Low-dose inhalation of interleukin-2 bio-chemotherapy for the treatment of pulmonary metastases in melanoma patients.

BACKGROUND: Interleukin-2 (IL-2) treatment for patients with metastatic melanoma has shown remarkable durable responses. Systemic administration of IL-2 may cause severe side effects, whereas local administration is considered to be a safe alternative. The lungs are common sites of metastases in melanoma patients causing considerable respiratory problems. We sought to evaluate the potential antitumoral effect of a low-dose inhalative IL-2 (lh-IL-2) regimen for patients with melanoma lung metastases. In addition, we explored the prophylactic potential of Ih-IL-2 after surgical removal of lung metastases in a study carried out in an outpatient setting. METHODS: Twenty patients with American Joint Committee on Cancer stage-IV (M1b and M1c) melanoma were enrolled in this study and treated with 3 × 3 million IU inhalative IL-2 q.d. together with monthly dacarbazine bolus injections. Five patients received lh-IL-2 after surgical resection of lung metastases to prevent recurrence of the disease (prophylaxis group, N=5). All other patients were enrolled in the treatment group (N=15). Clinical evaluations were carried out monthly and radiological follow-up was performed every third month. RESULTS: Nine patients in the treatment group had a clinical benefit with partial regression (27%) or stable disease (33%). Four patients had progression of lung metastases (26.7%) and two patients were not evaluable (13.3%). In the prophylaxis group, none of the patients developed new lung metastases during lh-IL-2 therapy. The median follow-up period was 7.8 months in the treatment group and 25.7 months in the prophylaxis group. In the majority of patients, treatment was well tolerated. CONCLUSIONS: Low-dose IL-2 inhalation might offer an effective and safe treatment option for lung metastases in melanoma patients. In addition, lh-IL-2 may have a prophylactic potential to prevent recurrence in the lungs after pulmonary melanoma metastasectomy. Administration can easily be performed in an outpatient setting, thus offering an attractive treatment option.

[Minor antigens - major impact. The role of minor histocompatibility antigens in allogeneic hematopoietic stem cell transplantation]. / Kleine Antigene - grosse Wirkung. Die Rolle von Minor-Histokompatibilitätsantigenen bei der allogenen Blutstammzelltransplantation.

Allogeneic hematopoietic cell transplantation (HCT) is often the only curative treatment option for patients with malignant and non-malignant hematological diseases. There is striking evidence that immunological Graft-versus-Leukemia (GvL)-reactions efficiently eradicate malignant cells after transplant. After HLA-matched HCT both the beneficial GvL-effect and the detrimental Graft-versus-Host Disease (GvHD) are mediated by donor derived T-cells specific for minor histocompatibility antigens (mHag) that differ between patient and stem cell donor. In addition, tumor-specific antigens can also be targeted and contribute to GvL-reactivity. This review summarizes the state-of-the-art knowledge on mHag and presents the potential therapeutical options on example of the mHag HA-1. HA-1 is currently the best characterized mHag and particularly attractive for immunotherapy due to the restricted expression on hematopoietic cells and on some solid tumors but not on cells involved during GvHD. This would allow amplifying the endogenous GvL-effect and selectively targeting malignant HA-1-positive cells without causing GvHD. HA-1-specific immunotherapy in eligible patient and donor pairs may range from vaccination with the immunogenic HA-1 peptide to the infusion of HA-1-specific cytotoxic T-cells (adoptive immunotherapy).

Children's use of mental state information in selecting persuasive arguments.

Whether and when children use information about others' mental states to invent or select persuasive strategies were examined. In Study 1, preschoolers, 3rd-graders, and 6th-graders (ns = 11, 12, and 16, respectively; 17 girls) were told about story characters' persuading parents to buy pets or toys. Children were either given or not given information about story parents' beliefs and asked to invent or select appropriate arguments. Older children, but not preschoolers, used belief information to select arguments. Results were replicated in Study 2 (16 kindergartners, 16 3rd-graders; 19 girls). In Study 3, kindergartners and 1st-graders (N = 16; 6 girls) reasoned well on false-belief tasks but not on persuasion tasks, suggesting that failure to consider mental states in persuasion was not due to lack of a belief concept. Findings suggest that mental state understanding may continue to develop after the preschool years; methodological qualifications are also considered.

