Subject(s)
Antineoplastic Combined Chemotherapy Protocols/adverse effects , Herpes Simplex/chemically induced , Opportunistic Infections/chemically induced , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Simplexvirus/drug effects , Virus Activation/drug effects , Adolescent , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Asparaginase/adverse effects , Asparaginase/therapeutic use , Bacterial Infections/chemically induced , Child, Preschool , Cyclophosphamide/adverse effects , Cyclophosphamide/therapeutic use , Cytarabine/adverse effects , Cytarabine/therapeutic use , Daunorubicin/adverse effects , Daunorubicin/therapeutic use , Encephalitis, Herpes Simplex/chemically induced , Encephalitis, Herpes Simplex/diagnosis , Fatal Outcome , Female , Hematopoietic Stem Cell Transplantation , Herpes Simplex/virology , Humans , Induction Chemotherapy , Infant , Male , Mercaptopurine/adverse effects , Mercaptopurine/therapeutic use , Methotrexate/adverse effects , Methotrexate/therapeutic use , Opportunistic Infections/virology , Prednisone/adverse effects , Prednisone/therapeutic use , Stomatitis, Herpetic/chemically induced , Stomatitis, Herpetic/virology , Vincristine/adverse effects , Vincristine/therapeutic useABSTRACT
Erucylphosphocholine (ErPC) exerts strong anticancer activity in vivo and in vitroand induces apoptosis even in chemoresistant glioma cell lines. We investigated the contribution of Apaf-1 and caspase-3 to the apoptotic response to ErPC using RNA interference (RNAi) in human glioblastoma cells. We could demonstrate that human glioma cell lines are susceptible to RNAi. Apaf-1 and caspase-3 are amenable to specific small interfering RNA (siRNA)-induced degradation resulting in a reduction of protein levels to 8-33% (Apaf-1) and to 30-50% (caspase-3). Transfection of siRNA directed to Apaf-1 and caspase-3 specifically reduced caspase-3 processing induced by ErPC treatment and yielded a reduction in cells that undergo ErPC-induced apoptosis to 17-33% (Apaf-1) and to 38-50% (caspase-3). The caspase-3 siRNA experiments were corroborated in caspase-3-deficient and -reconstituted MCF-7 breast cancer cells. Survival assays and morphological observations revealed that caspase-3 reconstitution significantly sensitized MCF-7 cells to ErPC. Exploring the caspase cascade responsible for ErPC-induced apoptosis MCF-7 cells provided evidence that caspase-3 is required for the activation of caspases-2, -6 and -8 and also participates in a feedback amplification loop. Our results provide evidence that Apaf-1 and caspase-3 are major determinants of ErPC-induced apoptosis and the possible use of ErPC in a clinical setting is discussed.
Subject(s)
Apoptosis/drug effects , Caspases/biosynthesis , Down-Regulation/physiology , Glioblastoma/physiopathology , Intracellular Signaling Peptides and Proteins/metabolism , Phosphorylcholine/analogs & derivatives , Proteins/metabolism , RNA Interference , Apoptosis/physiology , Apoptotic Protease-Activating Factor 1 , BH3 Interacting Domain Death Agonist Protein/metabolism , Breast Neoplasms/physiopathology , Caspase 3 , Cell Line, Tumor , Cell Proliferation/drug effects , Humans , Phosphorylcholine/pharmacology , Proto-Oncogene Proteins c-bcl-2/physiologyABSTRACT
Novel isolates of Achromobacter xylosoxidans, Tsukamurella wratislaviensis and a Rhodococcus sp. are described. These grew with short-chain alkanesulfonates as their sole source of carbon and energy. T. wratislaviensis strain SB2 grew well with C(3)-C(6) linear alkanesulfonates, isethionate and taurine, Rhodococcus sp. strain CB1 used C(3)-C(10) linear alkanesulfonates, taurine and cysteate, but neither strain grew with ethanesulfonate. In contrast, A. xylosoxidans strain AE4 grew well with ethanesulfonate, making it the first bacterium to be described which can grow with this compound. It also grew with unsubstituted C(3)-C(5) alkanesulfonates and isethionate. Hydrolysis was excluded as a mechanism for alkanesulfonate metabolism in these strains; and evidence is given for a diversity of uptake and desulfonatase systems. We provide evidence for an initial monooxygenase-dependent desulfonation in the metabolism of ethanesulfonate and propanesulfonate by A. xylosoxidans strain AE4.
