Biomedical subjects

E Schleyer

Publications and source records attributed to E Schleyer.

At least 37 records · Page 2Linked to original sources

PubMed1999

Leukaemic blasts differ from normal bone marrow mononuclear cells and CD34+ haemopoietic stem cells in their metabolism of cytosine arabinoside.

Different metabolites of cytosine arabinoside (AraC) contribute to its cytotoxicity including incorporation of AraCTP into DNA, the incorporation of AraUMP into RNA, inhibition of polymerase alpha and beta (AraCMP/CTP), an impairment of repair mechanisms (AraCTP), alterations of phospholipid metabolism (AraCDP-choline), a direct membrane interaction (AraC), the alteration of signal transduction pathways (AraCDP-choline, AraCTP) and the induction of apoptosis. Since little is known about the potential differences in AraC metabolism between leukaemic blasts and normal haemopoietic progenitor cells, the formation of all known AraC metabolites was determined in bone marrow samples from patients with acute myeloid leukaemia (AML), healthy volunteers and specimens of cellsorted CD34+ haemopoietic stem cells. Highly significant differences were found for phosphorylated AraC metabolites (AraCMP, -CDP, -CTP, AraUMP) between AML and normal mononuclear bone marrow (ng/107 cells respectively 1.30 v 2.66; 2.65 v 7.50; 33.68 v 99.0; 1.18 v 5.70). The highest differences were found for formation of AraCDP-choline (3.75 v 12.86) which might be relevant for the high efficacy of high-dose AraC regimens. In contrast, no differences were found in the deamination product AraU (2.01 v 2.91). Only minute amounts of phosphorylated AraU derivatives were detected, providing an explanation for the lacking contribution of AraU to cytosine arabinoside cytotoxicity. Results in normal CD34+ haemopoietic stem cells did not differ significantly from normal bone marrow mononuclear cells and therefore justify their use as a surrogate in determining AraC-induced haematotoxicity. These data suggest a metabolic basis for the relative selectivity of AraC cytotoxicity for AML blasts and provide a means to determine the role of different metabolites and their related mechanism of action for overall AraC cytotoxicity.

Acute Disease↗

PubMed1998

Oral cytarabine ocfosfate in acute myeloid leukemia and non-Hodgkin's lymphoma--phase I/II studies and pharmacokinetics.

Cytosine arabinoside (AraC) is rapidly inactivated via systemic deamination with half-lives ranging from 1 h (i.v.) to 4 h (s.c.) -- and cannot be applied orally due to its hydrophilic properties. These limitations might be overcome by YNK01 -- a lipophilic prodrug of AraC -- that is resistant to deoxycytidine deaminase and can be applied orally. In the present study the therapeutic activity, side-effects and pharmacokinetics of YNK01 were evaluated in a phase I/II study including patients with relapsed or refractory acute myeloid leukemia (AML) (n=23) or low-grade non-Hodgkin's lymphoma (NHL) (n=20). YNK01 was given by 14 day cycles with escalating doses starting with a daily dose of 50 mg/m2 (equivalent to 20 mg/m2 AraC on a molar basis). The maximum tolerated dose was reached at the 600 mg/m2 dose level with WHO grade 3-4 diarrhoea as the main toxicity. In the 23 patients with AML two complete remissions, four partial remissions and three patients with stable disease were observed. In the 23 patients with AML two complete remissions, four partial remissions and three patients with NHL two cases reached partial remission and six other patients mainained stable disease. Pharmacokinetic evaluations were performed during 34 treatment cycles in 28 patients. The data suggest that YNK01 was absorbed in the distal part of the small intestine and taken up into hepatocytes. After hepatic psi and subsequent beta-oxydation of YNK01 the released AraC (and its deamination product AraU) appeared in the systemic circulation. Time of maximum concentration (h), half-life (h) and area under the curve (ng x h/ml, at the 1200 mg dose level) were as follows (VC variation coefficient) YNK01: 1.0 (0.58), 10.1 (0.43), 12622 (0.65); AraC: 23.2 (0.57), 22.6 (0.36), 3496 (0.76); AraU: 19.2 (0.58) 22.3 (0.33) 15441 (0.66). Of the total dose of YNK01 15.8% was absorbed and metabolized to AraC and AraU, defining the metabolic bioavailability of this prodrug. A linear relationship was observed between YNK01 dose and YNK01 AUC and AraC AUC over the whole dose range tested. AraC was released from hepatocytes over a prolonged period of time resulting in long lasting plasma levels similar to a continuous i.v. infusion. After administration of YNK01 at a dosage of 100-150 mg/m2 plasma levels of AraC were comparable to those achieved after low-dose AraC treatment (20 mg/m2) while at doses of YNK01 of 450-600 mg/m2 concentrations of standard-dose AraC (100 mg/m2) were obtained. We conclude that YNK01 shows considerable activity against relapsed and refractory AML and NHL and that its pharmacokinetic properties offers advantages in comparison to (standard) i.v. or s.c. AraC in clinical practice.

