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Biomedical subjects

R M Mader

Publications and source records attributed to R M Mader.

At least 37 records · Page 2Linked to original sources
PubMed1998

Resistance to 5-fluorouracil.

1. Primary and secondary resistance to the widely used antimetabolite 5-fluorouracil (5-FU) are common phenomena in cancer chemotherapy. Because 5-FU still remains the agent of choice in the treatment of, for example, colorectal cancer, circumvention of resistance is of vital importance. 2. Resistance to fluoropyrimidines is a multifactorial event, which includes transport mechanisms, metabolism, molecular mechanisms, protection from apoptosis, and resistance via cell cycle kinetics. To date, the prediction of primary resistance to 5-FU in the clinic is limited to few studies focusing mainly on the key enzyme thymidylate synthase. To gain a deeper insight into the key events responsible for 5-FU resistance in vivo, the evaluation of additional parameters such as other (fluoro)pyrimidine converting enzymes, the mutational status of regulators of apoptosis, and tumour angiogenesis is currently under investigation. 3. Most studies on the circumvention of fluoropyrimidine resistance refer to preclinical investigations and were rarely confirmed in clinical trials. Although our understanding of resistance to 5-FU leaves many open questions, the fundamental insights accomplished during the last years provide a rational understanding to exceed the bounds of the actual therapeutic schedules.

Animals↗
PubMed1997

Transcription and activity of 5-fluorouracil converting enzymes in fluoropyrimidine resistance in colon cancer in vitro.

Cellular resistance to 5-fluorouracil (5-FU) is not completely understood. Since 5-FU shares the pyrimidine pathway with the physiological pyrimidines, we investigated the relationship between fluoropyrimidine metabolism, nucleic acid uptake and cytotoxicity of 5-FU in eight colon tumour cell lines including 5-FU-resistant subclones. The cytotoxicity of 5-FU was increased up to 423-fold when the anabolites 5-fluorouridine (FUrd), 5-fluorodeoxyuridine (FdUrd), and 5-fluorodeoxyuridine monophosphate (FdUMP) were compared with the parent drug in vitro. The enzymes uridine phosphorylase and thymidine phosphorylase were predictive for the cytotoxicity of 5-FU in 5/7 cell lines. Inhibition of uridine phosphorylase and thymidine phosphorylase by antisense strategies effectively antagonised 5-FU, abolishing 84% and 79% of its toxicity. The importance of thymidine phosphorylase was supported by a highly restricted enzyme activity in 5-FU-resistant cells. In 5-FU naive cells, a stimulating effect of 5-FU on thymidylate synthase mRNA and ribonucleotide reductase mRNA expression was observed. In these cells, antisense oligonucleotides to ribonucleotide reductase significantly reduced cell growth. Downregulation of ribonucleotide reductase mRNA in 5-FU-resistant subclones suggests different mechanisms in primary and secondary resistance to 5-FU. Most of the intracellular 5-FU was selectively incorporated into RNA (range: 45-91%) and generally spared DNA (range: 0.2-11%). In synthesising our data, we conclude that drug resistance could be overwhelmed through bypassing limiting steps in the activation of 5-FU. In the majority of colonic tumours, the activity of uridine phosphorylase and thymidine phosphorylase may have prognostic relevance for the cytotoxicity of 5-FU in vitro.

Antimetabolites, Antineoplastic↗
PubMed1997

5-fluorouracil kinetics in the interstitial tumor space: clinical response in breast cancer patients.

