Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.5.1.1 (asparaginase)
2,695 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The feasibility of the immobilization of Escherichia coli L-asparaginase into a hydrogel matrix made of poly-(ethylene glycol) (PEG) and BSA was demonstrated. After immobilization a 200-fold increase in the Km value was observed. The use of an L-aspartic acid analogue, carbobenzoxy-L-aspartic acid and surface modification by methoxy-PEG of molecular mass 5 kDa cause a only a slight gain in affinity of the enzyme for its natural substrate. The immobilized L-asparaginase has an optimal activity over a larger range of pH than the native enzyme, owing to the effect of the matrix. At a physiological pH of 7.3, the immobilized enzyme retained 90% of its activity compared with only 43% for the native form. The immobilized enzyme retained a high proportion of its initial activity, more than 90% after 50 days of incubation at 37 degrees C, even in the presence of its substrate. This may be compared with a half-life of 2 days observed for native enzyme incubated under the same conditions. These results suggest that the BSA-PEG matrix can be very useful for enzyme immobilization and, taking into account the good biocompatibility of the matrix, one can expect that this matrix will provide a functional bioreactor for use in vivo.
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PMID:Immobilization of L-asparaginase into a biocompatible poly(ethylene glycol)-albumin hydrogel: I: Preparation and in vitro characterization. 867 8

The L-asparaginase of Escherichia coli (ASNase) is currently used in combination with antineoplastic drugs to treat various lymphoblastic leukaemias. However, its use is limited by severe immunological reactions and the short serum half-life associated with the enzyme. Immobilization of ASNase into a biocompatible matrix can greatly decrease the immunogenicity of the enzyme, increase its half-life in vivo and its therapeutic index. Thus the E. coli ASNase was immobilized in a biocompatible hydrogel made of rat serum albumin and poly(ethylene glycol) (PEG; molecular mass 10 kDa). The effectiveness of this enzymic bioreactor to deplete serum L-asparagine was evaluated after its peritoneal implantation in rats. Seven units of immobilized ASNase/rat depleted serum asparagine to an undetectable level (< 1 microM) during 6 days, while 5 units of immobilized ASNase/rat decreased the level of serum asparagine by 85-90% during at least 2 days. Under both conditions asparagine levels returned to normal about 10 days after surgery, and hydrogels still retained 80% of their enzymic activity when assayed in vitro. After 10-14 days in vivo, hydrogels became opaque and surrounded by a fibrotic capsule with a few inflammatory sites. Nevertheless, the enzymic hydrogel showed great stability in vivo, and, after 4 months of implantation, 12% of the initial ASNase activity was still present. At 6 months, histological analysis showed stabilization of the fibrotic capsule thickness. Assays on the levels of ASNase and asparagine synthetase indicated an induction of the latter activity, mainly in the pancreas when compared with the level observed in spleen or liver. ELISA tests at 28 days and 120 days showed the presence of anti-ASNase (and, in lower amounts, anti-PEG) antibodies in sera of implanted rats. As observed with other enzyme-immobilization systems used in vivo, the formation of fibroblast-like cell layers around the implant, which block the translocation of the substrate into the enzymic matrix, is the major factor affecting the performance and longevity of the bioreactor.
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PMID:Immobilization of L-asparaginase into a biocompatible poly(ethylene glycol)-albumin hydrogel: evaluation of performance in vivo. 942 58

We attempted to administer PEG-L-asparaginase (PEG-L-A) following hematologic recovery to 38 patients undergoing autologous or allogeneic marrow transplantation for acute lymphoblastic leukemia (ALL). Twenty-four patients (12 of 22 receiving allogeneic and 12 of 16 receiving autologous transplants) received between one and 12 doses of PEG-L-A, including nine who completed the planned 12 doses of therapy. The toxicities encountered were similar to those observed in non-transplanted patients undergoing therapy with PEG-L-A and included allergic reactions, pancreatitis, weight loss, hypoalbuminemia, and low levels of anti-thrombin III. Of the 24 who received the drug, eight remain in remission. Of 12 patients in second remission at the time of transplantation who received PEG-L-A, five of seven who received allogeneic and two of five who received autologous transplants remain in remission, 16+ to 46+ months from transplant. While PEG-L-A could be administered to most of the patients undergoing marrow transplantation for ALL, most patients either relapsed while receiving the drug or developed toxicities which resulted in abbreviated courses. At this time, we cannot recommend PEG-L-A as single agent, post-BMT chemotherapy.
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PMID:Toxicity, pharmacology and feasibility of administration of PEG-L-asparaginase as consolidation therapy in patients undergoing bone marrow transplantation for acute lymphoblastic leukemia. 961 79