Acute Disease↗

PubMed1997

Activity of thymidine kinase and of polymerase alpha as well as activity and gene expression of deoxycytidine deaminase in leukemic blasts are correlated with clinical response in the setting of granulocyte-macrophage colony-stimulating factor-based priming before and during TAD-9 induction therapy in acute myeloid leukemia.

The present study was undertaken to assess the predictive value of pretherapeutic determinants of ara-C metabolism and proliferative activity of leukemic blasts for early response to antileukemic therapy in the setting of granulocyte-macrophage colony-stimulating factor (GM-CSF)-based priming before and during TAD-9 induction in 36 consecutive patients with de novo acute myeloid leukemia (AML). Ara-C metabolism was assessed by the activities of deoxycytidine kinase (DCK), deoxycytidine deaminase (DCD), DNA polymerase alpha (Poly alpha), and overall polymerase (overall Poly). The fraction of cells in S phase (%S phase) and thymidine kinase (TK) activity were determined as a measure of proliferative activity. Early response to therapy was defined by the percentage of leukemic blasts in the bone marrow 5 to 7 days after completion of TAD-9 with less than 5% signaling an adequate response and greater than 5% indicating an inadequate early reduction, respectively. While neither %S phase, DCK, nor overall Poly activity were predictive for early response, TK and Poly alpha activities were significantly higher for cases with adequate blast cell clearance. The respective median values were for TK 3.8 versus 1.85 pmol/min/mg protein (P = .012), and for Poly alpha 1.9 versus 0.69 pmol/min/mg protein (P = .014). An inverse relation was detected for DCD activity which was significantly lower in responding patients with a median of 0.33 nmol/min/mg protein (range, 0.0 to 29.5) as compared to a median of 5.1 nmol/min/mg protein (range, 0.11 to 8.45) in early nonresponders, (P = .009). Taking the respective median values as arbitrary cut-points for high or low enzyme activities, responders and nonresponders could be discriminated prospectively. Hence, 14 of 16 cases (88%) with DCD activities below the median of 1.56 nmol/min/mg protein responded as compared to only 3 of 14 (22%) patients with higher DCD activities (P = .0004). From the 15 patients with TK activity above the overall median of 3.2 pmol/min/mg protein, 11 cases (73%) achieved an adequate blast cell clearance while only 6 of 17 cases (35%) with lower values responded (P = .035). Similarly, 12 of 15 patients (80%) with high Poly alpha levels (>1.22 pmol/min/mg protein) responded to induction therapy as compared to only 5 of 14 patients (36%) with lower enzyme activities (P = .02). By logistic regression analysis of enzyme activities, DCD activity was found to be the most sensitive parameter to predict an adequate blast cell clearance (P = .032). Activities of DCD and TK were not only associated with initial response but were also found predictive for remission duration. Hence, from 11 patients with low TK levels 8 (73%) relapsed within 1 year, whereas only 2 of 11 (18%) patients with high TK activity experienced a recurrence of their disease (P = .015). Six of 9 (66%) patients with higher than median DCD levels relapsed within 1 year, whereas 10 of 14 patients (71%) with lower DCD levels had a longer remission duration (P = .085). Analysis of DCD gene expression at the mRNA level by a semi-quantitative reverse transcriptase-polymerase chain reaction method showed that a high transcription rate of the DCD gene was associated with high enzyme activities and vice versa. Hence, the observed intraindividual differences in DCD activity are a reflection of differences in gene activity and transcription rate rather than of variants in translation. Although further analyses are needed to elucidate the molecular mechanisms that determine the variation of enzyme activities in individual patients, the present study strongly suggests that pretherapeutic determination of TK and Poly alpha as well as of DCD allows to predict response to TAD-9 + GM-CSF induction therapy and may provide the means for the development of a risk adapted treatment strategy.

Acute Disease↗

PubMed1997

High antileukemic activity of sequential high dose cytosine arabinoside and mitoxantrone in patients with refractory acute leukemias. Results of a clinical phase II study.