Physiological optics in the hummingbird hawkmoth: a compound eye without ommatidia

The fast-flying day-active hawkmoth Macroglossum stellatarum (Lepidoptera: Sphingidae) has a remarkable refracting superposition eye that departs radically from the classical principles of Exnerian superposition optics. Unlike its classical counterparts, this superposition eye is highly aspherical and contains extensive gradients of resolution and sensitivity. While such features are well known in apposition eyes, they were thought to be impossible in superposition eyes because of the imaging principle inherent in this design. We provide the first account of a superposition eye where these gradients are not only possible, but also produce superposition eyes of unsurpassed quality. Using goniometry and ophthalmoscopy, we find that superposition images formed in the eye are close to the diffraction limit. Moreover, the photoreceptors of the superposition eyes of M. stellatarum are organised to form local acute zones, one of which is frontal and slightly ventral, and another of which provides improved resolution along the equator of the eye. This angular packing of rhabdoms bears no resemblance to the angular packing of the overlying corneal facets. In fact, this eye has many more rhabdoms than facets, with up to four rhabdoms per facet in the frontal eye, a situation which means that M. stellatarum does not possess ommatidia in the accepted sense. The size of the facets and the area of the superposition aperture are both maximal at the frontal retinal acute zone. By having larger facets, a wider aperture and denser rhabdom packing, the frontal acute zone of M. stellatarum provides the eye with its sharpest and brightest image and samples the image with the densest photoreceptor matrix. It is this eye region that M. stellatarum uses to fixate flower entrances during hovering and feeding. This radical departure from classical Exnerian principles has resulted in a superposition eye which has not only high sensitivity but also outstanding spatial resolution.

The development of biological explanation: are children vitalists?

Whether and how an understanding of biological explantation changes with development was explored in interviews with 24-first graders, 24 third graders, and 24 adults. Participants were asked about the changeability of biological and psychological characteristics and the causal mechanisms underlying biological, psychological, and mechanical phenomena (using both open-ended and forced-choice questions). In saying how characteristics might be changed, children and adults similarly distinguished between biology and psychology; they also responded similarly to questions about specific processes underlying biological change. Children's attributions of intention or agency to biological organs or body parts (i.e., vitalistic attribution) did not differ from adults', contrary to previous findings. The authors concluded that children's thinking about biology is not necessarily more vitalistic than adults'.

Maze learning by honeybees.

This study examines whether honeybees can learn to fly through complex mazes, in the presence or the absence of specific visual cues. The results are summarized as follows: 1. Bees can learn to fly through a complex maze by following a trail of colored marks. 2. Bees, initially trained to follow color marks through an initial part of the maze, are immediately able to use the same sign-tracking cue to find their way through the rest of the maze, which is unfamiliar to them. 3. Bees trained to follow color marks through a particular maze can use the same cue to negotiate a novel maze. 4. Bees trained to use a particular color to negotiate a maze can immediately use a novel color to negotiate the same maze or even a novel maze. 5. After learning to negotiate a maze by following colored marks, bees can find their way through the maze even when the marks are removed, albeit at reduced levels of accuracy. Thus, the trained bees do not rely solely on sign-tracking to find their way through the maze: they also acquire a spatial memory of the maze or at least a sequence of motor commands describing the correct path through it. 6. Bees can learn to use color as a signal even when it indicates the path through the maze in a symbolic way, for example, blue indicating a turn to the right and green a turn to the left. 7. Bees can learn an unmarked maze. Performance under these conditions is poorer than when marks are provided, but is still significantly better than chance level. 8. Control experiments rule out the use of external landmarks in all of these situations.

Stereospecific production of the herbicide phosphinothricin (glufosinate): purification of aspartate transaminase from Bacillus stearothermophilus, cloning of the corresponding gene, aspC, and application in a coupled transaminase process.