Several anticancer drugs fail to exhibit sufficient activity against solid tumors in vivo despite effective inhibition of tumor cell growth in vitro. This may be due to impaired drug transfer from plasma into solid tumors. The present study, therefore, aimed at measuring interstitial tumor 5-fluorouracil (5-FU) pharmacokinetics and 5-FU transfer rates from plasma into the tumor interstitium in breast cancer patients. Microdialysis probes were inserted into the primary tumor and the periumbilical s.c. adipose layer of 10 breast cancer patients (8 females and 2 males) scheduled to receive neoadjuvant chemotherapy due to locally advanced breast cancer. Thereafter, patients received 5-FU (600 mg/m2, i.v). 5-FU kinetics were followed in plasma and tumor and s.c. interstitial fluid. Mean interstitial 5-FU load, expressed as area under curve (AUC), in breast tumors was 61 +/- 11% (means +/- SE) of the mean plasma 5-FU load. 5-FU displayed similar kinetics in the interstitial space of s.c. adipose tissue and tumor tissue. A high interstitial tumor AUC was associated with increased tumor response. There was no association with tumor response for s.c. or plasma AUC of 5-FU. Measurement of interstitial drug concentrations in breast tumors by in vivo microdialysis may predict response to chemotherapy. This information may explain drug resistance in some patients and help to optimize dosing and administration schedules. In the future, selection of novel cytotoxic compounds with favorable tumor penetration characteristics may become possible.

Adipose Tissue↗
PubMed1997

The in vitro effects of interleukin-12 upon tumor-infiltrating lymphocytes derived from renal cell carcinoma.

Clinical trials utilising interleukin (IL)-2 with tumor-infiltrating lymphocytes (TIL) have demonstrated efficacy in the treatment of metastatic renal cell carcinoma (RCC). Several cytokines, as well as growth factors have demonstrated modulatory effects upon the biological properties of TIL from RCC, suggesting a potentially important role for cytokines other than IL-2 in the development of active and tumor-specific TIL. IL-12 was recently characterized as a natural-killer-cell-stimulatory factor or cytotoxic-T-cell-maturation factor. These properties of IL-12 prompted us to investigate the impact of this cytokine upon the activation of TIL from human RCC. In an attempt to enhance the in vitro growth and activity of renal TIL, we have grown eight renal TIL cultures in varying concentrations of IL-2 (8, 40, 80, 400 U/ml) and IL-12 (200 U/ml). In addition, IL-12 (200 U/ml) was added to TIL cultures pre-activated with IL-2 (400 U/ml). Growth, cell expansion, and the ability of TIL to release certain cytokines upon tumor stimulation were determined. Proliferation assays, phenotypic analysis, and cytotoxicity assays were performed at an early and a late culture stage. IL-12, alone and when added to suboptimal concentrations of IL-2, failed to induce TIL growth. While the addition of IL-12 to optimal doses of IL-2 suppressed TIL culture expansion, sequential culture exposure first to IL-2 and then to IL-2+IL-12 increased the number of cells expressing CD3+/CD56+ and these cultures demonstrated enhanced in vitro lysis of autologous tumor. IL-12 clearly demonstrated a sequence-dependent impact of the biological behaviour of TIL from RCC. The optimal use of IL-12 in the in vitro expansion of renal TIL may result in cells with an enhanced specific anti-tumor effect.

CD3 Complex↗
PubMed1996

Modulation of 5-fluorouracil resistance in human colon tumor cell lines by azidothymidine.

5-fluorouracil (5-FU), an inhibitor of thymidylate synthase (EC 2.1.1.45), is clinically used in the treatment of several solid tumors, including colorectal, head and neck, gastric, and pancreatic cancer. The drug effectively inhibits deoxynucleoside triphosphate de novo synthesis. However, this inhibition can be circumvented by increased thymidine kinase (EC 2.7.1.21) activity. In the present study we examined the effects of 5-FU combined with azidothymidine (AZT), a competitive inhibitor of thymidine kinase in human colon tumor cells in vitro, including three 5-FU resistant cell lines. The cells were simultaneously incubated with various concentrations of 5-FU (0.015 to 150 microM) and AZT (20 to 300 microM) for 6 days. 5-FU alone yielded an IC50 of 18 microM in the parental CCL 227 cell line and IC50s of 470 and 1100 microM in the 5-FU resistant cell lines as determined by a MTT chemosensitivity assay. Addition of 100 microM AZT alone, a drug concentration that can be achieved in patients, had no effect on the growth of the cell lines examined. However, when added simultaneously with 5-FU, the IC50s of 5-FU synergistically decreased to 10 microM in the sensitive and to 360 or 760 microM in the resistant cell lines, respectively. Our results demonstrate that the combination of 5-FU with AZT synergistically inhibited the growth of 5-FU resistant cells, suggesting the use of 5-FU in combination with AZT for the treatment of 5-FU sensitive as well as resistant human colon tumors.