BACKGROUND: The current study was initiated to assess the efficacy and side effects of a timed sequential application of high dose cytosine arabinoside (AraC) in combination with mitoxantrone (S-HAM) in patients with refractory acute myeloid leukemia (AML) or acute lymphoblastic leukemia (ALL). METHODS: Patients with refractory AML or ALL were eligible for S-HAM salvage therapy, which was comprised of AraC, 1 g/m2 or 3 g/m2 every 12 hours, on Days 1, 2, 8, and 9 and mitoxantrone, 10 mg/m2/day, given on Days 3, 4, 10, and 11. RESULTS: Of 22 fully evaluable patients, 14 patients (64%) achieved a complete remission whereas 5 patients (23%) succumbed to early death and 3 patients (14%) did not respond. Blood counts recovered at a median of 33.5 days after the start of treatment and complete remission was achieved after a median of 38 days. The median duration of complete remission was 4 months (range, 1-14 months) whereas overall survival time lasted for a median of 4.5 months (range, 1-30+ months). Treatment-associated toxicity was comprised predominantly of infection and diarrhea that reached World Health Organization Grades 3 and 4 in 64% and 32% of patients, respectively. Complementary pharmacokinetic evaluations of plasma AraC and AraU levels revealed no impact of initial AraC administration on the pharmacokinetics of subsequent AraC administrations and failed to demonstrate any evidence of self-potentiation. CONCLUSIONS: The clinical data show the S-HAM regimen to be a promising approach for the treatment of patients with advanced acute leukemias. However, further evaluation at earlier stages of treatment is needed.

Acute Disease↗

PubMed1997

Frequency and causes of refractoriness in multiply transfused patients.

The use of leukocyte-depleted blood components has become the standard therapy for multiply transfused patients during the past few years, as a measure to reduce the frequency of alloimmunization and refractoriness. We assessed frequency and causes of refractoriness, defined as a repeated 24-h post-transfusion platelet count below 20,000/microliters, in 145 consecutive patients who received three or more single-donor platelet concentrates during a 1-year period. Flow-cytometric detection of anti-platelet antibodies and a glycoprotein-specific ELISA were applied for the diagnosis of alloimmunization. Forty patients (27.6%) had at least one episode of refractoriness. In 25 of these 40 patients (62.5%), nonimmune factors (fever, sepsis, coagulopathy, splenomegaly) alone were the cause. In 15 refractory patients alloantibodies were detected. In seven patients (17.5%), alloimmunization alone caused an inadequate transfusion response, while in eight refractory patients (20.0%) alloimmunization and fever or sepsis were present. HLA antibodies were detected in 17 patients (11.7%); three patients (2%) had platelet-specific antibodies in addition to HLA antibodies; in two patients panreactive platelet antibodies were detectable. All 17 patients had a history of previous transfusions or pregnancy. We did not observe primary immunization in patients transfused exclusively with filtered (leukodepleted) blood products. Our data suggest that alloimmunization in patients with a negative risk history can be prevented by the exclusive use of leukodepleted blood components.

Antigens, Human Platelet↗

PubMed1997

Differential effect of GM-CSF pretreatment on intracellular Ara-C metabolism in normal bone marrow mononuclear cells vs acute myeloid leukemia (AML) blasts.

The effect of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) on the intracellular metabolism of cytosine arabinoside (Ara-C) was comparatively analyzed in normal bone marrow mononuclear cells (NBMMC) from eight healthy volunteers and in leukemic blasts from 50 patients with acute myeloid leukemia (AML). Pretreatment with GM-CSF (100 U/ml) for 48 h resulted in a significant enhancement of DNA synthesis in both cell types: 21 of 35 AML specimens were found to be responsive to GM-CSF as defined by an increase of 3H-TdR incorporation into the DNA > 1.5-fold while NBMMC from normal donors were responsive in all cases. In GM-CSF responsive AML blasts, overall DNA polymerase and DNA polymerase alpha activity increased from a median of 84.4 to 96.1 and from 3.45 to 5.2 pmol/min x mg as compared to a median of 96.7 to 189.9 and 1.2 to 2.2 pmol/min x mg in NBMMC (P < 0.05). Median Ara-C-mediated inhibition of DNA synthesis was significantly more effective in AML blasts as compared to NBMMC (76.5 vs 55.0% at 0.05 microM and 99.0 vs 96.0% at 5.0 microM Ara-C, P < 0.01) but was not influenced by GM-CSF pretreatment. Similarly, intracellular Ara-CTP levels were higher in AML blasts as compared to NBMMC (median of 46.9 vs 18.7 at 1 microM, 167.8 vs 48.0 at 10 microM and 337.5 vs 59.5 ng/10(7) cells at 100 microM extracellular Ara-C, P < 0.01) but showed no enhancement in the presence of GM-CSF. Median deoxycytidine (DCK) and thymidine kinase (TK) activity were only slightly increased in AML blasts after GM-CSF priming. In contrast, NBMMC revealed a significant increase in TK activity after GM-CSF pretreatment (from a median of 1.9 to 3.6 pmol/min x mg, P = 0.039). At low; intermediate and high extracellular Ara-C concentrations GM-CSF pretreatment resulted in a significant enhancement of the 3H-Ara-C incorporation into the DNA in both GM-CSF responsive AML blasts and NBMMC (median of 1.3 to 2.1- and 1.4 to 1.6-fold, P < 0.05). GM-CSF non-responsive AML blasts showed no change in 3H-Ara-C incorporation into the DNA in response to GM-CSF at low Ara-C concentrations but significant increases at intermediate and high extracellular Ara-C concentrations (median increases of 1.63-fold at 1.06 microM with P = 0.01 and 1.37-fold at 10 microM extracellular Ara-C with P = 0.0+005). NBMMC revealed significantly lower GM-CSF-induced increases of the 3H-Ara-C incorporation into the DNA as compared to the effect of GM-CSF priming on DNA synthesis (median increases of 1.4 to 1.7-fold vs 2.6-fold, P < 0.05). These data reveal a different effect of GM-CSF priming on the metabolism of Ara-C in normal vs leukemic cells which may cause a preferential increase in the antileukemic cytotoxicity of Ara-C in the presence of GM-CSF.

