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Biomedical subjects

J Preiss

Publications and source records attributed to J Preiss.

At least 91 records · Page 5Linked to original sources
PubMed1992

Interferon alpha-2b, 5-fluorouracil, and folinic acid combination therapy in advanced colorectal cancer: preliminary results of a phase I/II trial.

We conducted a phase I/II trial of 5-fluorouracil (5-FU)/folinic acid (FA) and alpha-2b interferon (IFN) in 43 previously untreated patients with measurable metastatic colorectal cancer. Patients had disease progression prior to therapy consistent of 5-FU 500 mg/m2 (level A) or 600 mg/m2 (level B) as starting dose administered as a 2-hour infusion, FA 200 mg/m2, and alpha-2b IFN 5MU/m2 subcutaneously. All drugs were given on days 1 to 5 and cycles were repeated after 3 to 4 weeks. The aim of the study was to define the maximal tolerable dose (MTD) of 5-FU for this schedule. In the absence of toxicity above MTD, defined as diarrhea and mucositis of World Health Organization grade 2 and leukopenia of World Health Organization, grade 3 5-FU was increased in increments of 100 mg/m2 for further cycles. Twenty-four patients were started on level A; 18 were started on level B. Forty-two patients were evaluable for toxicity, 32 for response. Three of 32 patients achieved a partial response; in 22 of 32 patients, tumor stabilization occurred. Forty percent of patients started on level A developed grade 2 diarrhea; 28% of patients developed grade 2 or 3 mucositis. Of 18 patients on level B, two patients experienced grade 4 mucositis (11%) and grade 3 or 4 diarrhea (11%). One drug-related death in the presence of sepsis occurred. Due to 11% of patients with grade 4 toxicity when started on 600 mg/m2 5-FU and 40% of patients with grade 2 diarrhea when started on level A, MTD as starting dose for 5-FU is 500 mg/m2 as a 2-hour infusion when used in combination with FA and IFN on a day 1 to 5 active schedule. We observed a wide range of 5-FU dose tolerated by individual patients. While some patients experienced severe, mainly gastrointestinal, toxicity on lower levels of 5-FU, others tolerated much higher 5-FU doses (11 patients, 700 mg/m2; 5 patients, 800 mg/m2; and one patient, 900 mg/m2). Our data suggest that double modulation of 5-FU by FA and IFN may not be superior to modulation of 5-FU with either drug alone.

Adult↗
PubMed1992

Fluorouracil plus interferon + folinic acid in regional and systemic therapy in colorectal cancer.

Modulation of fluorouracil (5-FU) therapy with folinic acid (FA) in advanced colorectal adenocarcinoma produces a doubled increase in remission rates, but one important study with another modulation of 5-FU with interferon alfa 2a (IFN-alpha-2a) had better results. In the fact that both modulations have different mechanisms of interaction, it seems hopeful that both have additive or synergistic effects and they give further increases of remission rates. Twenty-three patients with proven progressive advanced colorectal cancer were treated with 5-FU, FA, and IFN-alpha-2a in three different regimens. IFN-alpha-2a was given subcutaneously in a dose of 5 MioU per day during 5-FU treatment. In the first regimen FA (500 mg/m2) was administered in a short time infusion (30 minutes) while 5-FU (1,000 mg/m2) was given in a 24-hour continuous infusion with weekly intervals. In regimen 2 and 3 FA/5-FU treatment was given on days 1 to 5 every 3 to 4 weeks. The difference was time and dose of the 5-FU infusion (600 mg/m2 in 4-hour versus 350 mg/m2 by push injection). Three patients with isolated liver metastases and one patient with local recurrence of rectal carcinoma received regional therapy with higher 5-FU doses. Dose escalation was carried out in all regimens when possible (toxicity less than or equal to World Health Organization [WHO] grade 1). Objective response rates were reached in nine of 23 patients with two complete remissions (one in regional and systemic treatment) with a mean duration of 6 months. Eight patients had stable disease and six had therapy failure. In pretreated patients no complete remission was reached; three of seven had partial remission, two no change, and two progressive disease. Severe toxicity occurred in 12 of 23 patients with mucositis WHO grade 4 in one patient, grade 3 in two, hematotoxicity grade 3 in three, and cutaneous toxicity in two. Eight of twenty-three patients had alopecia grades 2 and 3. These early results of our phase II study with limited patients shows no significant advantage of double modulation in comparison to the well established results of FA/5-FU therapy.