We have isolated and characterized an aspartate transaminase (glutamate:oxalacetate transaminase, EC 2.6.1.1) from the thermophilic microorganism Bacillus stearothermophilus. The purified enzyme has a molecular mass of 40.5 kDa by sodium dodecyl sulfate gel analysis, a temperature optimum of 95 degrees C, and a pH optimum of 8.0. The corresponding gene, aspC, was cloned and overexpressed in Escherichia coli. The recombinant glutamate:oxalacetate transaminase protein was used in immobilized form together with 4-aminobutyrate:2-ketoglutarate transaminase (EC 2.6.1.19) from E. coli for the production of L-phosphinothricin [L-homoalanin-4-yl-(methyl)phosphinic acid], the active ingredient of the herbicide Basta (AgrEvo GmbH), from its nonchiral 2-keto acid precursor 2-oxo-4-[(hydroxy)(methyl)phosphinoyl]butyric acid (PPO). In this new coupled process conversion rates of ca. 85% were obtained with substrate solutions containing 10% PPO by using only slight excesses of the amino donors glutamate and aspartate. The contamination of the reaction broth with amino acid by-products was < 3%.

Between desires and beliefs: young children's action predictions.

1 view of children's developing understanding of mind contends that children adopt a succession of naive psychological theories, moving from a desire-focused theory to a mature theory that attributes a greater role to beliefs. Between these, a transition theory, in which desires are primary but beliefs play an auxiliary role, is characteristic. Novel predictions arising from this view were tested in 2 experiments utilizing within-participants designs. Preschool children (Ns = 20 and 24) were asked to predict the actions of story characters who believed a desired object to be in one of two containers (which were shown to actually contain the desired object, nothing, or a different object). In both experiments, children's predictions accorded with belief significantly more when objects of the type desired were in both containers than when containers held nothing or other sorts of objects, supporting a transition theory interpretation over competing interpretations.

Molecular organization of the Escherichia coli gab cluster: nucleotide sequence of the structural genes gabD and gabP and expression of the GABA permease gene.

We have determined the nucleotide sequences of two structural genes of the Escherichia coli gab cluster, which encodes the enzymes of the 4-aminobutyrate degradation pathway: gabD, coding for succinic semialdehyde dehydrogenase (SSDH, EC 1.2.1.16) and gabP, coding for the 4-aminobutyrate (GABA) transport carrier (GABA permease). We have previously reported the nucleotide sequence of the third structural gene of the cluster, gabT, coding for glutamate: succinic semialdehyde transaminase (EC 2.6.1.19). All three gab genes are transcribed unidirectionally and their orientation within the cluster is 5'-gabD-gabT-gabP-3'. gabT and gabP are separated by an intergenic region of 234-bp, which contains three repetitive extragenic palindromic (REP) sequences. The gabD gene consists of 1,449 nucleotides specifying a protein of 482 amino acids with a molecular mass of 51.7 kDa. The protein shows significant homologies to the NAD(+)-dependent aldehyde dehydrogenase (EC 1.2.1.3) from Aspergillus nidulans and several mammals, and to the tumor associated NADP(+)-dependent aldehyde dehydrogenase (EC 1.2.1.4) from rat. The permease gene gabP comprises 1,401 nucleotides coding a highly hydrophobic protein of 466 amino acids with a molecular mass of 51.1 kDa. The GABA permease shows features typical for an integral membrane protein and is highly homologous to the aromatic acid carrier from E. coli, the proline, arginine and histidine permeases from Saccharomyces cerevisiae and the proline transport protein from A. nidulans. Uptake of GABA was increased ca. 5-fold in transformants of E. coli containing gabP plasmids.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular analysis of two genes of the Escherichia coli gab cluster: nucleotide sequence of the glutamate:succinic semialdehyde transaminase gene (gabT) and characterization of the succinic semialdehyde dehydrogenase gene (gabD).

