Bernhard Erdlenbruch

Intensive chemotherapy improves survival in pediatric high‐grade glioma after gross total resection: results of the HIT‐GBM‐C protocol

The authors hypothesized that intensified chemotherapy in protocol HIT‐GBM‐C would increase survival of pediatric patients with high‐grade glioma (HGG) and diffuse intrinsic pontine glioma (DIPG).

Alkylglycerol opening of the blood-brain barrier to small and large fluorescence markers in normal and C6 glioma-bearing rats and isolated rat brain capillaries.

The blood–brain barrier (BBB) represents the major impediment to successful delivery of therapeutic agents to target tissue within the central nervous system. Intracarotid alkylglycerols have been shown to increase the transfer of chemotherapeutics across the BBB. We investigated the spatial distribution of intracarotid fluorescein sodium and intravenous lissamine‐rhodamine B200 (RB 200)–albumin in the brain of normal and C6 glioma‐bearing rats after intracarotid co‐administration of 1‐O‐pentylglycerol (200 mM). To elucidate the mechanisms involved in the alkylglycerol‐mediated BBB opening, intraluminal accumulation of fluorescein isothiocyanate (FITC)–dextran 40,000 was studied in freshly isolated rat brain capillaries using confocal microscopy during incubation with different alkylglycerols. Furthermore, 1‐O‐pentylglycerol‐induced increase in delivery of methotrexate (MTX) to the brain was evaluated in nude mice. Microscopic evaluation showed a marked 1‐O‐pentylglycerol‐induced extravasation of fluorescein and RB 200–albumin in the ipsilateral normal brain. In glioma‐bearing rats, increased tissue fluorescence was found in both tumor tissue and brain surrounding tumor. Confocal microscopy revealed a time‐ and concentration‐dependent accumulation of FITC–dextran 40,000 within the lumina of isolated rat brain capillaries during incubation with 1‐O‐pentylglycerol and 2‐O‐hexyldiglycerol, indicating enhanced paracellular transfer via tight junctions. Intracarotid co‐administration of MTX and 1‐O‐pentylglycerol (200 mM) in nude mice resulted in a significant increase in MTX concentrations in the ipsilateral brain as compared to controls without 1‐O‐pentylglycerol (P<0.005). In conclusion, 1‐O‐pentylglycerol increases delivery of small and large compounds to normal brain and brain tumors and this effect is mediated at least in part by enhanced permeability of tight junctions.

Erucylphosphocholine: Pharmacokinetics, biodistribution and CNS-accumulation in the rat after intravenous administration.

Abstract 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: Cmax= 1.6 ± 0.3 μmol/ml, T1/2α= 0.18 ± 0.09 h, T1/2β= 3.3 ± 0.88 h, clearance = 9.7 ± 1.2 ml/h, AUC = 2.5 ± 0.3 μmol/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.

Intracarotid administration of short‐chain alkylglycerols for increased delivery of methotrexate to the rat brain

The intracarotid administration of alkylglycerols has been reported previously by us to be a novel strategy for increased delivery of various chemotherapeutic drugs to the normal brain and brain tumors in rats. Effectiveness and structure–activity relations of the most promising pentyl‐ and hexylglycerol derivatives have been elucidated in vivo by analyzing the transfer of methotrexate (MTX) across the blood–brain barrier (BBB) in normal rats. The effects were compared with BBB disruption using hypertonic mannitol or intracarotid infusion of bradykinin. Furthermore, toxicity of the alkylglycerols has been studied in long‐term experiments. Apart from 1‐O‐pentyldiglycerol, all alkylglycerols induced a concentration‐dependent increase in MTX delivery to the brain varying from 1.1 to more than 300‐fold compared to intra‐arterial MTX alone. Enhanced barrier permeability rapidly approached baseline values within 5 and 120 min at the latest. Chemical structure, concentration, time schedule of injections and combination of different alkylglycerols were identified as instruments suited to regulate the MTX accumulation within a wide range. Mannitol 1.4 M resulted in very high MTX levels in the brain as observed using the highest concentrations of alkylglycerols. Intracarotid infusion of bradykinin had only a minor effect on the BBB. Using 1‐O‐pentylglycerol or 2‐O‐hexyldiglycerol, both cell culture experiments and long‐term in vivo analyses including clinical, laboratory and histopathological evaluations revealed no signs of toxicity. In summary, intracarotid short‐chain alkylglycerols constitute a very effective and low toxic strategy for transient opening of the BBB to overcome the limited access of cytotoxic drugs to the brain.