Adult↗
PubMed1991

Biosynthesis of bacterial glycogen. Mutagenesis of a catalytic site residue of ADP-glucose pyrophosphorylase from Escherichia coli.

Site-directed mutagenesis was used to explore the role of Lys-195 in ADP-glucose pyrophosphorylase from Escherichia coli. This residue, which is conserved in every bacterial and plant source sequenced to date, was originally identified as a potential catalytic site residue by covalent modification studies. Mutation of Lys-195 to glutamine produces an enzyme whose Km for glucose 1-phosphate is 600-fold greater than that measured for the wild-type enzyme. The effect on glucose 1-phosphate is very specific since kinetic constants measured for ATP, Mg2+, and the allosteric activator, fructose 1,6-bisphosphate, are unchanged relative to those measured for the wild-type enzyme. Furthermore, the catalytic rate constant, Kcat, for the glutamine mutant is similar to that of the wild-type enzyme. Taken together, the results suggest a role for Lys-195 in binding of glucose 1-phosphate and exclude its role as a participant in the rate-determining step(s) in the catalytic reaction mechanism. To further study the effect of charge, shape, size, and hydrophobicity of the amino acid residue at position 195, a series of mutants were prepared including arginine, histidine, isoleucine, and glutamic acid. In every case, the kinetic constants measured for ATP, Mg2+, and fructose 1,6-bisphosphate were similar to wild-type constants, reinforcing the notion that this residue is responsible for a highly localized effect at the glucose 1-phosphate-binding site and also suggesting that the protein can accommodate a wide range of substitutions at this position without losing its global folding properties. Thermal stability measurements corroborate this finding. The mutations did, however, produce a range of glucose 1-phosphate Km values from 100- to 10,000-fold greater than wild-type, which indicate that both size and charge properties of lysine are essential for proper binding of glucose 1-phosphate at the catalytic site. AMP binding was also affected by the nature of the mutation at position 195. A model for glucose 1-phosphate, ATP, and AMP binding is presented.

Base Sequence↗
PubMed1991

Molecular characterization of the gene encoding a rice endosperm-specific ADPglucose pyrophosphorylase subunit and its developmental pattern of transcription.

The gene encoding a rice endosperm-specific ADPglucose pyrophosphorylase (AGPP) subunit was isolated and its structure determined by nucleotide (nt) sequencing. A comparison of the genomic and cDNA nt sequences revealed a complex gene structure with ten exons and nine introns spanning over 6 kb. The exons ranged in size from 293 to 99 nt and the introns were between 1435 and 84 nt in size, with the first intron being the largest. All of the intron splice sites, except intron-2, contained GT/AG borders and were similar to the published splice site consensus sequences. Intron-2 had CA/CC borders at the 5' and 3' ends, but sequences adjacent to the splice site borders shared homology to the splice site consensus sequence, suggesting that the overall splice region, rather than the specific GT/AG sequence, determines the splice site. Several sequence motifs which may play a role in the regulation of plant genes were evident upstream from the transcriptional start point. Analysis of the developmental pattern of expression revealed a maximum level of mRNA transcripts for AGPP at five days after flowering coincident with starch accumulation. This result suggests that starch biosynthesis is controlled at the transcriptional level during seed development.

Amino Acid Sequence↗
PubMed1991

Comparison of the primary sequences of two potato tuber ADP-glucose pyrophosphorylase subunits.