We have characterized two genes of the Escherichia coli K-12 gab cluster, which encodes the enzymes of the 4-aminobutyrate degradation pathway. The nucleotide sequence of gabT, coding for glutamate:succinic semialdehyde transaminase (EC 2.6.1.19), alternatively known as 4-aminobutyrate transaminase, was determined. The structural gene consists of 1,281 nucleotides specifying a protein of 426 amino acids with a molecular mass of 45.76 kDa. The protein shows significant homologies to the ornithine transaminases from Saccharomyces cerevisiae and from rat and human mitochondria. Three functionally and structurally important amino acid residues of the transaminase were identified by sequence comparison studies, and evolutionary relationships of the aminotransferases are discussed. The gabD gene, encoding succinic semialdehyde dehydrogenase (EC 1.2.1.16), was cloned and shown to be located adjacent to the 5' end of gabT. Expression studies with subfragments of the initially cloned DNA region revealed a maximal size of 1.7 kb for gabD. Both genes are cotranscribed from a promoter located upstream of gabD.

Stereospecific production of the herbicide phosphinothricin (glufosinate) by transamination: isolation and characterization of a phosphinothricin-specific transaminase from Escherichia coli.

An aminotransferase capable of transaminating 2-oxo-4-[(hydroxy)(methyl)phosphinoyl]butyric acid to L-phosphinothricin [L-homoalanine-4-yl-(methyl)phosphinic acid], the active ingredient of the herbicide Basta (Hoechst AG), was purified to apparent homogeneity from Escherichia coli K-12. The enzyme catalyzes the transamination of L-phosphinothricin and various analogs with 2-ketoglutarate as the amino group acceptor. The transaminase has a molecular mass of 43 kilodaltons by sodium dodecyl sulfate-gel analysis and an isoelectric point of 4.35. The enzyme was most active in the high-pH region, with a maximum at pH 8.0 to 9.5, and had a temperature optimum of 55 degrees C. Heat stability was observed up to 70 degrees C. Substrate specificity studies suggested that the enzyme is identical with the 4-aminobutyrate:2-ketoglutarate transaminase (EC 2.6.1.19). The first 30 amino acids of the N terminus of the protein were determined by gas phase sequencing. The transaminase was immobilized by coupling to the epoxy-activated carrier VA-Biosynth (Riedel de Haen) and used in a column reactor for the continuous production of L-phosphinothricin. The enzyme reactor was operated for 7 weeks with only a slight loss of catalytic capacity. Production rates of more than 50 g of L-phosphinothricin per liter of column per h were obtained.

Stereospecific production of the herbicide phosphinothricin (glufosinate) by transamination: cloning, characterization, and overexpression of the gene encoding a phosphinothricin-specific transaminase from Escherichia coli.

We have cloned the gene encoding a 43-kilodalton transaminase from Escherichia coli K-12 with a specificity for L-phosphinothricin [L-homoalanine-4-yl-(methyl)phosphinic acid], the active ingredient of the herbicide Basta (Hoechst AG). The structural gene was isolated, together with its own promoter, and shown to be localized on a 1.6-kilobase DraI-BamHI fragment. The gene is subject to catabolite repression by glucose; however, repression could be relieved completely when 4-aminobutyrate (GABA) served as the sole nitrogen source. The regulation pattern obtained and a comparison of the restriction map of the initially cloned 15-kilobase SalI fragment with the physical map of the E. coli K-12 genome suggest that the cloned gene is identical with gabT, a locus on the gab gene cluster of E. coli K-12 which codes for the GABA:2-ketoglutartate transaminase (EC 2.6.1.19). A number of expression plasmids carrying the isolated transaminase gene were constructed. With these constructs, the transaminase expression in transformants of E. coli could be increased up to 80-fold compared with that in a wild-type control, and the transaminase constituted up to 20% of the total soluble protein of the bacteria. Thus, the protein crude extracts of the transformants could be used, after a simple heat precipitation step, for the biotechnological production of L-phosphinothricin in an enzyme reactor.

Young children's attribution of action to beliefs and desires.