Antimetabolites, Antineoplastic↗
PubMed1995

Pharmacokinetics of 4'-O-tetrahydropyranyladriamycin given on a weekly schedule in patients with advanced breast cancer.

Improved quality of life has gained importance over shortly lasting remissions in yet incurable metastatic breast cancer. Fractionation of drug administration is one of the possible approaches to reduce the concentration-dependent toxicity of anthracyclines. We evaluated the pharmacokinetics of 4'-O-tetrahydropyranyladriamycin (THP-ADM) under weekly administration in patients with advanced breast cancer (dose escalation, from 20 to 27 mg/m2 THP-ADM). The concentration-time curves of THP-ADM in plasma were best described by an open three-compartment model [half-life of the first disposition phase (t1/2 alpha), 3.15 min; terminal elimination half-life (t1/2 gamma), 13.9 h] with a mean area under the curve (AUC) of 12.2 ng h ml-1mg-1 m-2, resulting in a mean plasma clearance of 86.9 1h-1 m-2. Metabolism included the formation of Adriamycin (ADM), Adriamycinol (ADM-OH), 13-dihydro-4'-O-tetrahydropyranyladriamycin (THP-OH), 7-deoxyadriamycinone (7H-ADn), and 7-deoxy-13-dihydroadriamycinone (7H-ADn-OH), with maximal plasma concentrations ranging from 2.8 to 5.5 ng/ml. The mean total amount of cytotoxic anthracyclines excreted into urine, mainly as the parent drug, was 5% of the delivered dose. ADM and ADM-OH, but not the parent drug, were observed in urine at up to 4 weeks after the last therapeutic cycle. There was a significant correlation between the leukocyte nadir under therapy and the AUC of ADM-OH (r = 0.800, P < 0.05). Since no shift in the plasma kinetics was observed from the first to the sixth cycle, the favorable ratio of the AUCs of THP-ADM and ADM after fractionation of THP-ADM suggests lower toxic side effects attributable to ADM. This hypothesis was confirmed in a clinical study, where no severe cardiotoxicity and only mild alopecia were observed in 19 patients. Thus, pharmacokinetics studies might be helpful in both individualization of therapy with THP-ADM and optimization of the administration schedule.

Adult↗
PubMed1995

Cutaneous side effects in breast cancer patients treated with cytostatic polychemotherapy and rh GM-CSF: immune phenomena or drug toxicity?

The application of recombinant colony stimulating factors for chemotherapy induced granulocytopenia is becoming common in clinical oncology. Here we report on localized cutaneous side effects after subcutaneous administration of recombinant human granulocyte-macrophage colony-stimulating factor (rh GM-CSF) in 11 patients with breast cancer receiving cytostatic treatment. Seven patients suffering from inflammatory breast cancer received cytostatic chemotherapy with mitoxantrone/cyclophosphamide, whereas four patients suffering from noninflammatory breast cancer received high-dose epirubicin/cyclophosphamide, respectively. rh GM-CSF was applicated subcutaneously in a dose of 5 micrograms/kg/d for at least ten days. In all patients, sharply demarked, maculous itching and burning erythemas restricted to the injection sites occurred after three to four injections of rh GM-CSF. These eruptions cleared within 2 to 3 weeks, but reappeared after reexposure to rh GM-CSF. In contrast to previous sporadic reports, no generalized erythemas were observed. Because of this unexpected and subjectively intolerable side effect, rh GM-CSF administration had to be interrupted in all patients. Histopathological findings revealed skin infiltration with lymphocytes, monocytes/macrophages, neutrophils, and occasionally eosinophils, respectively. Since GM-CSF is known to alter immune functions, it seems likely that the eruptions were at least in part due to local immune reactions.