Oral topotecan in children with recurrent or progressive high-grade glioma: A Phase I/II study by the German Society for pediatric oncology and hematology

Continuous oral treatment with topotecan may be more effective than the typical 1‐day and 5‐day treatment schedules. In previous studies of continuous treatment with topotecan, increased intestinal side effects were reported in adult patients; however, the experience in pediatric patients and patients with high‐grade glioma is quite limited.

Cisplatin nephrotoxicity in children after continuous 72-h and 3×1-h infusions

Abstract Little is known about the association be- tween 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) re-ceiving cisplatin as 1-h bolus infusions on three consecutive days (3×40 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 α1-microglobulin, albumin and N-acetyl-β-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-β-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.

Erucylphosphocholine‐induced apoptosis in glioma cells: involvement of death receptor signalling and caspase activation

Erucylphosphocholine (ErPC) is a promising anti‐neoplastic drug for the treatment of malignant brain tumours. It exerts strong anti‐cancer activity in vivo and in vitro and induces apoptosis even in chemoresistant glioma cell lines. The purpose of this study was to expand on our previous observations on the potential mechanisms of ErPC‐mediated apoptosis with a focus on death receptor activation and the caspase network. A172 and T98G glioma cells were treated with ErPC for up to 48 h. ErPC effects on the expression of the tumour necrosis factor (TNF) and TNF‐related apoptosis‐inducing ligand (TRAIL) receptor system, and on caspase activation were determined. ErPC had no effect on the expression of TNFα or TRAIL. Inhibition of the TNF or TRAIL signalling pathway with antagonistic antibodies or fusion proteins did not affect apoptosis induced by ErPC, and a dominant‐negative FADD construct did not abolish ErPC‐induced effects. Western blot analysis indicated that ErPC‐triggered apoptosis resulted in a time‐dependent processing of caspases‐3, ‐7, ‐8 and ‐9 into their respective active subunits. Co‐treatment of A172 cells with different caspase inhibitors prevented apoptosis but did not abrogate cell death. These data suggest that A172 cells might have an additional caspase‐independent pathway that insures cell death and guarantees killing of those tumour cells whose caspase pathway is incomplete.

Blood-brain barrier opening with alkylglycerols: Biodistribution of 1-O-pentylglycerol after intravenous and intracarotid administration in rats

Short-chain alkylglycerols have been described to increase the penetration of drugs and macromolecules across the blood–brain barrier (BBB) into the central nervous system (CNS) and were considered to be of potential value in the pharmaceutical treatment of CNS disorders. Due to the lack of information on the pharmacological behavior of these compounds in vivo, pharmacokinetics and biodistribution of [14C]- and [3H]-labeled 1-O-pentylglycerol (49 mg/kg, 100 mM) was investigated in normal male Wistar rats after intravenous as well as intracarotid administration. There was a rapid and predominant renal elimination of 1-O-pentylglycerol and more than 70% of administered dose was found in the urine within 270 min. Analysis of the pharmacokinetic parameters after a single i.v. bolus injection of 1-O-pentylglycerol resulted in a peak blood concentration of 0.58±0.06 μmol/ml, an initial half life of 23±7 min and a terminal half life of 18.8±4.1 h. No accumulation of 1-O-pentylglycerol was observed in the brain or other organs while highest concentrations were found in liver and thymus. This was confirmed by autoradiographic studies. Five minutes after intracarotid administration, high radioactivity was found in the ipsilateral brain, whereas after 30 min radioactivity in the brain has dramatically decreased. Autoradiographic images gave evidence of biliary excretion in addition to the renal elimination. There were no signs of cleavage of the O-alkyl bond in vivo as demonstrated by HPLC analysis. In conclusion, 1-O-pentylglycerol is characterized by pharmacological properties appearing very favorable for in vivo use as a permeabilizing drug for increased drug delivery to the brain.