Subject(s)
Actinomycetales/metabolism , Alcaligenes/enzymology , Alkanesulfonates/metabolism , Oxygenases/metabolism , Rhodococcus/metabolism , Actinomycetales/growth & development , Alcaligenes/growth & development , Alkanesulfonic Acids/metabolism , Oxidation-Reduction , Rhodococcus/growth & development , Species Specificity , Sulfites/metabolismABSTRACT
BACKGROUND: Cisplatin is a highly effective and frequently used drug in the chemotherapy of solid tumours in children, but only limited data are available on the pharmacokinetics of cisplatin and its associated nephrotoxicity in paediatric patients. METHODS: We investigated the pharmacokinetics of free platinum (Pt) in 12 children (25 courses) receiving cisplatin (75-120 mg/m2) either as a continuous 72-h infusion, prolonged single 6-h infusion or repetitive 1-h infusions. Plasma and urinary Pt concentrations were analysed using atomic absorption spectroscopy. Cisplatin-induced nephrotoxicity was determined using creatinine clearance and several glomerular and tubular marker proteins. RESULTS: Using a two-compartment model the pharmacokinetic parameters for free Pt were: initial half-life 21.6 +/- 9.6 min, terminal half-life 25.9 +/- 16.2 h, area under the plasma concentration-time curve (AUC) 13.5 +/- 4.97 (microg/ml) x h/(100 mg/m2) and cumulative renal elimination(infinity) 41.7 +/- 6.6% of dose. Higher cisplatin delivery rates led to higher peak concentrations of free Pt in plasma and urine and to lower cumulative renal Pt elimination (P < 0.01). During all courses, increases of urinary albumin and alpha1-microglobulin excretion were documented. The creatinine clearance decreased significantly to 70% of baseline values. Correlations were found between both peak free Pt concentrations in plasma and in urine and the maximum of urinary excretions of albumin and of N-acetyl-beta-D-glucosaminidase and the nadir of the glomerular filtration rate (P < 0.05). CONCLUSIONS: With respect to nephrotoxicity, long-term infusions of cisplatin seem to be preferable over intermittent bolus administration in paediatric patients. The best predictive pharmacokinetic parameters for cisplatin-associated nephrotoxicity in children are peak free Pt concentrations in plasma and urine.
Subject(s)
Antineoplastic Agents/adverse effects , Antineoplastic Agents/pharmacokinetics , Cisplatin/adverse effects , Cisplatin/pharmacokinetics , Kidney Diseases/chemically induced , Trypsin Inhibitor, Kunitz Soybean , Adolescent , Adult , Albuminuria/blood , Albuminuria/chemically induced , Albuminuria/urine , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/blood , Area Under Curve , Biomarkers/blood , Biomarkers/urine , Child , Child, Preschool , Cisplatin/administration & dosage , Creatinine/blood , Creatinine/urine , Drug Administration Schedule , Female , Half-Life , Humans , Infusions, Intravenous , Kidney/drug effects , Kidney/metabolism , Kidney Diseases/blood , Kidney Diseases/urine , Male , Membrane Glycoproteins/urine , Neoplasms/complications , Neoplasms/metabolismABSTRACT
Induction of differentiation represents a promising concept for chemotherapy of malignant gliomas, which are often refractory even to the combined treatment with surgery, irradiation and chemotherapy. Since anti-neoplastic alkylphosphocholines can induce differentiation of leukemic cell lines, the effects of the intravenously applicable alkylphosphocholine-derivative erucylphosphocholine (ErPC) on proliferation, morphology and differentiation of the rat glioma cell line C6 was examined in vitro. Short-term exposure to ErPC induced accumulation of the cells in the G2/M-phase of the cell cycle and apoptotic cell death. In contrast, continuous exposure of C6 rat glioma cells to sublethal ErPC doses (30 and 50 microM) caused both the formation of a slower growing tetraploid cell population and astrocytic differentiation. No resistance to in vivo obtainable ErPC concentrations was observed during this treatment. We conclude that ErPC-induced differentiation might be beneficial for a long-term adjuvant chemotherapy of low grade glioma.