Adenocarcinoma↗
PubMed1995

Stereospecific pharmacokinetics of rac-5-methyltetrahydrofolic acid in patients with advanced colorectal cancer.

1. The pharmacokinetics and toxicity of racemic 5-methyltetrahydrofolic (rac-5-MTHF) acid after i.v. infusion were investigated in 18 patients with advanced colorectal cancer. Doses of 100-600 mg rac-5-MTHF/m2 were administered over 2 h together with a bolus of 500 mg/m2 5-fluorouracil (5-FU) as a midpoint injection. 2. The pharmacokinetics of both diastereoisomers were linear in the range from 100-600 mg 5-MTFH/m2. Independent of the administered dose, the maximal plasma concentration of [R]-5-MTHF was nearly twice that of [S]-5-MTHF. The elimination of [S]-5-MTHF from plasma was considerably faster than that of the [R]-isomer (elimination half-life: 3.1 +/- 1.0 h vs 8.3 +/- 3.2 h). No metabolites were detected in plasma and in urine samples. 3. The plasma protein binding was stereoselective ([R]-5-MTHF bound: 88.2 +/- 2.7%; [S]-5-MTHF bound: 59.9 +/- 6.8%; P < 0.001), causing a significantly higher renal clearance for [S]-5-MTHF when compared with the [R]-isomer (37.5 +/- 23.7 ml min-1 vs 12.7 +/- 11.2 ml min-1, P < 0.001). There was no dose dependence, but gender influenced renal clearance (CLren[R]-5-MTHF: male vs female: 20.5 +/- 14.5 ml min-1 vs 7.8 +/- 4.7 min-1, P = 0.03; CLren [S]-5-MTHF: male vs female: 57.2 +/- 21.7 ml min-1 vs 25.7 +/- 16.2 ml min-1, P = 0.006). 4. Toxic side effects of the combination 5-FU/5-MTHF were rare and generally mild, and included stomatitis, nausea/emesis, diarrhoea, anaemia, leukopenia, and thrombocytopenia. 5. In combination with 500 mg 5-FU/m2 a single dose of 600 mg rac-5-MTHF/m2 can safely be administered to patients with colorectal cancer. A similar therapeutic benefit of 5-MTHF to folinic acid in the biochemical modulation of 5-FU is supported by the comparison of in vitro and in vivo data.

Aged↗
PubMed1994

Different stereospecific protein binding of tetrahydrofolates to human serum albumin.

The protein binding of the tetrahydrofolates folinic acid (FA) and its metabolite 5-methyltetrahydrofolic acid (5-MTHF) to human serum albumin (HSA) is stereoselective. At therapeutically relevant concentrations of tetrahydrofolate (range, 5-100 microM), the protein binding was stereoselective to the (R)-isomers of FA and 5-MTHF. The binding of (R)-FA and (R)-5-MTHF was saturated at a concentration of 7% HSA [(R)-tetrahydrofolate bound, ca. 80%]. In contrast to (S)-FA, which was not bound to HSA, (S)-5-MTHF was bound to 45% under physiological conditions. (R)-FA did not influence the protein binding of (S)-FA. Hypoalbuminemia is common in patients with advanced colorectal cancer and affects differentially the protein binding of the diastereoisomers of FA and 5-MTHF. Thus, an influence on the biochemical modulation of 5-fluorouracil by tetrahydrofolates should be taken into consideration.

Chromatography, High Pressure Liquid↗
PubMed1994

Double modulation of 5-fluorouracil by high-dose leucovorin and interferon alpha 2b in advanced colorectal cancer: a phase I and a phase II study of weekly administration.