Near-full-length cDNA clones to the small and large subunit of the heterotetrameric potato tuber ADP-glucose pyrophosphorylase have been isolated and characterized. The missing amino terminal sequence of the small subunit has also been elucidated from its corresponding genomic clone. Primary sequence comparisons revealed that each potato subunit had less identity to each other than to their homologous subunit from other plants. It also appeared that the smaller subunit is more conserved among the different plants and the larger subunit more divergent. Amino acid comparisons of both potato tuber sequences to the Escherichia coli ADP-glucose pyrophosphorylase sequence revealed conserved regions important for both catalytic and allosteric function of the bacterial enzyme.

Amino Acid Sequence↗
PubMed1991

[Problems in recognizing recurrences of craniocervical malignancies in CT].

A retrospective study was carried out involving 39 patients with malignant tumours of the head and neck in order to differentiate by CT changes due to local tumour or lymph node recurrences and various post-operative or post-irradiation changes. In these patients, two or more CT examinations of the neck were available, derived from a fourth generation scanner. The suspected lesions were classified according to their type and the appearances correlated with the treatment and clinical findings. There was frequent overlap between the appearances of benign and malignant changes, so that in individual cases a definite decision often proved impossible.

Adult↗
PubMed1991

Mutants of Arabidopsis with altered regulation of starch degradation.

Mutants of Arabidopsis thaliana (L.) Heynh. with altered regulation of starch degradation were identified by screening for plants that retained high levels of leaf starch after a period of extended darkness. The mutant phenotype was also expressed in seeds, flowers, and roots, indicating that the same pathway of starch degradation is used in these tissues. In many respects, the physiological consequences of the mutations were equivalent to the effects observed in previously characterized mutants of Arabidopsis that are unable to synthesize starch. One mutant line, which was characterized in detail, had normal levels of activity of the starch degradative enzymes alpha-amylase, beta-amylase, phosphorylase, D-enzyme, and debranching enzyme. Thus, it was not possible to establish a biochemical basis for the phenotype, which was due to a recessive mutation at a locus designated sex1 at position 12.2 on chromosome 1. This raises the possibility that hitherto unidentified factors, altered by the mutation, play a key role in regulating or catalyzing starch degradation.

Journal Article↗
PubMed1991

Regulatory and Structural Properties of the Cyanobacterial ADPglucose Pyrophosphorylases.

ADPglucose pyrophosphorylase (EC 2.7.7.27) has been purified from two cyanobacteria: the filamentous, heterocystic, Anabaena PCC 7120 and the unicellular Synechocystis PCC 6803. The purification procedure gave highly purified enzymes from both cynobacteria with specific activities of 134 (Synechocystis) and 111 (Anabaena) units per milligram protein. The purified enzymes migrated as a single protein band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with molecular mass corresponding to 53 (Synechocystis) and 50 (Anabaena) kilodaltons. Tetrameric structures were determined for the native enzymes by analysis of gel filtrations. Kinetic and regulatory properties were characterized for the cyanobacterial ADPglucose pyrophosphorylases. Inorganic phosphate and 3-phosphoglycerate were the most potent inhibitor and activator, respectively. The Synechocystis enzyme was activated 126-fold by 3-phosphoglycerate, with saturation curves exhibiting sigmoidicity (A(0.5) = 0.81 millimolar; n(H) = 2.0). Activation by 3-phosphoglycerate of the enzyme from Anabaena demonstrated hyperbolic kinetics (A(0.5) = 0.12 millimolar; n(H) = 1.0), having a maximal stimulation of 17-fold. I(0.5) values of 95 and 44 micromolar were calculated for the inhibition by inorganic phosphate of the Synechocystis and Anabaena enzyme, respectively. Pyridoxal-phosphate behaved as an activator of the cyanobacterial enzyme. It activated the enzyme from Synechocystis nearly 10-fold with high apparent affinity (A(0.5) = 10 micromolar; n(H) = 1.8). Phenylglyoxal modified the cyanobacterial enzyme by inactivating the activity in the presence of 3-phosphoglycerate. Antibody neutralization experiments showed that anti-spinach leaf (but not anti-Escherichia coli) ADPglucose pyrophosphorylase serum inactivated the enzyme from cyanobacteria. When the cyanobacterial enzymes were resolved on sodium dodecyl sulfate- and two-dimensional polyacrylamide gel electrophoresis and probed with Western blots, only one protein band was recognized by the anti-spinach leaf serum. The same polypeptide strongly reacted with antiserum prepared against the smaller spinach leaf 51 kilodalton subunit, whereas the anti-54 kilodalton antibody raised against the spinach subunit reacted weakly to the cyanobacterial subunit. Regulatory and immunological properties of the cyanobacterial enzyme are more related to the higher plant than the bacterial enzyme. Despite this, results suggest that the ADPglucose pyrophosphorylase from cyanobacteria is homotetrameric in structure, in contrast to the reported heterotetrameric structures of the higher plant ADPglucose pyrophosphorylase.