Owing to the high efficacy of L-asparaginase in the treatment of acute lymphatic leukaemia the enzyme was introduced into the chemotherapy schedules for remission induction of this disease shortly after results of large-scale clinical trials had become available. Since asparaginase monotherapy was associated with a high response rate but short remission duration, the enzyme is currently employed within the framework of combination chemotherapy schedules which achieve treatment response in about 90% and long-term remissions in the majority of patients. Recently initiated clinical trials have still confirmed the eminent value of asparaginase in the combination chemotherapy of acute lymphatic leukaemia and of some subtypes of non-Hodgkin lymphoma, and its important role as an essential component of multimodal treatment protocols. Despite the unique mechanism of action of this cytotoxic substance which shows relative selectivity with regard to the metabolism of malignant cells, some patients experience toxic effects during asparaginase therapy. Immunological reactions toward the foreign protein include enzyme inactivation without any clinical manifestations as well as anaphylactic shock. Severe functional disorders of organ systems result from the impaired homeostasis of the amino acids asparagine and glutamine. The changes affecting the proteins of the coagulation system have considerable clinical impact as they may induce bleeding as well as thromboembolic events and may be associated with life-threatening complications when the central nervous system is involved. Risk factors predisposing to thromboembolic complications are hereditary resistance against activated protein C and any other hereditary thrombophilia. Other organ systems potentially affected by relevant functional disorders are the central nervous system, the liver, and the pancreas, with patients who have a history of pancreatic disorders carrying an especially high risk of developing pancreatitis. Studies on the mechanisms of action and the occurrence of resistance phenomena have shown that a treatment response may only be expected if the malignant cells are unable to increase their asparagine synthetase activity to an extent providing enough asparagine to the cell; one may thus conclude that the enzyme-induced asparagine depletion of the serum constitutes the decisive cytotoxic mechanism. Independent of the asparagine depletion related cytotoxicity however, there are other mechanisms of clinical relevance like induction of apoptosis. Besides this, further influences on signal transduction cannot be excluded. Only few publications have dealt with the question of minimum trough activities to be ensured before each subsequent asparaginase dose in order to maintain uninterrupted asparagine depletion under treatment, and answers to this problem are not definitive. Clinical studies using enzymes from E. coli strains indicate that a trough activity of 100 U/l will suffice for complete asparagine depletion of the fluid body compartments with the preparations studied. These findings have been transferred to enzymes from other E. coli strains as well as those isolated from Erwinia chrysanthemi and to the PEG-conjugated E. coli asparaginases. It might be desirable to countercheck the results for confirmation or correction. The dosage and administration schedule of the various enzyme preparations required for complete asparagine depletion over a period of time have been insufficiently defined. While pharmacokinetic studies showed clinically relevant differences in biological activity and activity half-lives for enzymes from different biological sources, the findings of recently published clinical trials indicate that the therapeutic efficacy is affected when different asparaginase preparations are given by identical therapy schedules. (ABSTRACT TRUNCATED)
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PMID:Use of L-asparaginase in childhood ALL. 976 45

Asparaginase (ASP) is a standard component of the antileukemia armamentarium. There are currently 3 preparations of asparaginase available: (1) E. coli (ASP, Elspar); (2) the enzyme derived from Erwinia chrysanthemi (ERW, Erwinase); (3) pegaspargase (PEG, Oncaspar), the E. coli enzyme modified by covalent attachment of polyethylene glycol. This report describes the findings of 3 pharmacologic end points: ASP enzyme activity in patients' sera, depletion of asparagine and the development of anti-ASP antibodies. Pharmacokinetics and pharmacodynamic studies in a group of naive children with newly diagnosed ALL demonstrate a significant difference in apparent half-life (1.24 days E. coli vs. 0.65 ERW vs. 5.73 PEG; p < 0.001) and days of asparagine depletion (14-23 E. coli vs. 7-15 ERW vs. 26-34 PEG; p < 0.01) for the 3 different preparations. Data from Pediatric Oncology Group (POG) Protocol #8866 show that high antibody levels correlated with rapid ASP clearance and a significantly lower response rate (NR = 26% vs. CR + PR + 64%). The pharmacologic characteristics of ASP in terms of clearance of enzyme activity and ability to deplete serum asparagine was dependent upon the nature of the enzyme and are significantly altered in patients who develop anti-ASP antibodies regardless of their clinical status. In addition, these data demonstrate that ASP pharmacokinetics are directly related to its anti-leukemic effect. In order to maximize the therapeutic benefits of ASP, the optimal dose and schedule of treatment should be determined based on pharmacologic testing rather than by clinical criteria alone. Future studies will focus on the role of "silent hypersensitivity" as a mechanism of resistance to ASP and strategies to maximize the therapeutic efficacy of ASP as part of ALL therapy.
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PMID:The three asparaginases. Comparative pharmacology and optimal use in childhood leukemia. 1050 Aug 42