Adult↗

PubMed1997

The influence of 1-beta-D-arabinofuranosylcytosine on the metabolism of phosphatidylcholine in human leukemic HL 60 and Raji cells.

We report that high-dose 1-beta-D-arabinofuranosylcytosine (Ara-C) treatment leads to substantial changes of membrane lipid composition in human leukemic cell lines. HL 60 cells are at least 10- to 20-fold more sensitive to Ara-C than Raji cells. After 4 h incubation with 50 microM Ara-C, both cells show deviations in their phosphatidylcholine (PC) and triglyceride (TG) contents, starting as early as 8 h after treatment. After 24 h, the Ara-C-induced changes in lipid metabolism are accompanied by a severe loss of viability in HL 60 cells but not in Raji cells. At this time point the HL 60 cells show a 20% depletion of PC with a concomitant increase in TG of 25%, whereas in Raji cells both PC and TG are increased 20 and 22%, respectively. The addition of lysophosphatidylcholine (lysoPC) antagonizes Ara-C-induced cell death in various leukemic cell lines and primary AML blasts from patients. Since lysoPC is a direct precursor for PC and increases the PC content of the membrane, we assume that the loss of PC in the sensitive cell line HL 60 and in other cells plays a role in Ara-C-induced toxicity. Further evidence for this mechanism is presented by the observation that hexadecylphosphocholine, an inhibitor of PC synthesis shows synergistic antiproliferative effects with Ara-C. We conclude that the rapid cell lysis described during high-dose Ara-C treatment seems to be mediated by reduction of cell membrane PC content.

Antimetabolites, Antineoplastic↗

PubMed1997

Oral idarubicin pharmacokinetics--correlation of trough level with idarubicin area under curve.

Idarubicin is the first anthracycline that can be successfully administered via the oral route and thus may facilitate antineoplastic chemotherapy in an improved quality of life. These perspectives are somewhat hampered by the large variation in bioavailability between individual patients and the obvious requirement to monitor plasma concentration and area-under-the-curve values (AUC) for an appropriate adjustment of idarubicin dose. In this study we describe the pharmacokinetics of idarubicin and its main metabolite idarubicinol in 12 patients after oral application of 20 mg/m2 idarubicin on 3 consecutive days and demonstrate that the 24 h trough levels show a high correlation with AUC and may thus allow a rapid and easy determination of individual drug concentrations and an appropriate dose adjustment. The average terminal half-life was 30.5 h for idarubicin and 66.9 h for idarubicinol. The AUC for idarubicin and its main metabolite idarubicinol revealed a substantial interpatient variation with AUC values ranging from 25.7 to 114 ng x h/ml (average 58.1 ng x h/ml) for idarubicin and from 109.4-445.2 ng x h/ml (average 287.3 ng x h/ml) for idarubicinol. However, the ratio of idarubicin/idarubicinol differed only two folds from 1:3.7 to 1:7.7 with an average of 1:5.1. Both idarubicin and idarubicinol concentrations were highly reproducible, however, upon measurements after repeated applications within individual patients. Moreover, idarubicinol and idarubicin AUCs showed a good correlation with r = 0.78, indicating that the interindividual variation of idarubicin AUC reflects differences in absorption rather than in metabolism. In order to describe the interindividual bioavailability of idarubicin - represented by the respective AUC - measurement of a single data point with a high correlation with the AUC would be ideal. Our study demonstrates that the 24 h trough level shows such an excellent correlation (r = 0.96) with AUC, making it the perfect candidate for fast estimates of the individual bioavailability in a given patient. On this basis, the longitudinal measurement of the 24 h trough level may allow assessment of the impact of interindividual variations in AUC on clinical outcome and toxicity.

