Implantable biodegradable polymers for IUdR radiosensitization of experimental human malignant glioma

A geometric model for diffusion-limited radiate accretive growth is used to simulate the emergence of growth and form of a certain class of organisms and crystals. In the model, layered structures are generated by the construction of new layers on top of the previous ones, the local growth velocities are determined by the local nutrient concentration gradients. The simulation of nutrient-limited growth in three dimensions (3D) is computationally very expensive. This paper presents a methodology to formulate the geometrical problem in terms of a 3D Laplace solver and maps this solver on a parallel platform. The aim of this paper is to demonstrate that this hybrid method is capable of simulating two typical properties of radiate accretive growth: (1) the indeterminateness of the growth process; and (2) the strong impact of the physical environment on the growth process.     

Interstitial delivery of carboplatin via biodegradable polymers is effective against experimental glioma in the rat

PURPOSE: Carboplatin has shown promise experimentally as an antineoplastic agent against both primary central nervous system (CNS) tumors and several solid tumors that frequently metastasize to the brain. Unfortunately, carboplatin is limited in its clinical use for tumors in the CNS by systemic toxicity and poor penetration through the blood brain barrier. Recent advances in polymer technology have made feasible the intracranial implantation of a biodegradable polymer capable of local sustained delivery of chemotherapy for brain neoplasms. This study assessed the toxicity and efficacy of carboplatin delivered from intracranial sustained release polymers in the treatment of experimental gliomas in rodents. METHODS: Two biodegradable anhydride polymer systems were tested: a copolymer of 1,3-bis-(p-carboxyphenoxy propane) and sebacic acid, and a copolymer of fatty acid dimer and sebacic acid. The polymers were loaded with carboplatin and dose escalation studies evaluating toxicity were performed by implanting carboplatin-loaded polymers into the brains of rats. Next, efficacy was tested. F-98 glioma cells were injected intracranially into rats, and 5 days later polymers containing the highest tolerated doses were implanted at the site of tumor growth. The survival of animals receiving carboplatin-loaded polymer was compared with that of animals receiving intraperitoneal doses of the same agent. RESULTS: Carboplatin-polymer was well tolerated at doses up to 5% loading in both polymer systems. Locally delivered carboplatin effectively prolonged survival of rats with F98 gliomas. Maximal treatment effect was seen with 5% loading of either polymer, with median survival increased threefold over control (P < 0.004). Systemic carboplatin also significantly prolonged survival, but the best intracranial polymer dose was significantly more effective than the best systemic dose tested. CONCLUSIONS: Carboplatin can be safely delivered intracranially by biodegradable sustained- release polymers. This treatment improves survival in rodents with experimental gliomas, with locally delivered carboplatin being more effective than systemic carboplatin.     

Iodo-2'-deoxyuridine (IUdR) and 125IUdR loaded biodegradable microspheres for controlled delivery to the brain

The aim of this work was to develop sustained local release systems for radioiodinated iodo-2'-deoxyuridine (125IUdR) from biodegradable polymeric microspheres to facilitate the controlled delivery of 125IUdR to brain tumours. The selective uptake of IUdR into the cell nucleus results in cell disruption over the short range of the low energy Auger electrons. The biodegradable microspheres can be precisely implanted in the brain by stereotactic techniques and the IUdR within the microspheres is protected from degradation and thus a sustained source of radiolabelled IUdR is available in the vicinity of the residual tumour cells. Poly(lactic-co-glycolic acid), PLGA (85:15), microspheres containing cold IUdR and the Auger-electron emitter 125I, as 125IUdR were prepared using the O/W, O/O and W/O/W emulsion-solvent evaporation methods. The W/O/W emulsion method was most effective in achieving good drug loading with the use of bovine plasma in the internal water phase. Also effective in improving the drug loading was the use of 20% acetone in the dichloromethane and the presence of Span 40 in the organic phase. Electrolytes (NaCl and IUdR) in the external aqueous phase also improved drug loading. After an initial rapid release from the microspheres, a sustained release was observed over 15 days for the 'cold' IUdR. The sustained release portions of the release curves showed Higuchi (t1/2), diffusion controlled release kinetics. The radiolabelled IUdR microspheres showed a burst release effect of 30-40% followed by a sustained release over 35 days.     

Saccharide-assisted delivery of cytotoxic liposomes to human malignant cells

The overexpression of lectins by malignant cells was applied for in vitro targeting of liposomes equipped with a saccharide vector and loaded in the lipid phase with a lipid derivative of anticancer agent sarcolysine. The lectin specificity of human leukemia HL-60 and human lung adenocarcinoma ACL cells was revealed by tests with fluorescein-labeled sugar probes. With the help of fluorescent lipid dye it was shown that active saccharide ligands increased the level of the vectored liposome binding to malignant cells by 50-80% as compared to liposomes without vector or with inactive one. The degree of liposome/cell membrane fusion was monitored fluorometrically and was shown to be complete and independent of the vectors. The targeted drug-loaded liposomes had the cytotoxic activity 2-4 times higher as compared to the vector-free ones.     

Beta-galactosidase gene transfer to human malignant glioma in vivo using replication-deficient retroviruses and adenoviruses

Telomere maintenance has been proposed as an essential prerequisite to human tumor development. The telomerase enzyme is itself a marker for tumor cells, but the genetic alterations that activate the enzyme during neoplastic transformation have remained a mystery. Here, we show that Myc induces telomerase in both normal human mammary epithelial cells (HMECs) and normal human diploid fibroblasts. Myc increases expression of hEST2 (hTRT/TP2), the limiting subunit of telomerase, and both Myc and hEST2 can extend the life span of HMECs. The ability of Myc to activate telomerase may contribute to its ability to promote tumor formation.     