Polymer conjugation is of increasing interest in pharmaceutical chemistry for delivering drugs of simple structure or complex compounds such peptides, enzymes and oligonucleotides. For long time drugs, mainly with antitumoral activity, have been coupled to natural or synthetic polymers with the purpose of increasing their blood permanence time, taking advantage of the increased mass that reduces kidney ultrafiltration. However only recently complex constructs were devised that exploit the 'enhanced permeability and retention' (EPR) effect for an efficient tumor targeting, the high molecular weight for adsorption or receptor mediated endocytosis and finally a lysosomotropic targeting, taking advantage of acid labile bonds or cathepsin susceptible polypeptide spacers between polymer and drug. New original, very active conjugates of this type, as those based on poly(hydroxyacrylate) polymers, are already in advanced state of development. Labile oligonucleotides, including antisense drugs, were also successfully coupled to polymers in view of an increased cell penetration and stabilization towards nucleases. However, the most active research activity resides in the field of polypeptides and proteins delivery, mainly for the two following reasons: first of all because a great number of therapeutically interesting compounds are now being produced by genetic engineering in large quantity and, secondly, because these products are difficult to administer to patients for several inherent drawbacks. Proteins are in fact easily digested by many endo- and exo-peptidases present in blood or in other body districts; most of them are immunogenic to some extent and, finally, they are rapidly excreted by kidney ultrafiltration. Covalent polymer conjugation at protein surface was demonstrated to reduce or eliminate these problems, since the bound polymer behaves like a shield hindering the approach of proteolytic enzymes, antibodies, or antigen processing cell. Furthermore, the increase of the molecular weight of the conjugate allows to overcome the kidney elimination threshold. Many successful results were already obtained in peptides and proteins, conjugated mainly to water soluble or amphiphilic polymers like poly(ethylene glycol) (PEG), dextrans, or styrenemaleic acid anhydride. Among the most successful are the conjugates of asparaginase, interleukin-2 or -6 and neocarcinostatin, to remind some antitumor agents, adenosine deaminase employed in a genetic desease treatment, superoxide dismutase as scavenger of toxic radicals, hemoglobin as oxygen carrier and urokinase and streptokinase as proteins with antithrombotic activity. In pharmaceutical chemistry the conjugation with polymers is also of great importance for synthetic applications since many enzymes without loss of catalytic activity become soluble in organic solvents where many drug precursors are. The various and often difficult chemical problems encountered in conjugation of so many different products prompted the development of many synthetic procedures, all characterized by high specificity and mild condition of reaction, now known as 'bioconjugation chemistry'. Bioconjugation developed also the design of new tailor-made polymers with the wanted molecular weight, shape, structure and with the functional groups needed for coupling at the wanted positions in the chain.
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PMID:Bioconjugation in pharmaceutical chemistry. 1051 Aug 47

L-asparaginase is widely used in the treatment of acute lymphoblastic leukemia in children and adults. Use of L-aspa E. Coli as well as Erwinase is not possible in all cases because of the side effects, mainly allergic reactions and disfunction of pancreas. Recently, the new form of the enzyme PEG-L-asparaginase was introduced. Binding L-asparaginase E. coli to polyethylene glycol a decreased its toxicity, extended its plasma half-live, not significantly affecting the efficacy. The aim of the study was to examine the results of PEG-L-asparaginase administration in five children with acute lymphoblastic leukemia, and the symptoms of intolerance to L-aspa E. Coli or Erwinase. There were three children with newly diagnosed ALL and two children with first relapse of ALL, treated according to New York Protocol and BFM-90 Protocol for ALL relapses respectively. PEG-L-asparaginase (Oncaspar) was administered in the dose of 2500 IU/m2. According to the protocol four children received 11 courses of treatment with the full dose of the drug. The number of doses for individual patient varied from one to six. The short-lived nettlerash was observed in one patient during two subsequent infusions of the drug. Hydrocortisone and antihistamine drugs were administered. Treatment with PEG-asparaginase was discontinued in one child, who developed dyspnea, nausea, vomiting and face rash during the third dose of the drug. Oncaspar is the valuable drug, which enabled continuation of treatment according to protocol in four out of five children with bad tolerance to routinely used L-asparaginase preparations.
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PMID:[New possibilities of treatment with PEG-L-asparaginase in patients with acute lymphoblastic leukemia sensitized to l-asparaginase E.coli and erwinase]. 1073 74