In an attempt to evaluate the feasibility of 5-fluorouracil (FU) treatment modulated by (R,S)-leucovorin (LV) and interferon alpha (IFN alpha) in patients with advanced colorectal cancer, we conducted a phase I trial with increasing doses of subcutaneous IFN alpha (3 x 1 x 10(6) U, 3 x 3 x 10(6) U, 3 x 3 x 10(6) U, 3 x 5 x 10(6) U and 3 x 10 x 10(6) U/week) and 500 mg/m2 LV i.v. as a 2-h infusion with 600 mg/m2 FU i.v. as a midpoint injection. Unacceptable side-effects occurred in all 3 patients at the highest dose level of IFN alpha, while toxicity was tolerable at 3 x 5 x 5 x 10(6) U IFN alpha/week. Thus, this dose was defined as the maximal tolerable dose for IFN alpha in combination with FU and LV. In a subsequent phase II study a total of 83 treatment courses (median: 2.8, range: 2-10) were administered to 30 evaluable patients. Side-effects were acceptable with no WHO grade IV toxicities. Grade III toxicities consisted in thrombopenia (2/30), stomatitis (2/30), diarrhoea (3/30) and nausea/vomiting (4/30). After a median observation time of 17 months (range: 8-22 months), no complete remission was observed and 9 patients experienced a partial response lasting for a median of 6.6 months (range: 3-13+ months), for an overall response rate of 30% (95% confidence interval: 15%-49%). These results show that the described regiment of FU doubly modulated by LV and IFN alpha is active in colorectal cancer and can be safely administered in an out patient setting with acceptable toxicity. Thus, this regimen is suitable to be used for further evaluation in clinical trials.

Adult↗
PubMed1994

Pharmacokinetics of rac-leucovorin vs [S]-leucovorin in patients with advanced gastrointestinal cancer.

1. The pharmacokinetics of [R]-leucovorin ([R]-LV), [S]-leucovorin ([S]-LV) and the circulating metabolite [S]-5-methyltetrahydrofolate ([S]-5-MTHF) were studied after administration of racemic LV and [S]-LV in 21 subjects. 2. After intravenous infusion of 600 mg m-2 rac-LV (group 1, n = 7) or 300 mg m-2 [S]-LV (group 3, n = 7), the decay of [S]-LV in plasma was biexponential with a distribution half-life of 0.8 to 1 h and an elimination half-life of 11 to 23 h. When rac-LV was administered as a 2 h i.v. infusion (400 mg m-2) following a loading dose of 200 mg m-2 (group 2, n = 7), the plasma concentrations of [R]-LV and [S]-5-MTHF decayed monoexponentially with mean (+/- s.d.) half-lives of 10 +/- 3 h and 7 +/- 2 h, respectively. 3. The AUC of [S]-5-MTHF was significantly higher after infusion of 300 mg m-2 [S]-LV than after infusion of 600 mg m-2 rac-LV (83 +/- 22 micrograms ml-1 h vs 53 +/- 22 micrograms ml-1 h; P = 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1994

Folate absorption in Crohn's disease.

An oral folate absorption test was performed in 100 consecutive patients with Crohn's disease (CD) and 20 healthy individuals to investigate the frequency of abnormal folate absorption in regard to the site of the disease and to investigate the possibility of defining a subgroup of patients requiring parenteral folate supplementation. The described oral folate absorption test can be performed quickly on an outpatient basis and is capable of distinguishing patients with altered folate absorption from those with normal folate absorption. In 25 patients, abnormal folate absorption was detected. 16 patients showed impaired folate absorption as indicated by a marked but insufficient increase in serum folate levels after oral folate intake, whereas no increase of the serum folate levels was detected in the remaining 9 patients. Abnormal folate absorption was not correlated with disease extent or activity. In patients with only impaired folate absorption, it might be sufficient to increase dietary intake of folates. In the remaining patients with no measurable increase of serum folate levels after oral folate intake, i.e. about 10% of all patients with CD, parenteral folate supplementation could be considered.

Administration, Oral↗
PubMed1994

Comparison of an immunoradiometric and an immunoluminometric assay for the evaluation of the tumour associated antigens CA 19-9 and CA 125.