Journal Article↗
PubMed1991

[Epilepsy and rehabilitation using psychological methods (psychotherapy)].

41 subjects (39 patients and two mothers) participated in 1983-1988 in group psychotherapy for ambulatory adult patients focused on psychosocial problems and questions of the therapeutic regime. According to the independent evaluation by the attending physician the individually outlined goal was achieved in 38% of the patients. 31% of the patients left without a special effect, however, participation in the group meant probably a certain enrichment of their life. Another 31% of the patients had a negative attitude to participation in the group. Successful patients attended much longer, had a significantly higher verbal IQ and all responded reliably in Washington's psychosocial seizure inventory--WPSI. Conversely, 27% of the other patients gave invalid responses to WPSI. Psychological procedures should be part of a wider scale of rehabilitation approaches to epilepsy, including occupational rehabilitation and club activities.

Adult↗
PubMed1991

[The Halstead-Reitan Neuropsychological Battery. Initial experience with its use in Czechoslovakia].

The authors present information on the basic, classical neuropsychological battery and its Czech application in adults (15 years and older). Its principles are adopted also by more recent partial methods. The validity of new methods is usually assessed by comparison with the former method. Its position in neuropsychology can be compared with that of Wechsler's intelligence scales in the area of assessment of intelligence. The author describes 11 original tests which form the battery. Although individual tests are used also separately, it is better not to deprive oneself of the comprehensive evaluation of cerebral functions and psychic consequences of neurological disease or damage. The authors assembled encouraging experience when examining the first 72 Czech patients--ambulatory psychiatric patients (n = 12), patients rated for invalidity (n = 20), epileptic patients (n = 33) and patients from the clinic for occupational diseases (n = 7).

Czechoslovakia↗
PubMed1990

Identification of lysine 15 at the active site in Escherichia coli glycogen synthase. Conservation of Lys-X-Gly-Gly sequence in the bacterial and mammalian enzymes.

Glycogen synthases from Escherichia coli and mammalian muscle differ in many respects including regulation, sugar nucleotide specificity, and primary sequence. To compare the structure of the active sites in these enzymes, the affinity-labeling study of the E. coli enzyme was carried out using adenosine diphosphopyridoxal as the reagent. The E. coli enzyme was inactivated in a time- and dose-dependent manner when incubated with the reagent followed by sodium borohydride reduction. The inactivation was markedly protected by ADP-glucose and ADP, suggesting that the reagent was bound to the substrate-binding site. The stoichiometry of the bound reagent to the enzyme was approximately 1:1. Sequence analysis of the labeled peptide isolated from a proteolytic digest of the modified protein revealed that Lys15 is labeled. Based on the geometry of the reagent, the epsilon-amino group of this residue might be located close to the pyrophosphate moiety of ADP-glucose bound to the E. coli enzyme, like that of Lys38 in the rabbit muscle enzyme, which is labeled by uridine diphosphopyridoxal (Tagaya, M., Nakano, K., and Fukui, T. (1986) J. Biol. Chem. 260, 6670-6676; Mahrenholz, A. M., Wang, Y., and Roach, P. J. (1988) J. Biol. Chem. 263, 10561-10567). The importance of the conserved sequence of Lys-X-Gly-Gly is discussed in connection with the glycine-rich region found in many nucleotide-binding proteins.

