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Biomedical subjects

E Holler

Publications and source records attributed to E Holler.

At least 145 records · Page 8Linked to original sources
PubMed1989

Investigations on the antiproliferative effects of amino acid antagonists targeting for aminoacyl-tRNA synthetases. Part II--The antileukemic effect.

Amino acid antagonists with proven or potential inhibitory activities on aminoacyl-tRNA synthetases were tested for their antiproliferative effect against the murine leukemic cell line P388D1. Micromolar concentrations of the compounds S-tritylcysteine (18), fenitropan and beta-chloroalanine gave strong growth inhibition. In the mouse only 18 was effective against leukemia P388 (T/C = 211%). The inhibitory effect on aminoacyl-tRNA synthetases and the antiproliferative action on P388D1 or P388 could not be correlated.

Amino Acids↗
PubMed1989

Bone marrow transplantation for the treatment of leukaemia-results of the Munich Cooperative Group.

This report summarizes the results of marrow transplantation from HLA-identical siblings and syngeneic twins for treatment of acute myelogenous leukaemia, chronic myelogenous leukaemia, acute lymphoblastic and undifferentiated leukaemia from 1975 until December 1986. Three conditioning regimens and treatment of the marrow graft in vitro with absorbed antithymocyte globulin or the monoclonal antibody "Campath 1" for prophylaxis of graft-versus-host disease (GVHD) have been studied and analyzed retrospectively. The regimen of total body irradiation in large fractions of 4 Gy and of cyclosphosphamide (200 mg/kg) has achieved the most favorable results. Inactivation of T-cells by treatment of the marrow "in vitro" has decreased the severity of GVHD without improving survival. The antileukaemic effect of the graft may be important for control of the disease and may be improved by better immunosuppression of the recipient.

Adolescent↗
PubMed1989

Improved survival following HLA-incompatible bone marrow transplantation. Munich Cooperative Group of Bone Marrow Transplantation.

In most centers allogeneic bone marrow transplantation is restricted to patients with HLA-identical siblings as donors. We have transplanted 16 patients with marrow of donors other than HLA-identical siblings. Seven patients were grafted in the years 1978 until 1984. Six died of transplant complications and one of recurrent leukemia. More recently 9 patients were transplanted following an improved immunosuppressive conditioning treatment derived from experimental studies in dogs. Four are alive and in continuous remission between more than 2 months and 2 years. 5 patients died, 3 from fungal infections, one from recurrent leukaemia and one early from endothelial leakage syndrome. Our results indicate that intensified immunosuppressive conditioning may improve the results of marrow transplantation from HLA-haploidentical donors.

Adolescent↗
PubMed1989

Myeloablative conditioning for marrow transplantation in myelodysplastic syndromes and paroxysmal nocturnal haemoglobinuria.

Paroxysmal nocturnal haemoglobinuria (PNH) and myelodysplastic syndromes (MDS) are disorders of pluripotent stem cells resulting in haematopoietic insufficiency which can be cured by marrow transplantation. The extent of myeloablative conditioning necessary for elimination of the non-malignant and premalignant clones is not known. We report our results of marrow transplantation with and without myeloablative conditioning in two patients with PNH and seven patients with MDS. Conditioning was not used in a patient with PNH and a monozygotic twin as donor. In this patient the disease remained unchanged. Myeloablative treatment with busulphan (BUS) in addition to immunosuppression with cyclophosphamide (CY) was used for conditioning in a patient with PNH and a 2-year-old boy with chronic myelomonocytic leukaemia (CMML). Fractionated total body irradiation (FTBI) and CY was used in six patients with refractory anaemia with excess of blasts (RAEB) and RAEB in leukaemic transformation (RAEB-T). Haematopoiesis was fully restored in all patients conditioned with myeloablative treatment except for a patient in leukaemic transformation with myelofibrosis and a HLA-DR-incompatible donor. Chimerism was complete in all patients except for the 2-year-old boy conditioned with BUS and CY. Our results and those reviewed in the literature indicate that myeloablative conditioning with either BUS or FTBI is advantageous for marrow transplantation in PNH and MDS.