Administration, Oral↗

PubMed1996

Detection and determination of the major metabolites of [3H]cytosine arabinoside by high-performance liquid chromatography.

An ion-pair high-performance liquid chromatographic assay involving solid-phase scintillation detection was established for the rapid identification and determination of all major metabolites of tritium-labelled cytosine arabinoside (Ara-C) in an in vitro system. In a single run of 50 min, Ara-C, Ara-CMP, Ara-CDP-choline, Ara-CDP, Ara-U, Ara-UMP, Ara-CTP, Ara-UDP and Ara-UTP can be measured. The method is fast, sensitive, with limits of detection ranging from 40 to 200 pg (absolute), and highly reproducible.

Calibration↗

PubMed1996

Response to cytarabine ocfosfate (YNK01) in a patient with chronic lymphocytic leukemia refractory to treatment with chlorambucil/prednisone, fludarabine, and prednimustine/mitoxantrone.

Cytarabine ocfosfate (YNK01) is a novel orally applicable prodrug of cytosine arabinoside. Recent pharmacokinetic studies have revealed a prolonged release of the cytotoxic agent cytosine arabinoside (araC) from hepatocytes into the systemic circulation, resulting in a half-life of approximately 24 h for araC. The specific pharmacokinetic characteristics of cytarabine ocfosfate lead to a prolonged exposure of leukemic cells to this antineoplasstic agent during the 14-day cycle. the oral applicability during outpatient treatment and the sustained antineoplastic activity of araC against slowly proliferating leukemic B-cells suggest that cytarabine ocfosfate might be a useful drug in the treatment of chronic lymphocytic leukemia. Four years after diagnosis of B-CLL, a 50-year-old patient was started on cytarabine ocfosfate. Sequentially, the patient's disease had proved refractory to treatment with chlorambucil/prednisone (31 months), fludarabine (5 months), and prednimustine/mitoxantrone (3 months). These established regimens were discontinued because of increasing lymphocytosis, significant thrombocytopenia, and progressive B-symptoms. Following three cycles of cytarabine ocfosfate B-symptoms resolved, lymphadenopathy disappeared, and thrombocytopenia was significantly reduced. The patient has been free of these symptoms on a dosage of 1500 mg cytarabine ocfosfate/day (cycle of 14 days with intervals of 14-21 days) for 24 months and remains in an ongoing partial remission.

Administration, Oral↗

PubMed1996

Optimizing therapy for acute myeloid leukemia based on differences in intracellular metabolism of cytosine arabinoside between leukemic blasts and normal mononuclear blood cells.

The increasing insights into the pharmacokinetics and the metabolism of arabinoside C (AraC) have improved the rationale for its application in leukemia therapy and have led to a pharmacologically directed design of antileukemic treatment. The current study aims at adding to this approach by detecting differences in the intracellular metabolism of AraC 5'-triphosphate (AraCTP) between leukemic and normal mononuclear blood cells. Measurements of intracellular AraCTP levels were complemented by determinations of plasma AraC and arabinoside uridine (AraU) concentrations and were performed in 26 patients with acute myeloid leukemia (AML) who were undergoing combination therapy, including high-dose (1.0 or 3.0 g/m2 x 2/day) AraC. Plasma AraC concentrations showed a linear relationship to the applied AraC dose but did not correlate with intracellular AraCTP levels. Substantial differences in AraCTP retention times were revealed, during 3-h infusions of either 1.0 or 3.0 g/m2 AraC in leukemic blasts from 10 patients with t1/2 values of 1.60-7.63 h (median, 2.42 h). In addition, AraCTP levels declined in only one patient by > 10% within the first hour after the end of therapy and remained constant or even increased up to 1.5-fold during a posttreatment period of 1-2.5 h in the other nine cases. In contrast, AraCTP retention times were relatively uniform in normal mononuclear blood cells from 11 patients, with t1/2 values of 3.34-5.29 h (median, 3.85 h). More importantly, AraCTP levels dropped by > 10% within the first hour after the end of the high-dose AraC infusion in eight of 11 cases. A posttherapeutic increase of > 10% was not observed in any patient. These differences in AraCTP pharmacokinetics between leukemic and normal blood cells provided the basis for a modified timing of AraC administration with the aim of selectively maintaining cytotoxic AraCTP levels in leukemic blasts while allowing an intermittent drop of AraCTP levels in normal cells. This modification may result in higher antileukemic activity without increasing the damaging effect on normal cells and may, thus, improve the therapeutic index for AraC.