Subject(s)
Antineoplastic Agents/pharmacology , Astrocytes/drug effects , Brain Neoplasms/drug therapy , Cell Differentiation/drug effects , Glioma/drug therapy , Phosphorylcholine/analogs & derivatives , Phosphorylcholine/pharmacology , Ploidies , Tumor Cells, Cultured/drug effects , Animals , Apoptosis/drug effects , Brain Neoplasms/metabolism , Cell Division/drug effects , Flow Cytometry , Glioma/metabolism , Rats , Sensitivity and SpecificityABSTRACT
Little is known about the association between the rate of cisplatin administration and the severity of cisplatin-induced renal damage in children. The purpose of this study was to compare severity and reversibility of renal damage in children after continuous and repetitive bolus administration of cisplatin and to correlate these data with pharmacokinetic parameters. Study subjects included six children (ten courses) receiving cisplatin as 1-h bolus infusions on three consecutive days (3x40 mg/m2) and four children (eight courses) receiving 72-h continuous infusions (120 mg/m2). In all courses, signs of glomerular and tubular damage were seen, as evidenced by elevated urinary excretion of alpha1-microglobulin, albumin and N-acetyl-beta-D-glucosaminidase and decreased glomerular filtration rate (GFR). Comparing the two infusion regimens, the 1-h bolus administration of cisplatin was followed by significantly higher peak free platinum concentrations in plasma and urine (P<0.001), resulting in lower nadirs of the GFR (P<0.005). Correlations were found between both peak free platinum concentrations in plasma and urine and maxima of urinary albumin and N-acetyl-beta-D-glucosaminidase excretion. Within 12 months after completion of cisplatin therapy, children in the 1-h bolus group had recovered only partially from subclinical nephrotoxicity, with five out of six showing pathological proteinuria. The results provide clear evidence that long-term ciplatin infusions are less nephrotoxic than repetitive bolus infusions.
Subject(s)
Antineoplastic Agents/adverse effects , Cisplatin/adverse effects , Kidney Diseases/chemically induced , Adolescent , Antineoplastic Agents/pharmacokinetics , Area Under Curve , Biomarkers , Child , Child, Preschool , Cisplatin/pharmacokinetics , Female , Half-Life , Humans , Infusions, Intravenous , Kidney Diseases/pathology , Kidney Glomerulus/pathology , Male , Platinum/blood , Proteinuria/chemically inducedABSTRACT
Hereditary methemoglobinemia due to reduced nicotin amide adenine dinucleotide (NADH)-cytochrome b5 reductase (b5r) deficiency is classified into an erythrocyte type (I) and a generalized type (II). We investigated the b5r gene of three unrelated patients with types I and II and found four novel mutations. The patient with type I was homozygous for a c.535 G-->A exchange in exon 6 (A179T). The patients with type II were found to be homozygous for a c.757 G-->A transition in exon 9 (V253M) and compound heterozygous for two mutations, respectively. One allele presented a c.379 A-->G transition (M127V). The second allele carried a sequence difference at the invariant 3' splice-acceptor dinucleotide of intron 4 (IVS4-2A-->G) resulting in skipping of exon 5. To characterize a possible effect of this mutation on RNA metabolism, poly(A)(+) RNA was analyzed by RT-PCR and sequencing. The results show that RNA is made from the allele harboring the 3'-splice site mutation. Furthermore, western blot analysis revealed a complete absence of immunologically detectable b5r in skin fibroblasts of this patient. The compound heterozygosity for the splice site and the missense mutations apparently caused hereditary methemoglobinemia type II in this patient. Hum Mutat 17:348, 2001.