Adult↗
PubMed1988

Purification and characterization of DNA polymerase alpha from plasmodia of Physarum polycephalum.

DNA polymerase alpha from Physarum polycephalum has been purified from freshly harvested microplasmodia. An inhibitory activity was removed by precipitation with poly(ethyleneimine) and interfering type-beta-like DNA polymerase by chromatography on phosphocellulose. The preparation was free of endonucleases and exonucleases. The DNA-polymerizing polypeptide had a molecular mass of 140 kDa by polyacrylamide gel electrophoresis under denaturing conditions. It was contained in purified samples and in crude cell extracts. Peptides of smaller size that reacted with antibodies against this protein were generated during purification and prolonged standing. Molecular sizing under non-denaturing conditions resulted in high-molecular-mass forms. The type of isolated DNA polymerase was established on the basis of inhibition and template-primer utilization experiments underlying the classification of DNA polymerases from higher eucaryotes. The majority of the DNA-polymerizing activity was contained in the cell nucleus fraction and was inhibited by aphidicolin. The isoelectric point (pI) was 6.7 +/- 0.2, the pH optimum at pH 6.8, and the temperature optimum at 40 degrees C. Monovalent salts, Li+, Na+, NH+4, K+, were inhibitory except for small activation maxima at 10 mM, 75 mM and 100 mM in the case of Na+, NH+4 and K+ respectively. The bivalent cations Mg2+ and Mn2+ had broad activity maxima at 3-20 mM concentrations, which were shifted to 0.05-0.1 mM in the case of Mn2+ and synthetic DNA homopolymers. The numbers of molecules of DNA polymerases in Physarum nuclei were calculated and compared with the established number of replicons in plasmodia and with the number of molecules of DNA polymerases in higher eucaryotes.

Cell Nucleus↗
PubMed1988

Effects of coordination of diammineplatinum(II) with DNA on the activities of Escherichia coli DNA polymerase I.

The effects of the reaction of cis- and trans-diamminedichloroplatinum(II) with DNA have been measured with regard to DNA synthesis, 3'-5' exonuclease (proofreading), and 5'-3' exonuclease (repair) activities of Escherichia coli DNA polymerase I. Both isomers inhibit DNA synthetic activity of the polymerase through an increase in Km values and a decrease in Vmax values for platinated DNA but not for the nucleoside 5'-triphosphates as the varied substrates. The inhibition is a consequence of lowered binding affinity between platinated DNA and DNA polymerase, and of a platination-induced separation of template and primer strands. Strand separation enhances initial rates of 3'-5' excision of [3H]dCMP from platinated DNA (proofreading), while total excision levels of nucleotides are decreased. In contrast to proofreading activity, the 5'-3' exonuclease activity (repair) discriminates between DNA which had reacted with cis- and with trans-diamminedichloroplatinum(II). While both initial rates and total excision are inhibited for the cis isomer, they are almost not affected for the trans isomer. This differential effect could explain why bacterial growth inhibition requires much higher concentrations of trans- than cis-diamminedichloroplatinum(II).

Antineoplastic Agents↗
PubMed1988

Synthesis of diadenosine 5',5'''-P1,P4-tetraphosphate (AppppA) from adenosine 5'-phosphosulfate and adenosine 5'-triphosphate catalyzed by yeast AppppA phosphorylase.

