulocyte macrophage colony stimulating factor (GM-CSF) in myeloma patients as well as the potential effect on the plasma cell labeling index (PCLI). Relapsed patients with multiple myeloma were eligible. METHODS: The first phase of this trial assessed the toxicity (including the effect on disease progression) of escalating doses (125-500 microg/m2 SC, days 1-5) of GM-CSF, and the effects of this cytokine on PCLI. Patients whose PCLI doubled and increased to > or = 1.7% were treated with chemotherapy including cyclophosphamide, vincristine, prednisone, and GM-CSF. Twenty-two patients were enrolled. RESULTS: The toxicity of GM-CSF was mild, and no dose-limiting side effects were seen. Twenty-five percent of patients (5/20) achieved the target PCLI, and 4/5 proceeded to receive chemotherapy. No relationship of GM-CSF dose to increases of the PCLI was noted. All patients who received chemotherapy responded. CONCLUSIONS: GM-CSF has acceptable toxicity in patients with multiple myeloma and produced increases of PCLI in selected individuals. Further studies of GM-CSF alone or in combination with chemotherapy are indicated.

Characterisation of Haemophilus spp. isolated from infant conjunctivitis.

Haemophilus strains isolated from children under the age of 11 months with conjunctivitis were characterised by biotype, sugar fermentation, plasmid pattern and outer-membrane-protein profiles. H. influenzae was the most common species identified and was separated into 14 groups based on sugar fermentation and biotype patterns and into more than 20 groups when plasmid and outer-membrane-protein profiles were included. Small (mol. wt less than 10 X 10(6)) plasmids were identified in 11 of 34 (32%) H. influenzae isolates, 1 of 2 H. haemolyticus and 4 of 6 (67%) H. parainfluenzae isolates. Examination of sugar-fermentation and plasmid patterns increased the ability to distinguish between strains isolated at different times from recurrent disease and may have general applications in the study of Haemophilus strains isolated from a single anatomical site.

Electrophoretic mobility of salbutamol drug powder in mixed propellant solvents.

The influence of lipids on the dispersion properties of micronized Salbutamol base drug in liquid fluorocarbons has been characterized by electrophoretic mobility measurements and by particle size measurements. A modified Malvern ps26 microelectrophoretic cell was employed, allowing pressurized samples to be analyzed. The measurements were carried out at 25 degrees C in 100:0, 50:50, 40:60, and 30:70 blends of trichlorofluoromethane (P11) and dichlorodifluoromethane (P12) as a function of oleic acid concentration. A limited number of measurements were also done with soybean lecithin or synthetic dipalmitoylphosphatidylcholine (DPPC). A solvent series based on the polarizability (alpha) and on the dipole moment (mu) of the solvent molecules is constructed in order to estimate the acid-base character of the propellants. The results indicate that the type and the amount of lipids and also the type of fluorocarbon mixture plays an important role in the formation of surface charge. The dispersion stability with respect to the measured particle size does not always correlate with the measured electrophoretic mobility, and hence, the surface charge cannot alone explain the dispersion stability. Instead, the wettability of the powders seems to be important as well. Positive surface charge is obtained with the oleic acid or with synthetic dipalmitoylphosphatidylcholine, but negative surface charge exists with soybean lecithin.

Fetal exposure from endovaginal ultrasound examinations in the first trimester.

Ultrasonic obstetrical examinations during the first trimester are now often performed endovaginally with higher-frequency (5-7.5 MHz) transducers operating closer to the fetus than for transabdominal examinations. To estimate exposure to the fetus, propagation distances were obtained from a retrospective study of 100 normal first-trimester endovaginal B-mode examinations. No significant dependence of attenuation on gestational age was observed. The range of the attenuation estimates was 1.8-10.4 dB. A mean attenuation of 5.0 dB at 5 MHz for an average depth of 2.8 cm resulted in an attenuation coefficient of .36 dB/cm/MHz. Exposure (ISPTA) to the fetus at each gestational week from three ultrasound units was very similar: worst-case values of the 100 cases ranged from 1.2-1.9 mW/cm2, well within the Food and Drug Administration (FDA) guidelines of 94 mW/cm2 for derated focused transducers. Energy density deposited to the anterior surface of the fetus during a typical examination, assuming that the transducer is kept stationary over one area for the entire period of the examination (which is unlikely), ranged from 143-217 mJoules/cm2, within the American Institute of Ultrasound in Medicine (AIUM) recommendations.

The predictive power of combined neuropsychological measures for attention-deficit/hyperactivity disorder in children.