Pregnancy and delivery of a healthy baby in autoimmune Lambert-Eaton myasthenic syndrome

Sirs: Autoimmune diseases are not uncommon in young women in the child bearing age. In myasthenia gravis, an autoimmune disorder of the neuromuscular synapse, during pregnancy worsening of the mother’s condition and complications like preterm rupture of amniotic membranes, delivery problems and arthrogryposis or neonatal myasthenia gravis in the newborn due to passive antibody transfer from the mother to the child may occur. [1–4, 6, 7] Lambert-Eaton myasthenic syndrome (LEMS), another autoimmune disease of the neuromuscular synapse, is associated with antibodies (AB) directed at presynaptic voltage-gated Ca channels (VGCC) of the neuromuscular junction. LEMS may occur in the course of small lung cell cancer and as a non-paraneoplastic disorder. [5] In the literature pregnancies have not been reported in this disease. We here describe a pregnancy and delivery of a healthy baby in a 30 year old woman with a 5-year history of non-paraneoplastic LEMS. The 30 year old female patient had been diagnosed with LEMS at the age of 25 years, five years before her pregnancy. The diagnosis was based on typical clinical and laboratory findings, i.e. predominantly proximal leg muscle weakness, rapid exhaustion after exercise, mouth dryness, an increment of about 400% of the ulnar CMAP amplitude after maximal voluntary contraction for 30 s, and positive testing for VGCC serum AB. Examinations for cancer and for AB against acetylcholine receptors were negative. Thus, the diagnosis of a non-paraneoplastic LEMS was made. Three plasma exchanges were performed and therapy with pyridostigmine 4 · 60 mg, 3,4 diaminopyridine (3, 4 DAP) 4 · 5 mg, glucocorticosteroids and azathioprine 125 mg/day was started. Azathioprine and 3, 4 DAP could be stopped three years after the diagnosis had been made, whilst oral methylprednisolone 8mg/day and pyridostigmine 2 · 60 mg were continued. VGCC AB titers were between 108 and 124 pmol/l (normal range 0–25 pmol/l) before pregnancy. During pregnancy the patient remained stable. VGCC AB titers were 33 pmol/l in the first and 42 pmol/l in the last trimester. In the 33 gestational week routine tocolysis with fenoterol 0.04 mg/h and magnesium sulphate 1500 mg/h for treatment of premature labor induced rapidly evolving muscle weakness MRC grade 2 in proximal leg and arm muscles with inability to stand or to raise the arms. This condition improved slowly but continuously within one week after discontinuation of magnesium sulphate. In the 34 gestational week delivery was initiated by prostaglandines i.v.. Despite normal labor the male baby had to be delivered by forceps (weight at birth 2080 g, Apgar values 8/9/9, umbilical artery pH values 7.36 and 7.37). Testing for VGCC AB was negative at days 0, 5 and 14. Thorough clinical examination by the coauthoring pediatrician (B.E.) of the newborn revealed no weakness or respiratory insufficiency. Blood gas analyses were performed twice daily during the first four days and once weekly thereafter and all examinations were normal. In the umbilical cord vein blood VGCC AB titers were 30.4 pmol/l. VGCC AB titers in native breast milk were negative. To the best of our knowledge this is the first report on a pregnancy and delivery of a baby in non-paraneoplastic autoimmune LEMS with elevated VGCC AB titers. Intrauterine development of the baby was normal. Postnatally, the newborn did not show any clinical signs of neonatal LEMS and VGCC AB testing was negative. It might be expected that in LEMS – by analogy with MG [4, 6, 7] – VGCC IgG ABs C. Wessig Æ K. Toyka Dept. of Neurology University of Würzburg Würzburg, Germany