A novel way of enzymatic synthesis of diadenosine 5',5"'-P1,P4-tetraphosphate (AppppA), which does not involve aminoacyl-tRNA synthetases, has been discovered. Yeast AppppA alpha, beta-phosphorylase catalyzes irreversible conversion of adenosine 5'-phosphosulfate (APS) and ATP into AppppA according to the equation APS + ATP----AppppA + sulfate. In this reaction, the enzyme exhibits a broad pH optimum (between 6 and 8) and requires Mn2+, Mg2+, or Ca2+ ions for activity, with Mn2+ being twice as effective as Mg2+ or Ca2+ at optimal concentration (0.5 mM). The Km values computed for APS and ATP are 80 microM and 700 microM, respectively. The rate constant for the AppppA synthesis is 3 s-1 (pH 8.0, 30 degrees C, 0.5 mM MgCl2). Some ATP analogues like ppppA, GTP, adenosine 5'-(alpha, beta-methylenetriphosphate), and adenosine 5'-(beta, gamma-methylenetriphosphate), but not dATP, UTP, or CTP, are also substrates for AppppA phosphorylase and accept adenylate from APS with the formation of AppppA, AppppG, Appp(CH2)pA, and App(CH2)ppA, respectively. Functional versatility of yeast AppppA phosphorylase may provide a link between metabolism of AppppA on one hand and metabolism of APS and phosphate on the other and raises the possibility of participation of AppppA in regulation of metabolism of APS and/or inorganic phosphate in yeast.

Adenine Nucleotides↗
PubMed1987

The binding of adenosine(5')tetraphospho(5')adenosine to calf thymus histones measured by non-equilibrium dialysis.

Binding of adenosine(5')tetraphospho(5')adenosine (Ap4A) to histones of calf thymus was investigated by non-equilibrium dialysis. Histone H1 interacts with the dinucleotide via two strong sites and competes with Mg2+ ions. Intrinsic dissociation constants were 1.6 +/- 0.1 microM and 11 +/- 1 microM for zero and 0.4 mm-Mg2+ concentration respectively. Binding of poly(dT) and of other nucleotides to histone H1 was measured in an [3H]Ap4A-competition assay. The tendency to form complexes among nucleotides was highest for bisnucleoside tetraphosphates and decreased in the order poly(dT) greater than or equal to Ap4A approximately Gp4G greater than Ap4 much greater than Ap3A approximately Ap5A greater than or equal to ATP, GTP and dTTP. The co-ordination complex derived from Ap4A and cis-diammine-dichloroplatinum(II) was not reactive. The other histones of calf thymus also bound Ap4A with affinities decreasing in the order H4 approximately H3 greater than H1 greater than H2b greater than H2a. Ap4A stimulated the exchange of histone H1 between nucleosomes, but this effect was referred to ionic strength. It did not bind to assembled nucleosomes. Binding of Ap4A to histone H1 was decreased by salt (NaCl). At physiological saline concentration the value of the dissociation constant is commensurable with the value of the Ap4A concentration in the nucleus and thus indicative of complex-formation in vivo.

Adenine Nucleotides↗
PubMed1987

Mode of inhibition of the DNA polymerase of Methanococcus vannielii by aphidicolin.

The mode of action of aphidicolin on DNA synthesis catalysed by the DNA polymerase of Methanococcus vannielii is competitive for dCTP, noncompetitive for dATP, dGTP and dTTP and uncompetitive for activated DNA. The kinetic data are accounted for by a mechanism in which dCTP and aphidicolin compete for the dCTP-specific binding site on the DNA polymerase. The dissociation constant for the aphidicolin--DNA-polymerase complex is 0.04-0.07 microM. Similar modes of inhibition of DNA synthesis exist for DNA polymerase alpha of higher eucaryotes but not for eubacteria or viruses and suggests a close functional relationship between the DNA polymerase of eucaryotes and of the archaebacterium M. vannielii.

Aphidicolin↗
PubMed1987

A DNA polymerase with unusual properties from the slime mold Physarum polycephalum.