We evaluated the levels of CA 19-9 and CA 125 in the sera of healthy individuals, patients with non-neoplastic diseases known to produce elevated serum concentrations of CA 19-9 and CA 125, and patients with malignant tumours. The serum concentrations determined with an immunoradiometric assay (IRMA) and an immunoluminometric assay (ILMA) were compared. The accuracy was determined (as far as this is possible in the absence of reference method values), as well as the precision (intra-assay variation and inter-assay variation), using internal and external controls. The serum concentrations were comparable in both test systems (coefficient of correlation, Kendall's Tau of CA 19-9: 0.88, p < 0.001; CA 125: 0.87, p < 0.001). The linearity of both assays was excellent when serum samples were diluted (r > 0.98 in all assays tested). The intra-assay variation of CA 19-9 IRMA was less than that of CA 19-9 ILMA, and was comparable for CA 125 in both assays. The coefficients of variation of duplicates were nearly independent of the antigen concentration within the range 20-1000 kU/l (mean CV CA 19-9: 3.7% and 3.9% for IRMA and ILMA, respectively; mean CV CA 125: 3.9% and 5.6% for IRMA and ILMA, respectively). In the assay of external controls, the performance of the IRMAs was found to be slightly better than that of the ILMAs, but it was not satisfactory. We conclude that, with some improvements in quality control, luminescence assays are a possible alternative to isotopic assays in the clinical laboratory.

Antigens, Tumor-Associated, Carbohydrate↗
PubMed1993

Determination of methotrexate in human urine at nanomolar levels by high-performance liquid chromatography with column switching.

An high-performance liquid chromatographic method with column switching for the detection of less than 4 ng of methotrexate in the urine of oncologic nurses is described. Urine samples were purified by solid-phase extraction on silica-bonded phenyl columns, eluting impurities with ethyl acetate. After elution from the column, the analyte was concentrated ten-fold, evaporating the solvent. On a strong anion-exchange column (Nucleosil 100 SB), methotrexate was separated from the remaining interfering substances, was then switched to a reversed-phase column (LiChrospher 100 RP-18e), and finally eluted by a linear gradient in a solvent system consisting of ammonium formate buffer (pH 2.7) and acetonitrile. Absorbance was monitored at 310 nm. This method has proved to be suitable for detecting traces of methotrexate in urine in order to individualize risks and to reduce further the occupational safety hazard for hospital personnel.

Bone Neoplasms↗
PubMed1993

GM-CSF in the treatment of a patient with severe methotrexate intoxication.

This case report describes the successful treatment of severe methotrexate intoxication in a 72-year-old female patient. Following two prior uneventful courses of a polychemotherapy regimen including low-dose intravenous (i.v.) methotrexate, the patient presented with fever, polymucositis, incipient pyodermia, acute renal failure and pancytopenia 9 days after the third application. Severe methotrexate overdose was confirmed by serum levels. Using a polypragmatic treatment approach focusing on renal function and including granulocyte-macrophage-colony-stimulating factor (GM-CSF) this life threatening and nearly fatal intoxication was successfully treated. This case report demonstrates that GM-CSF might contribute to rapid reconstitution of leukopoiesis once methotrexate serum levels are in the subtoxic range.

Acute Disease↗
PubMed1991

Instability of the anticancer agent etoposide under in vitro culture conditions.

Degradation of etoposide is rapid under in vitro culture conditions. At pH 7.4 and 37 degrees C, the isomerisation of trans-etoposide to the inactive compound cis-etoposide has a half-life of 2 days in Dulbecco's modified Eagle's medium and results in the loss of 90% of the active drug within 1 week. As a consequence, prolonged incubations with etoposide in in vitro assays may lead to erroneous interpretations ignoring the real in vitro situation. The degradation is not influenced by organic compounds such as bovine serum albumin or amino acids but depends strongly on the pH value and, to a lesser degree, on the ionic strength of the medium. Therefore, we propose a mathematical correction based on the pH value so as to obtain the real exposure of cells to trans-etoposide during in vitro assays.

Cells, Cultured↗