Subject(s)
Alternative Splicing/genetics , Cytochrome Reductases/genetics , Exons/genetics , Genes, Recessive/genetics , Methemoglobinemia/congenital , Methemoglobinemia/genetics , Mutation, Missense/genetics , Alleles , Child, Preschool , Consensus Sequence/genetics , Cytochrome-B(5) Reductase , DNA Mutational Analysis , Female , Fibroblasts , Genotype , Humans , Infant, Newborn , Introns/genetics , Male , Methemoglobinemia/classification , Methemoglobinemia/enzymology , Middle Aged , Polymorphism, Single-Stranded Conformational , RNA Splice Sites/genetics , RNA, Messenger/analysis , RNA, Messenger/geneticsABSTRACT
Intrinsic chemoresistance constitutes a major problem in the therapy of malignant gliomas. In vitro experiments with four astrocytoma/glioblastoma (AC/GBM) cell lines revealed that the chemoresistance of two cell lines, A172 and T98G, to cisplatin and etoposide was due to resistance to drug-induced apoptosis. In contrast, all the AC/GBM cell lines tested were sensitive to treatment with the lipophilic ether lipid erucylphosphocholine, ErPC. ErPC-induced apoptosis was independent of wild-type p53-signaling and triggering of the CD95/CD95 ligand (CD95L) system. Inhibition of protein and RNA synthesis by cycloheximide and actinomycin D did not abrogate ErPC-induced apoptosis. However, expression of members of the bcl-2 protein family was modulated during ErPC treatment. Activation of caspase 3 and mitochondrial alterations were central to ErPC-induced apoptosis. We conclude that ErPC-induced activation of the mitochondrial pathway enables cell death in the chemoresistant AC/GBM cells.
Subject(s)
Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Astrocytoma/pathology , Caspases/metabolism , Glioblastoma/pathology , Mitochondria/drug effects , Phosphorylcholine/analogs & derivatives , Phosphorylcholine/pharmacology , Animals , Apoptosis/physiology , Astrocytoma/drug therapy , Astrocytoma/enzymology , Caspase 3 , Cisplatin/pharmacology , Down-Regulation/drug effects , Drug Resistance, Multiple , Drug Resistance, Neoplasm , Enzyme Activation/drug effects , Etoposide/pharmacology , Fas Ligand Protein , Glioblastoma/drug therapy , Glioblastoma/enzymology , Humans , Intracellular Membranes/drug effects , Intracellular Membranes/physiology , Membrane Glycoproteins/physiology , Membrane Potentials/drug effects , Membrane Potentials/physiology , Mitochondria/physiology , Proto-Oncogene Proteins c-bcl-2/biosynthesis , Proto-Oncogene Proteins c-bcl-2/physiology , Rats , Signal Transduction/drug effects , Tumor Cells, Cultured , bcl-X Protein , fas Receptor/physiologyABSTRACT
The blood-brain barrier hinders progress in the chemotherapy of brain tumors due to insufficient penetration of anticancer drugs into the brain tissue. Short-chain alkylglycerols affect the physicochemical properties of biological membranes. The enhancement of the blood-brain barrier permeability by intra-arterial administration of alkylglycerols was investigated in tumor-free and C6 astroglioma bearing rats. The antineoplastic agents cisplatin and methotrexate and the antibiotics vancomycin and gentamicin were selectively injected into the right internal carotid artery in the absence and presence of various alkylmono-, alkyldi-, and alkyltriglycerols. In normal rats the intra-arterial administration of the drugs without alkylglycerols resulted in low drug concentrations in brain tissue. In the presence of alkylglycerols (0.01-0.3 M) a reversible (within minutes) and concentration-dependent enrichment of the coinjected agents was found, preferentially in the ipsilateral hemisphere. The extent of drug accumulation in the brain was modified by changes in the chemical structure of the alkylglycerols. The effect increased with the chain length of the alkyl group, decreased with the number of glycerols, and varied from 2- to more than 230-fold compared to controls. In rats with C6 tumors 1-O-pentylglycerol increased the delivery of methotrexate 18-fold in the tumor, 28-fold in the surrounding brain, 18-fold in the contralateral brain, and 19-fold in the cerebellum compared to controls with methotrexate in the absence of pentylglycerol. In conclusion, the intra-arterial administration of alkylglycerols represents a novel and well controllable method for enhanced drug delivery to the brain and to brain tumors.