Amino Acid Sequence↗
PubMed1990

Escherichia coli E-39 ADPglucose synthetase has different activation kinetics from the wild-type allosteric enzyme.

Kinetic and binding studies have shown that Lys39 of Escherichia coli ADPglucose synthetase is involved in binding of the allosteric activator. In order to study structure-function relationships at the activator binding site, this lysine residue was substituted by glutamic acid (Lys39----Glu) by site-directed mutagenesis. The resultant mutant enzyme (E-39) showed activation kinetics different from those of the wild-type enzyme. The level of activation of the E-39 enzyme by the major activators of E. coli ADPglucose synthetase, 2-phosphoglycerate, pyridoxal phosphate, and fructose-1,6-phosphatase was only approximately 2-fold compared to activation of 15- to 28-fold respectively, for the wild-type enzyme. NADPH, an activator of the wild-type enzyme, was unable to activate the mutant enzyme. In addition, the concentrations of the above activators necessary to obtain 50% of the maximal stimulation of enzyme activity (A0.5) were 5-, 9-, and 23-fold higher, respectively, than those for the wild-type enzyme. The E-39 enzyme also had a lower apparent affinity (S0.5) for the substrates ATP and MgCl2 than the wild-type enzyme and the values obtained in the presence or absence of activator were similar. The concentration of inhibitor giving 50% of enzyme activity (I0.5) was also similar for the E-39 enzyme in the presence or absence of activator. These results indicate that the E-39 mutant enzyme is not effectively activated by the major activators of the E. coli ADPglucose synthetase wild-type enzyme, and that this amino acid substitution also prevents the allosteric effect that the activator has on the wild-type enzyme kinetics, either increasing its apparent affinity for the substrates or modulating the enzyme's sensitivity to inhibition.

Allosteric Regulation↗
PubMed1990

Molecular Characterization of the Brittle-2 Gene Effect on Maize Endosperm ADPglucose Pyrophosphorylase Subunits.

Activity of the enzyme ADPglucose pyrophosphorylase is known to be reduced in maize (Zea mays L.) endosperm mutants at two independent loci, Shrunken-2 (Sh(2)) and Brittle-2 (Bt(2)). Spinach leaf ADPglucose pyrophosphorylase has previously been shown to comprise two subunits of 51 and 54 kilodaltons. Anti-bodies raised to each of the two subunits of spinach leaf ADPglucose pyrophosphorylase were found to cross-react to different bands on Western blots prepared from polyacrylamide gel electrophoresis separated wild-type maize endosperm proteins. The anti-spinach leaf 51 kilodalton subunit antibody cross-reacted with a 55 kilodalton maize endosperm protein and the anti-spinach leaf 54 kilodalton subunit antibody cross-reacted with a 60 kilodalton maize endosperm protein. These immunological reactions were observed in maize endosperm extracts and with a highly purified preparation of maize endosperm ADPglucose pyrophosphorylase. Mutant bt(2) endosperm lacked the 55 kilodalton subunit while mutant sh(2) endosperm lacked the 60 kilodalton subunit on Western blots. These results suggest that the maize endosperm ADPglucose pyrophosphorylase is made up of two immunologically dissimilar subunits and that the bt(2) and sh(2) mutations cause reduction in ADPglucose pyrophosphorylase activity through the lack of one of these two subunits. An ADPglucose pyrophosphorylase cDNA clone antigenically selected from a rice seed cDNA expression library was found to hybridize strongly with a cDNA corresponding to a maize endosperm transcript which is absent in a W64A bt(2) mutant. Thus, the bt(2) mutant causes the absence not only of the small subunit but of the corresponding transcript. Bt(2) is implicated as the structural gene for the small (54 kilodalton) subunit of maize endosperm ADPglucose pyrophosphorylase.

Journal Article↗