Acute Disease↗

PubMed1996

Oral idarubicin pharmacokinetics--correlation of trough level with idarubicin area under curve.

Idarubicin is the first anthracycline that can be successfully administered via the oral route and thus may facilitate antineoplastic chemotherapy at an improved quality of life. These perspectives are somewhat hampered by the large variation in bioavailability between individual patients and the obvious requirement to monitor plasma concentration and area-under the curve values (AUC) for an appropriate adjustment of idarubicin dose. In this study we describe the pharmacokinetics of idarubicin and its main metabolite idarubicin in 12 patients after oral application of 20 mg/m2 idarubicin on 3 consecutive days and demonstrate that the 24-h trough levels shows high correlation with AUC and may thus allow a rapid and easy determination of individual drug concentrations and an appropriate dose adjustment. The average terminal half-life was 30.5h for idarubicin and 66.9 h for idarubicinol. The AUC for idarubicin and its main metabolite idarubicinol revealed a substantial interpatient variation with AUC values ranging from 25.7 to 114 ng x h/ml (average 58.1 ng x h/ml) for idarubicin and from 109.4 - 445.2 ng x h/ml (average 287.3 ng x h/ml) for idarubicinol. However, the ratio of idarubicin/idarubicinol differed only two-fold from 1:3.7 to 1:7.7 with an average of 1:5.1. Both idarubicin and idarubicinol concentrations were highly reproducible, however, upon measurements after repeated applications within individual patients. Moreover, idarubicinol and idarubicin AUCs showed a good correlation with r=0.78, indicating that the interindividual variations of idarubicin AUC reflects differences in absorptions rather than metabolism. In order to describe the interindividual bioavailability of idarubicin - represented by AUC - measurement of a single data point with a high correlation with the AUC would be ideal. Our study demonstrates that the 24-h trough level shows such an excellent correlation (r=0.96) with AUC, making it the perfect candidate for fast estimates of the individual bioavailability in a given patient. On this basis, the longitudinal measurement of the 24-h trough level may allow the assessment of the impact of interindividual variations in AUC of clinical outcome and toxicity.

Acute Disease↗

PubMed1995

Highly sensitive coupled-column high-performance liquid chromatographic method for the separation and quantitation of the diastereomers of leucovorin and 5-methyltetrahydrofolate in serum and urine.

A column-switching chiral HPLC assay was developed that allows the separation and quantitation of the diastereomers of leucovorin (LV, 5-formyltetrahydrofolic acid) and its metabolite 5-methyltetrahydrofolate (METHF) in serum and urine by means of fluorescence detection. The analysis procedure consists of an on-line concentration of the folates in the HPLC system which is followed by the elution and separation of folates on an achiral 3-microns Microbore C18 column in (6R,S)-LV and (6R,S)-METHF. (6R,S)-LV and (6R,S)-METHF are subsequently transferred on-line onto a chiral 7-microns bovine serum albumin column through a Rheodyne valve system and are separated into their diastereometers. Time of analysis is 70 min. Detection limit is 5 ng/ml for each diastereometer. The within-day variation ranges between 3.2 and 15.8% in relation to the measured concentration. Between-day variation is 4.4-12.1% for a concentration of 100 ng/ml for each diastereometer. (6R,S)-LV and (6S)-LV pharmacokinetics were assessed by analyzing serum and urine samples of four-healthy volunteers.

Antineoplastic Agents↗

PubMed1995

Highly sensitive high-performance liquid chromatographic assay for 1-beta-D-arabinofuranosylcytosine-5'-stearyl phosphate (cytarabine-ocfosfate).

An ion-pair HPLC method for the determination of 1-beta-D-arabinofuranosylcytosine-5'-stearyl phosphate (cytarabine-ocfosfate I) was developed, using a phenyl-bonded column under reversed-phase conditions with a mobile phase of acetonitrile-buffered water (pH 6.8) (50:50) for isocratic elution. A reproducible sample clean-up was achieved by solid-phase extraction. In order to reach the low limit of detection of 2 ng/ml, an enrichment switching system was used. The present validation leads to a limit of quantification of 5 ng/ml with a coefficient of variation (C.V.) of 10%. The total time of measurement was shortened by a back-flush procedure to restore the conditions after each run. UV detection at 275 nm was applied. The recoveries for plasma samples ranged from 56.4 to 64.1%, regardless of drug concentrations. The intra-assay C.V. was about 4% (40 measurements at four different concentrations). The inter-assay recovery (ten measurements over ten days) at a plasma concentration of 50 ng/ml was 57% with a C.V. of 8.25%. Based on this HPLC method, the pharmacokinetics of I were measured during a clinical phase I/II study.