The present study explores the predictive power of seven neuropsychological assessment tools used in combination in classifying children with attention-deficit/hyperactivity disorder (ADHD). Twenty-one ADHD boys and 22 community control children participated. Group differences were significant on the continuous performance test only; however, battery analysis did increase overall predictive power, which was moderate. This study highlights the difficulty in identifying consistent mean differences on tests of frontal/executive functioning across studies, as well as the need to assess the predictive validity of these tests in classifying children with ADHD. The study suggests that these tests may provide greater predictive validity when used in combination. Inconsistencies in the literature are discussed, with consideration of research methodology, the heterogeneity of the ADHD population, and comorbid diagnoses.

Descriptions of functioning and health and relations to a gross motor classification in adults with cerebral palsy.

PURPOSE: The aim of this study was to describe functioning and health, and explore the use of the Gross Motor Function Classification System (GMFCS) in an adult population with cerebral palsy (CP). METHODS: From a cohort of 199 persons, 48 persons were selected for structured interviews and functional assessments regarding activities of daily living, motor function, range of motion, pain and general health. RESULTS: A third of the population had deteriorated in function from adolescence to adulthood according to the GMFCS. The majority were independent in personal ADL, but many of those were dependent in instrumental ADL. Motor function scores reflected problems in walking ability, and limited ROM and pain were common in all functional levels. General health was lower than in a general population. GMFCS seems valid for classifying adults with CP since it is correlated with instruments measuring motor function and ADL in terms of dependence. CONCLUSION: Decreased functional ability and secondary musculoskeletal problems are common in adults with CP and general health can be associated with those problems. It is important to further explore health aspects and relations between health status and self-perceived health. The GMFCS is a useful tool, especially for comparisons throughout the life span, but in order to use in an adult population further development is needed.

A novel CD44v6 targeting antibody fragment with improved tumor-to-blood ratio.

The chimeric monoclonal antibody U36 (cMAb U36) recognizes the CD44v6 antigen. Its potential as a radioimmunotargeting agent, as well as its safety, has been shown in previous studies in head and neck cancer patients. However, intact MAbs have long circulation time in the blood and tumor targeting may also be hampered due to the slow and incomplete diffusion into solid tumors. In comparison, smaller monovalent Fab' and divalent F(ab')2 fragments are expected to exhibit shorter circulating half-lives, better tumor penetration and are thus more likely to yield better imaging results. In this study, novel F(ab')2 and Fab' fragments from cMAb U36 were radiolabeled with 125I and the characteristics of the conjugates in vitro were examined. The biodistribution of the conjugates were then evaluated in nude mice bearing CD44v6-expressing xenograft tumors. Furthermore, the penetration depth and distribution in tumor tissue was assessed by autoradiography in selected tumor samples. The in vitro experiments showed that the conjugates were stable and had intact affinity to CD44v6. The biodistribution study demonstrated superior tumor-to-blood ratio for the novel cMAb U36 fragment 125I-F(ab')2 compared with both the intact MAb and the monovalent fragment form. Autoradiography also revealed better tumor penetration for 125I-F(ab')2. This study demonstrates that the use of antibody fragments may improve radioimmunotargeting and possibly improve the management of head and neck malignancies.

Herschel detects a massive dust reservoir in supernova 1987A.

We report far-infrared and submillimeter observations of supernova 1987A, the star whose explosion was observed on 23 February 1987 in the Large Magellanic Cloud, a galaxy located 160,000 light years away. The observations reveal the presence of a population of cold dust grains radiating with a temperature of about 17 to 23 kelvin at a rate of about 220 times the luminosity of the Sun. The intensity and spectral energy distribution of the emission suggest a dust mass of about 0.4 to 0.7 times the mass of the Sun. The radiation must originate from the supernova ejecta and requires the efficient precipitation of all refractory material into dust. Our observations imply that supernovae can produce the large dust masses detected in young galaxies at very high redshifts.

Searching for information, understanding, and self-value: the utilization of peer support groups by gay men with HIV/AIDS.

The present study examined the patterns and variations characterizing the support group involvements of 25 gay men with HIV/AIDS. Results indicated that those men who participated in support groups on a long-term basis (i.e., one year or more) were less likely to have access to other social support networks. They were also most interested in receiving and exchanging emotionally-oriented forms of support, such as empathy, acceptance and camaraderie. By contrast, the men who participated in support groups for a brief period of time (i.e., six months or less) had greater access to alternative support networks and were more interested in receiving and exchanging instrumental forms of support, such as illness-related information and examples of effective coping. Those men who elected not to participate in support groups emphasized their relatively good health, the strength of their existing support systems and their reluctance to see others with life-threatening symptoms. Finally, regardless of whether and how they participated in formal support groups, the majority of men in this study benefitted from interacting regularly with peers. Through these interactions, they received helpful understanding, information and friendship. In addition to this, they often experienced a revitalizing sense of purpose, efficacy and mutuality which enabled them to cope more successfully with their illness.