When and how children understand beliefs and desires is central to whether they are ever childhood realists and when they evidence a theory of mind. Adults typically construe human action as resulting from an actor's beliefs and desires, a mentalistic interpretation that represents a common and fundamental form of psychological explanation. We investigated children's ability to do likewise. In Experiment 1, 60 subjects were asked to explain why story characters performed simple actions, such as looking under a piano for a kitten. Both preschoolers and adults gave predominantly psychological explanations, attributing the actions to the actor's beliefs and desires. Even 3-year-olds attributed actions to beliefs and false beliefs, demonstrating an understanding of belief not evident in previous research. In Experiment 2, 24 3-year-olds were tested further on their understanding of false belief. They were given both false belief prediction and explanation tasks. Children performed well on explanation taks, attributing an anomalous action to the actor's false belief, even when they failed to predict correctly what action would follow from a false belief. We concluded that 3-year-olds and adults share a fundamentally similar construal of human action in terms of beliefs and desires, even false beliefs.

Initial steps in the degradation of phosphinothricin (glufosinate) by soil bacteria.

Three hundred bacterial isolates from soil were tested for resistance against phosphinothricin [PPT; dl-homoalanin-4-yl(methyl)phosphinic acid], the active ingredient of the herbicide BASTA. Eight resistant bacterial strains and Escherichia coli were analyzed for PPT-transforming activities. At least three different enzymatic reactions could be detected in cell extracts. In six strains an acetyltransferase was active, synthesizing N-acetyl-PPT in the presence of PPT and acetyl coenzyme A. All strains could degrade PPT to its corresponding 2-oxoacid {2-oxo-4-[(hydroxy)(methyl)phosphinoyl] butyric acid} by transamination. Rhodococcus sp., the only tested strain that was able to utilize PPT as a sole source of nitrogen, formed 2-oxo-4[(hydroxy)(methyl)phosphinoyl]butyric acid by oxidative deamination. This enzymatic activity was inducible by l-glutamic acid or PPT itself but not in the presence of NH(4). d-PPT transformation was not detectable in any of the investigated strains.

Protection against the staphylococcal enterotoxin-induced intestinal disorder in the monkey by anti-idiotypic antibodies.

The staphylococcal enterotoxin serotype B (SEB)-induced enteric intoxication and the immediate-type reaction in the skin of unsensitized monkeys was used to define whether agents competing with SEB for target cell receptors may inhibit pathophysiological effects. For this purpose a duodenal provocation test was developed by use of a pediatric gastroscope, allowing the evaluation of the influence of antagonists on the intestinal disorder upon SEB challenge at the same duodenal site. First, carboxymethylation of histidine residues of SEB caused a complete loss of emetic and skin-sensitizing activity without changing the immunological specificity. However, carboxymethylated SEB is a strong inhibitor of enteric intoxications and immediate-type skin reactions upon SEB challenge. Second, after immunization of BALB/c mice with monoclonal anti-SEB antibodies, monoclonal antiidiotypic antibodies (anti-Id) were obtained by the "hybridoma technique" and purification by idiotype-affinity chromatography. Anti-Id specifically inhibited the binding of horseradish peroxidase-labeled anti-SEB to the ligand, and SEB blocked as well the interaction of these two antibody species, indicating a high degree of binding-site selectivity. Anti-Id completely protected against emetic response and diarrhea upon duodenal provocation with SEB and inhibited immediate-type skin reactions as well. Further, anti-Id acted as an antagonist without triggering biologic functions themselves. This shows that anti-Id constitute a useful tool to protect against a bacterial toxin-induced intestinal disorder.

Psychogenic habit cough: review and case reports.

Psychogenic habit cough--a condition that can be debilitating if it extends over a period of years--has been described in both pediatric and adolescent populations, but not in adults. The authors review the cases of 4 adult patients with this condition, review the available pediatric/adolescent literature, and make suggestions for the direction of future research. In some cases, psychogenic habit cough in adults can be successfully treated with a combination of psychotherapy, relaxation therapy, and speech therapy.

Infant search and object permanence: a meta-analysis of the A-not-B error.