Subject(s)
Anti-Bacterial Agents/pharmacokinetics , Antineoplastic Agents/pharmacokinetics , Blood-Brain Barrier/drug effects , Glycerides/pharmacology , Animals , Brain/metabolism , Brain Neoplasms/metabolism , Glioma/metabolism , Male , Neoplasm Transplantation , Rats , Rats, Wistar , Structure-Activity Relationship , Tumor Cells, CulturedABSTRACT
The clinical use of alkylphosphocholines (APC) in cancer patients is restricted because of the high gastrointestinal toxicity and the need for oral administration. Therefore we evaluated the clinical pharmacology of erucylphosphocholine (ErPC), the first derivative of the APC family suitable for intravenous administration with strong antineoplastic activity, in vitro and in vivo in rats. The pharmacokinetic parameters after a single intravenous dose of 40 mg/kg were calculated using a two-compartment model: C(max) = 1.6 +/- 0.3 micromol/ml, T(1/2alpha) = 0.18 +/- 0.09 h, T(1/2beta) = 3.3 +/- 0.88 h, clearance = 9.7 +/- 1.2 ml/h, AUC = 2.5 +/- 0.3 micromol/ml per h and Vss = 40.4 +/- 7.9 ml. Biodistribution studies were performed after repeated ErPC administration at different doses. Intravenous injections of 20 mg/kg given at intervals of 48 h for up to 4 weeks were well tolerated. Neither clinical evaluation nor laboratory parameters (haematology and clinical chemistry) revealed toxic side effects. In contrast, higher doses of ErPC (40 mg/kg per 48 h) led to weight loss. After 2 and 4 weeks of therapy with 20 mg/kg per 48 h a high ErPC accumulation was found in the adrenal glands, small intestine and brain. The brain to serum concentration ratios averaged 2.1 after 2 weeks and 4.5 after 4 weeks. Significant leucocytosis and thrombocytosis were observed after 4 weeks of ErPC treatment. The findings suggest that ErPC is a suitable candidate for clinical trials. In particular, owing to the high accumulation in brain tissue, ErPC is a potential agent for chemotherapy against malignant brain tumours.
Subject(s)
Antineoplastic Agents/pharmacokinetics , Brain/metabolism , Phosphorylcholine/analogs & derivatives , Animals , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/pharmacology , Blood Glucose/metabolism , Body Weight/drug effects , Injections, Intravenous , Leukocyte Count/drug effects , Male , Metabolic Clearance Rate , Models, Biological , Phosphorylcholine/administration & dosage , Phosphorylcholine/pharmacokinetics , Phosphorylcholine/pharmacology , Rats , Rats, Wistar , Tissue DistributionABSTRACT
A potential benefit of alkylphosphocholines in brain tumor therapy was evaluated. The in vitro effects of the intravenously applicable erucylphosphocholine (ErPC) on proliferation, viability, morphology and cell cycle distribution of a rat glioma, four human astrocytoma/glioblastoma and a human medulloblastoma cell line were analyzed daily after continuous drug-exposure for up to six days. ErPC exerted strong cytostatic and direct cytotoxic effects on all cell lines tested at drug concentrations that are achieved in the rat brain after repeated intravenous injections of nontoxic drug doses. Concentrations of 70 microM (T98G, A172, 85HG66, 86HG39) and 110 microM (C6, D283 Med) led to complete cell death within 48-96 h. Particular characteristics of ErPC action are i) the accumulation of cells with a 4n DNA content corresponding to the G2/M-phase of the cell cycle, ii) the formation of two- and multinucleated cells and iii) the induction of apoptosis.
Subject(s)
Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Brain Neoplasms/drug therapy , Phosphorylcholine/analogs & derivatives , Animals , Blood-Brain Barrier , Brain/metabolism , Brain Neoplasms/pathology , DNA Fragmentation/drug effects , Humans , Phosphorylcholine/pharmacokinetics , Phosphorylcholine/pharmacology , Rats , Tumor Cells, CulturedABSTRACT
The antineoplastic activity of erucylphosphocholine (ErPC) on C6 rat glioma cells and the human glioblastoma cell lines A172 and T98G was studied in vitro. ErPC exerted concentration- and time-dependent cytostatic and cytotoxic actions. The effects of ErPC on C6 cells were stronger than hexadecylphosphocholine and similar to 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine. The human cell lines were more sensitive to ErPC (LC50 = 36 microM and 29 microM) than C6 cells (LC50 = 70 microM) (48 hours, WST-1-test). Morphological characteristics of apoptotic cell death were observed. Rats bearing intracerebral and subcutaneous gliomas were treated with intravenous ErPC for 30 days to determine the antitumor effects of ErPC in vivo. ErPC accumulation in subcutaneous tumors was higher than in brain tissue. Peripheral tumors revealed a better response than CNS-tumors. There was a strong negative relationship between the concentration of ErPC and the weight of peripheral tumors. ErPC is considered to be promising for chemotherapy of malignant brain tumors.