Effects of propofol on desipramine-sensitive [3H]-noradrenaline uptake kinetics in rat femoral artery.

BACKGROUND: The intravenous anaesthetic propofol inhibits the neuronal uptake of noradrenaline (uptake1) from the vascular sympathetic neuromuscular junction, resulting in an enhancement of the sympathetic neurotransmission. This could be important for maintenance of blood pressure during propofol anaesthesia. The aim of the present study was to determine how propofol influences the kinetics of uptake1. METHODS: Isolated segments of rat femoral arteries were incubated with [3H]-noradrenaline in the presence or absence of propofol and the radioactivity taken up was measured in a scintillation counter. The uptake1 inhibitor, desipramine, was used to delineate the specific neuronal uptake. RESULTS: Desipramine and 10 microM propofol significantly reduced the uptake in segments incubated with 0.1 microM [3H]-noradrenaline. Propofol at 1 microM and 100 microM did not affect the uptake. Non-linear regression analysis of specific uptake yielded Km 0.50 microM, Vmax 1.6 pmol mg(-1) 15 min(-1) and Hill coefficient 1.1. Propofol (1-10 microM) increased the Km value and propofol (10-100 microM) increased the Vmax value concentration-dependently, while the Hill coefficient was not affected. CONCLUSION: Propofol seems to have a biphasic effect on the uptake of noradrenaline in the vascular sympathetic neuromuscular junction. At lower propofol concentrations there is a decrease in the affinity of the noradrenaline transporters. The resulting uptake inhibition is counteracted at higher propofol concentrations by an increase in the efficacy of the uptake.

Inhibition of the CD28-CD80 co-stimulation signal by a CD28-binding affibody ligand developed by combinatorial protein engineering.

CD28 is one of the key molecules for co-stimulatory signalling in T cells. Here, novel ligands (affibodies) showing selective binding to human CD28 (hCD28) have been selected by phage display technology from a protein library constructed through combinatorial mutagenesis of a 58-residue three-helix bundle domain derived from staphylococcal protein A. Analysis of selected affibodies showed a marked sequence homology and biosensor analyses showed that all investigated affibodies bound to hCD28 with micromolar affinities (KD). No cross-reactivity towards the related protein human CTLA-4 could be observed. This lack of cross-reactivity to hCTLA-4 suggests that the recognition site on hCD28 for the affibodies resides outside the conserved MYPPPYY motif. The apparent binding affinity for hCD28 could be improved through fusion to an Fc fragment fusion partner, resulting in a divalent presentation of the affibody ligand. For the majority of selected anti-CD28 affibodies, in co-culture experiments involving Jurkat T-cells and CHO cell lines transfected to express human CD80 (hCD80) or LFA-3 (hLFA-3) on the cell surface, respectively, pre-incubation of Jurkat cells with affibodies resulted in inhibition of IL-2 production when they were co-cultured with CHO (hCD80+) cells, but not with CHO (hLFA-3+) cells. For one affibody variant denoted Z(CD28:5) a clear concentration-dependent inhibition was seen, indicating that this affibody binds hCD28 and specifically interferes in the interaction between hCD28 and hCD80.

Intraoperative glucose administration influences respiratory quotient during paediatric anaesthesia.

BACKGROUND: Oxidation of carbohydrates and fat yields respiratory quotients (RQ) of 1.0 and 0.7 respectively. Maintained or increased blood glucose concentrations are usually seen during paediatric anaesthesia and surgery even without glucose administration. The aim of the present study was to evaluate whether an intraoperative glucose infusion influences the RQ as an indication of a different metabolic preference in comparison to a glucose-free fluid regime. METHODS: Eighteen children between 0.5 and 24 months of age were studied during anaesthesia with controlled ventilation, oxygen in air, isoflurane, thiopentone, atracurium and fentanyl. Oxygen consumption and carbon dioxide production were measured using indirect calorimetry All children received Ringer acetate as needed; in addition, nine children were given glucose 10%, 3 ml.kg-1.h-1, corresponding to 300 mg.kg-1.h-1. Blood samples for analyses of glucose, lactate, free fatty acids and ketones were taken before and during surgery. RESULTS: RQ was significantly higher in the children given glucose 0.92 +/- 0.08, compared to 0.81 +/- 0.06 in the children without glucose (P < 0.01). Oxygen consumption tended to be higher, although not significantly so, in patients without glucose infusion. Energy expenditure was 1.70 +/- 0.29 kcal.kg-1.h-1, without significant group differences. Higher blood glucose concentrations during surgery were found in the children given glucose. CONCLUSIONS: Our results indicate a higher glucose oxidation rate in patients given glucose during surgery.