Periplasmic Escherichia coli L-asparaginase II with an Ser58Ala mutation in the active-site cavity has been crystallized in a new orthorhombic form (space group P2(1)2(1)2). Crystals of this polymorph suitable for X-ray diffraction have been obtained by vapour diffusion using two sets of conditions: (i) 1% agarose gel using MPD as precipitant (pH 4.8) and (ii) liquid droplets using PEG-MME 550 (pH 9.0). The crystals grown in agarose gel are characterized by unit-cell parameters a = 226.9, b = 128.4, c = 61.9 A and diffract to 2.3 A resolution. The asymmetric unit contains six protein molecules arranged into one pseudo-222-symmetric homotetramer and an active-site competent dimer from which another homotetramer is generated by crystallographic symmetry.
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PMID:Crystallization and preliminary crystallographic studies of a new crystal form of Escherichia coli L--asparaginase II (Ser58Ala mutant). 1073 36

The development of antibodies to asparaginase may attenuate the pharmacologic effect of asparaginase treatment, may be associated with hypersensitivity reactions, and may necessitate switching to a different commercial asparaginase preparation for current or future therapy. Thus, development of an ELISA for measurement of anti-asparaginase antibody levels is important in the clinical setting. An anti-asparaginase antibody reference was established by screening 65 plasma samples from six patients with acute lymphoblastic leukemia (ALL) who had recently developed a hypersensitivity reaction to Escherichia coli or Erwinia chrysanthemi asparaginase therapy. Twenty-one plasma samples were selected for the anti-asparaginase antibody reference pool. Five micrograms per milliliter of commercial E. coli and Erwinia asparaginase and 10 microg/ml of E. coli asparaginase conjugated with polyethylene glycol (PEG asparaginase) were found to be optimal as coating antigen concentrations. Anti-asparaginase antibody concentrations were determined using a commercial polyclonal goat anti-human IgG horseradish peroxidase conjugate. The antibody reference curves were linear in a range of absorbance from 0.1 to 1. 5 O.D. units for dilutions from 1:1600 to 1:51,200. Inter-assay coefficients of variation were 9.04, 14.7 and 13.0%, and intra-assay coefficients of variation were 1.44, 4.43 and 3.28% for antibodies against E. coli, Erwinia, and PEG L-asparaginase, respectively. The cut-off for positivity in plasma was determined as mean+2 S.D. of the optical density values for plasma from untreated healthy volunteers. Measurement of specific IgG by this ELISA allows for the evaluation of plasma anti-asparaginase antibody concentrations in patients receiving one or more of the multiple commercial L-asparaginase preparations.
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PMID:ELISA to evaluate plasma anti-asparaginase IgG concentrations in patients with acute lymphoblastic leukemia. 1082 49

The relapse rate in childhood acute lymphoblastic leukemia (ALL) is approximately 30% but few reinduction regimens have investigated the intensive use of polyethylene glycol Escherichia coli asparaginase (PEG-Asp). Therefore, we assessed the pharmocokinetics and efficacy of PEG-Asp in this setting. Children with B-precursor ALL, in first marrow and/or extramedullary relapse were eligible. Reinduction included doxorubicin on day 1, prednisone for 28 days, vincristine weekly for 4 weeks, and PEG-Asp either weekly or biweekly by randomization. Asparaginase levels and antibody to both E coli asparaginase and PEG-asp were measured weekly just before each PEG-asp dose. Overall, 129 of 144 patients (pts) (90%) achieved a complete remission (CR). There was a highly significant difference in CR rates between weekly (69 of 71; 97%) and biweekly (60 of 73; 82%) PEG-Asp dosing (P =.003). Grade 3 or 4 infectious toxicity was common (50%), but only 4 pts died of sepsis during induction. Other toxicities were infrequent and hypersensitivity was rare (6 of 144; 4%). Low asparaginase levels were associated with high antibody titers to either native (P =.024) or PEG asp (P =.0013). The CR rate was significantly associated with higher levels of asparaginase (P =. 012). Patients with ALL in first relapse receiving weekly PEG-Asp had a higher rate of second remission compared with biweekly dosing. Low levels of asparaginase were associated with high antibody titers. Increased asparaginase levels may correlate with an improved CR rate. The use of intensive PEG-Asp should be explored further in the treatment of ALL. (Blood. 2000;96:1709-1715)
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PMID:Weekly polyethylene glycol conjugated L-asparaginase compared with biweekly dosing produces superior induction remission rates in childhood relapsed acute lymphoblastic leukemia: a Pediatric Oncology Group Study. 1096 68


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