Antineoplastic Agents↗

PubMed1995

[Pharmacokinetics of buprenorphine in subcutaneous administration].

UNLABELLED: In this study investigations were carried out on pharmacokinetics of buprenorphine after it had been applied subcutaneously. Up to now no pharmacokinetic data exist using this kind of application with relevant clinical doses. METHODS: With 6 patients, who had major general surgery under balanced anaesthesia, buprenorphine was given in a single subcutaneous injection of 5 micrograms/kg bodyweight for postoperative pain relief. Over a period of 48 hours the level of buprenorphine in blood plasma was measured. The analyses were done with a modified HPLC method with electrochemical detection which is able to achieve measurements as low as 40 pg/ml of plasma. The pharmacokinetic values were calculated with the "Topfit" pharmacokinetic computerised programme. RESULTS: Whereas the period of time (tmax) to reach peak plasma concentration (Cmax) after subcutaneous application is much longer than with other parenteral application forms, the plasma concentration levels tend to be similar after 20 minutes. The most important finding of this study is that the mean terminal half-life (t1/2c) of buprenorphine is 23 hours. However, the presently available data for other application forms in respect to terminal half-life did not produce the same results. This can be explained by the use of insufficiently sensitive analytical procedures and by too short periods of observation of 3 and 13 hours respectively, leading to values of t1/2c of 2 and 5 hours. CONCLUSIONS: These results indicate that the terminal half-life (t1/2c) of buprenorphine is much longer than had been supposed. It confirms clinical findings of sufficient pain relief by showing measurable uptake of buprenorphine after subcutaneous application.

Aged↗

PubMed1995

Blast cell proliferative activity and sensitivity to GM-CSF in vitro are associated with early response to TAD-9 induction therapy in acute myeloid leukemia.

The current study was undertaken to determine the relevance of leukemic blast cell proliferative activity, cellular parameters of Ara-C metabolism and the in vitro sensitivity to GM-CSF in association with the clinical response to TAD-9 induction therapy in 66 patients with de novo acute myeloid leukemia (AML). Proliferative activity was assessed by 3H-thymidine (3H-TdR) incorporation and thymidine kinase (TK) activity, parameters of Ara-C metabolism comprised the activities of deoxycytidine kinase (DCK) and DNA polymerase alpha (poly alpha) as well as Ara-CTP concentrations and 3H-Ara-C uptake into DNA. GM-CSF sensitivity was determined by in vitro incubation of blasts for 48 h with or without GM-CSF (100 U/ml) followed by an additional 4 h concurrent exposure to GM-CSF and 3H-TdR (0.5 microCi/ml). The following results were obtained as expressed by median values and ranges: 3H-TdR incorporation: 1.07 pmol/10(5) cells (0.0-10.1), TK: 7.3 pmol/min/mg protein (1.3-56.0), DCK: 9.3 pmol/min/mg protein (0.77-47.1), poly alpha: 1.7 pmol/min/mg protein (0.00-28.9), Ara-CTP: 53.3 ng/10(7) cells (13.3-211.0), 3H-Ara-C uptake: 0.06 pmol/10(5) cells (0.0-0.57). 3H-Ara-C uptake was correlated with 3H-TdR incorporation (r = 0.74) and with the (S-phase dependent) activities of TK (r = 0.73) and poly alpha (r = 0.71, but not with DCK activity or intracellular Ara-CTP content. Blast cells of 37 from 55 analyzed patients were found to be sensitive to GM-CSF stimulation as defined by an increase in 3H-TdR incorporation > or = 1.5-fold over control values after the 48 h GM-CSF exposure. In vitro data were related with clinical response to TAD-9 induction therapy in 43 patients with newly diagnosed AML, taking the blast cell reduction at day 10 or 16 to < 5% or > or = 5% residual blasts as early parameter for adequate or inadequate response, respectively. While neither 3H-Ara-C uptake, nor intracellular Ara-CTP concentration, TK nor DCK activity were predictive for response, a high 3H-TdR incorporation and a high poly alpha activity were associated with adequate blast cell reduction. Median values of 3H-TdR incorporation were 2.26 pmol/10(5) cells for patients with adequate blast cell clearance and 0.80 pmol/10(5) cells for patients with inadequate blast cell clearance (P = 0.11), the respective values for poly alpha were 3.22 pmol/min/mg protein for responders and 1.1 pmol/min/mg protein for non-responders (P = 0.0085).(ABSTRACT TRUNCATED AT 400 WORDS)

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1995

Pharmacokinetics of Ara-CMP-Stearate (YNK01): phase I study of the oral Ara-C derivative.