Two forms of a DNA polymerase have been purified from microplasmodia of Physarum polycephalum by poly(ethyleneimine) precipitation and chromatography on DEAE-Sephacel, phosphocellulose, heparin Sepharose, hydroxyapatite, DNA-agarose, blue-Sepharose. They were separated from DNA polymerase alpha on phosphocellulose and from each other on heparin-Sepharose. Form HS1 enzyme was 30-40% pure and form HS2 enzyme 60% with regard to protein contents of the preparations. Form HS2 enzyme was generated from form HS1 enzyme on prolonged standing of enzyme preparations. The DNA polymerases were obtained as complexes of a 60-kDa protein associated with either a 135-kDa (HS1) or a 110-kDa (HS2) DNA-polymerizing polypeptide in a 1:1 molar stoichiometry. The biochemical function of the 60-kDa protein remained unknown. The complexes tended to dissociate during gradient centrifugation and during partition chromatography as well as during polyacrylamide gradient gel electrophoresis under nondenaturing conditions at high dilutions of samples. Both forms existed in plasmodia extracts, their proportions depending on several factors including those which promoted proteolysis. The DNA polymerases resembled eucaryotic DNA polymerase beta by several criteria and were functionally indistinguishable from each other. It is suggested that lower eucaryotes contain repair DNA polymerases, which are similar to those of eubacteria on a molecular mass basis.

Centrifugation, Density Gradient↗
PubMed1987

Kinetic investigation of the DNA platination reaction: evidence for a transient adduct between deoxyribonucleic acid and cis-platinum(II).

Kinetics of the synthesis of adducts between salmon testis DNA and platinum(II) compounds were measured by their effects on DNA synthesis, circular dichroism, and ethidium bromide dependent fluorescence. Transient incorporation of [14C]cyanide into DNA adducts of of cis-diammineaquochloroplatinum(II) and respectively cis-diamminediaquoplatinum(II) compounds but not of trans-diammineaquochlorplatinum(II) was observed. A minimal kinetic scheme is derived, in which a transient monodentate DNA-platinum(II) adduct is formed in a bimolecular reaction between DNA and aquated platinum(II) compounds. Second-order rate constants are 2000-3000 M-1 min-1 for cis-diamminediaquoplatinum(II) and 280-400 M-1 min-1 for cis- and trans-diammineaquochloroplatinum(II), respectively. The dependence of pseudo-first-order rate constants is not linear for high concentrations of DNA, suggesting competitive formation of more than one primary adduct. The monodentate adducts inhibit DNA polymerase catalyzed DNA synthesis. The biomolecular reaction is followed by a rearrangement (rate constant 0.22 min-1) that gives rise to most of the decrease in the fluorescence intensity and that depends on the state of aquation of the DNA-bound platinum(II) complex. By exchange of coordinated water with a second nucleotide, the monodentate adduct can form cross-links in a reaction joining the rearrangement. Adducts containing a chloro group liberate it by hydrolysis prior to cross-linking. In the case of the trans-platinum(II) adduct, the hydrolysis is aided by the trans effect of the bound first nucleotide.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

In vitro competition between adenosine(5')tetraphospho(5')adenosine and deoxyribonucleic acid in the reaction with diamminedichloroplatinum(II).

Competition between adenosine(5')tetraphospho(5')adenosine (Ap4A) and DNA for the synthesis of adducts with the cis or trans isomer of diamminedichloroplatinum(II) was measured in the presence and absence of magnesium and spermidinium ions. Reaction products were analysed by circular dichroism, poly(ethyleneimine) thin-layer chromatography and reversed-phase chromatography. Competition was affected by the oligovalent cations that bound specifically to the dinucleotide. Platination of DNA was favoured under all conditions. Chromatin was less competitive. The mechanism was kinetic competition, DNA reacting considerably faster than Ap4A. Platinum(II) did not exchange between adducts and free DNA and Ap4A, respectively. On that basis only low amounts of Ap4A adducts were estimated to be formed under conditions of clinical chemotherapy. The cis and trans isomers of diamminedichloroplatinum(II) were equally effective. Platinum(II) adducts of Ap4A were neither degraded by Ap4A-specific pyrophosphohydrolases nor by phosphodiesterase nor in the presence of unfractionated extract of calf thymus. Unphysiologically high concentrations of Crotalus durissus phosphodiesterase I were required for hydrolytic splitting, the amount of which was similar for both platinum(II) isomer adducts. The results suggest that Ap4A platinum(II) adducts might accumulate during chemotherapy of cancer treatment.

Adenine Nucleotides↗