Research on Piaget's stage 4 object concept has failed to reveal a clear or consistent pattern of results. Piaget found that 8-12-month-old infants would make perserverative errors; his explanation for this phenomenon was that the infant's concept of the object was contextually dependent on his or her actions. Some studies designed to test Piaget's explanation have replicated Piaget's basic finding, yet many have found no preference for the A location or the B location or an actual preference for the B location. More recently, researchers have attempted to uncover the causes for these results concerning the A-not-B error. Again, however, different studies have yielded different results, and qualitative reviews have failed to yield a consistent explanation for the results of the individual studies. This state of affairs suggests that the phenomenon may simply be too complex to be captured by individual studies varying 1 factor at a time and by reviews based on similar qualitative considerations. Therefore, the current investigation undertook a meta-analysis, a synthesis capturing the quantitative information across the now sizable number of studies. We entered several important factors into the meta-analysis, including the effects of age, the number of A trials, the length of delay between hiding and search, the number of locations, the distances between locations, and the distinctive visual properties of the hiding arrays. Of these, the analysis consistently indicated that age, delay, and number of hiding locations strongly influence infants' search. The pattern of specific findings also yielded new information about infant search. A general characterization of the results is that, at every age, both above-chance and below-chance performance was observed. That is, at each age at least 1 combination of delay and number of locations yielded above-chance A-not-B errors or significant perseverative search. At the same time, at each age at least 1 alternative combination of delay and number of locations yielded below-chance errors and significant above-chance correct performance, that is, significantly accurate search. These 2 findings, appropriately elaborated, allow us to evaluate all extant theories of stage 4 infant search. When this is done, all these extant accounts prove to be incorrect. That is, they are incommensurate with one aspect or another of the pooled findings in the meta-analysis. Therefore, we end by proposing a new account that is consistent with the entire data set.

Anti-idiotypic antibodies that inhibit immediate-type skin reactions in unsensitized monkeys on challenge with staphylococcal enterotoxin.

The staphylococcal enterotoxin B (SEB)-induced immediate-type skin reaction in unsensitized monkeys was used as a nonimmunological mast cell stimulus to examine whether the toxin exerts its effect via specific receptors on the target cell membrane. Anti-idiotypic antibodies (anti-Id) were raised in BALB/c mice against monoclonal anti-SEB antibodies (anti-SEB) and purified by idiotype affinity chromatography. The anti-Id nature of the antibody was demonstrated by its ability to inhibit the binding of 125I-labeled anti-SEB to the ligand in a concentration-dependent manner. Moreover, binding of anti-SEB to anti-Id was antagonized by the SEB ligand in a competitive way. These antibodies completely abolished skin reactions in unsensitized monkeys on challenge with SEB and impeded those provoked by staphylococcal enterotoxins A and C1 but did not have the biological activity of the toxin. These data are compatible with the view that receptors for staphylococcal enterotoxins may exist on the membrane of mast cells in the skin of unsensitized monkeys. The data suggest an experimental approach for producing anti-cell receptor antibodies that are of potential value to influence the course of staphylococcal enterotoxin-mediated effects.

Molecular cloning of a pea mRNA encoding an early light induced, nuclear coded chloroplast protein.

cDNA clones were isolated for a chloroplast protein, the mRNA of which is induced to maximum levels within 2-4 h after onset of illumination in five day old, etiolated pea seedlings.The cDNA library was constructed from poly(A)(+)-mRNA which was isolated from 4 h illuminated seedlings. The extremely short induction period of the early light induced protein(ELIP)-mRNA established the basis of our screening procedure. Colony hybridization experiments were performed with(32)P-labelled cDNA probes, synthesized from RNA of seedlings which had been exposed to different programs of illumination. Plasmid DNAs were isolated from colonies showing strong hybridization signals exclusively with cDNA corresponding to the 4 h-mRNA. Hybrid released translation of preselected plasmids p 17/C2 and p17/C4 revealed a peptide of Mr 24 000. After posttranslational importin vitro, the processed product of Mr 17 000 appears in the chloroplast. Using these clones, the expression of the ELIP-mRNA was investigated by DOT-hybridization. The ELIP-mRNA reaches maximum levels within 2-4 hours after onset of illumination. Our results correspond precisely to thein vivo characteristics and indicate positive identification of the sought clones.