Ara-CMP-Stearate (1-beta-D-arabinofuranosylcytosine-5'-stearylphosphate, YNK 01, Fosteabine) is the orally applicable prodrug of cytosine-arabinoside (Ara-C). During a phase I study in patients with advanced low-grade non-Hodgkin lymphomas or acute myeloid leukemia, the pharmacokinetic parameters of Ara-CMP-Stearate (kindly provided by ASTA Medica, Frankfurt, Germany) were determined by HPLC analysis. Seventy-two hours after a first starting dose which served for the determination of baseline pharmacokinetic parameters, Ara-CMP-Stearate was administered over 14 days by daily oral application. Ara-CMP-Stearate was started at a dose of 100 mg/day and was escalated in subsequent patients to 200 mg/day and 300 mg/day. Plasma and urine concentrations of Ara-CMP-Stearate, Ara-C and Ara-U were measured during the initial treatment phase and within 72 h after the end of the 14-day treatment cycle. So far six patients have been treated with 100 mg/day, three with 200 mg/day and another six with 300 mg/day. One patient was treated consecutively with 100 mg, 300 mg and 600 mg. Fitting the results of the plasma concentration measurements of Ara-CMP-Stearate to a one-compartment model, the following pharmacokinetic parameters were obtained (average and variation coefficient VC). Ara-CMP-Stearate dose-independent parameters: lag time = 1.04 h (0.57); tmax = 5.72 h (0.30); t1/2 = 9.4 h (0.36). Dose-dependent parameters: at 100 mg: AUC = 1099 ng/h/ml (0.31); concentration(max) = 53.8 ng/ml (0.28); at 200 mg: AUC = 2753 ng/h/ml (0.32); concentration(max) = 154.8 ng/ml (0.46); at 300 mg: AUC = 2940 ng/h/ml (0.66); concentration(max) = 160.0 ng/ml (0.59). The long lag time and late tmax can be explained by resorption in the distal part of the small intestine. No Ara-CMP-Stearate was detected in urine samples (limit of detection = 500 pg/ml). Pharmacokinetic parameters of Ara-C following Ara-CMP-Stearate application showed the following characteristics: t1/2 = 24.3 h (0.39); AUC (100 mg) = 262 ng/h/ml (0.93); AUC (200 mg) = 502 ng/h/ml (0.87); AUC (300 mg) = 898 ng/h/ml (1.07). Since Ara-CMP-Stearate causes intravascular hemolysis after intravenous administration, it was not possible to determine its bioavailability by comparing the AUC after oral and i.v. application. Instead, the renal elimination of Ara-U, as the main metabolite of Ara-C was measured during the first 72-h period and after the last application.(ABSTRACT TRUNCATED AT 400 WORDS)

Administration, Oral↗

PubMed1994

New perspectives in the treatment of acute myeloid leukemia by hematopoietic growth factors.

The application of hematopoietic growth factors in the treatment on acute myeloid leukemia (AML) may principally aim at shortening the period of treatment associated neutropenia and reducing the rate of infectious complications by their post-therapeutic administration but may also be used to increase the sensitivity of leukemic blasts to antileukemic therapy by pretherapeutic growth stimulation. Both aspects were addressed in subsequent clinical phase II studies and preclinical investigations. In a first clinical trial, 36 patients with high-risk AML received granulocyte-macrophage colony-stimulating factor (GM-CSF) after successful cytoreductive chemotherapy and experienced a shortening of the period of post-therapeutic neutropenia by 6 to 9 days, leading to a significant reduction of treatment-associated deaths from 39% to 14%. In preclinical studies an enhancement of the cytotoxicity of cytosine arabinoside (AraC) on leukemic blasts could be shown by pretreatment with GM-CSF or IL-3. Investigations on the impact of hematopoietic growth factors on the intracellular metabolism of AraC indicated that this effect was primarily mediated by an increase in the activity of DNA-polymerase-alpha. The evaluation of different doses of AraC showed the most marked increase after the combination of GM-CSF with conventional rather than high doses of AraC. Based on these preclinical experiments, a prospective randomized trial was subsequently initiated investigating the effect of GM-CSF before and during induction, consolidation, and the first two cycles of maintenance chemotherapy in newly diagnosed AML. This ongoing trial has enrolled 67 patients at the current time. An early interim analysis showed no differences in remission rates but a tendency toward a longer remission duration in patients receiving GM-CSF. These data indicate that hematopoietic growth factors like GM-CSF in particular may provide a new perspective in the treatment of acute myeloid leukemia with the possibility of reducing treatment associated mortality and perhaps of increasing the efficacy of antileukemic treatment.

Acute Disease↗