Interferon-alpha enhances CD95L-induced apoptosis of human malignant glioma cells

A new method of time-frequency analysis, based on the Matching Pursuit (MP) algorithm, was used to extract and quantify EEG 'driving' or frequency-following responses produced in human primary somatosensory cortex (SI) by 33 Hz vibrotactile stimulation of the right index fingertip in a single subject. EEG signals were recorded from a 5 x 5 array of electrodes centered over the left hand area, time-locked to repeated presentations of four vibratory stimulus amplitudes. The MP algorithm was used to decompose the edited and and filtered EEG signals into waveforms selected from a large and redundant dictionary. Statistical discrimination of the vibratory stimulus amplitudes was then readily achieved in terms of trial-by-trial measures of response amplitude constructed in automated fashion from the calculated MP parameters. The results were orderly and physiologically coherent, and potentially open the way to correlation of psychophysical magnitude estimates with measures of neurophysiological response on a trial-by-trial basis. The approach developed here appears well suited to detection and characterisation of time dependent or transient target signals embedded in a noisy background.     

Growth-inhibitory and differentiation-inducing activity of dimethylformamide in cultured human malignant glioma cells

OBJECTIVE: To determine the growth-inhibitory and differentiation-inducing activity of dimethylformamide (DMF) on a human glioma cell line (SHG-44). DMF is a type of polar solvent and a potent differentiation-inducing agent in many kinds of human solid tumors, yet its effect on human glioma remains unclear. METHODS: The effects of DMF on cell proliferation using 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, cell cycle distribution (with flow cytometry), colony-forming efficiency in double-layer soft agar, tumorigenicity in athymic nude mice, morphological changes, and glial fibrillary acidic protein expression were studied. RESULTS: At dose ranges of 0.25, 0.5, 0.75, and 1.0%, DMF caused a dose-dependent proliferation inhibitory effect in monolayers and a marked dose-dependent suppression of colony-forming efficiency in double-layer soft agar with a complete loss of colony-forming ability in cells exposed to 0.75 and 1.0% DMF. Accumulation of cells in G0/G1 phases was observed in DMF-treated (0.5 and 1.0%) cells, also in a dose-dependent manner. SHG-44 cells exposed to DMF (0.5 and 1.0%) for 15 days changed morphologically from small spindle-shaped to large polygonal and flattened stellate cells with multiple slender processes. These cells were still tumorigenic in athymic nude mice, but the growth of xenografts was remarkably reduced, especially in the 1.0% DMF-treated group. The expression of glial fibrillary acidic protein was notably increased by DMF (0.5 and 1.0%). Washout experiments revealed that the effects of DMF on cell proliferation and cell cycle distribution were reversible. CONCLUSION: Our results suggest that DMF drove the SHG-44 cells to a more mature phenotype with inhibited growth.     

Gemcitabine cytotoxicity of human malignant glioma cells: modulation by antioxidants, BCL-2 and dexamethasone

Gemcitabine is a novel antimetabolite drug that acts by multiple mechanisms, including inhibition of ribonucleoside diphosphate reductase, of dCMP deaminase and of dCTP incorporation into DNA and RNA. Here, we report that gemcitabine induces cytotoxic and clonogenic death of 12 human malignant glioma cell lines at clinically relevant concentrations around 1 microM. Gemcitabine is thus approximately 100-fold more active than the congener drug, cytarabine. Gemcitabine cytotoxicity of glioma cells does not require wild-type p53 activity: (i) there was no difference in the susceptibility to gemcitabine between cell lines with wild-type p53 and cell lines with mutant or deleted p53; (ii) ectopic expression of a temperature-sensitive p53 protein either at wild-type (32.5 degrees C) or at mutant (38.5 degrees C) conformation had no significant influence on gemcitabine-induced cell death. Gemcitabine cytotoxicity was unaffected by the antioxidants, N-acetylcysteine and phenyl-N-tert-butyl-alpha-phenylnitrone. There was no correlation between the susceptibility to gemcitabine and the endogenous expression of the B cell lymphoma-2 (BCL-2)-family proteins BCL-2, BCL-XL, myeloid cell leukemia-1 (MCL-1), BCL-2-associated X protein (BAX), BCL-2 homologous antagonist/killer (BAK) and BCL-XS. Ectopic expression of BCL-2 moderately attenuated gemcitabine-induced cell death. Similarly, preexposure to the synthetic steroid, dexamethasone, which is commonly used to control cerebral edema in brain tumor patients, reduced gemcitabine cytotoxicity. We conclude that the clinical evaluation of gemcitabine for the adjuvant chemotherapy of malignant glioma is warranted.     

Pharmacokinetic MRI for assessment of malignant glioma response to stereotactic radiotherapy: initial results

The parasites of the red rockfish Sebastes capensis off northern Chile are described quantitatively for the first time and compared with those of congeneric species of the Northern Hemisphere as well as of other Chilean marine fishes. Sixteen species were recorded, including 8 ectoparasites (2 copepods, 2 isopods, 1 turbellarian, and 3 monogeneans) and 8 endoparasites (2 acanthocephalans, 3 digeneans, and 3 nematodes). The ectoparasites Lepeophtheirus chilensis and Caligus cheilodactylus, and the endoparasites Pseudopecoelus sp. and Corynosoma sp. were predominant. Eighty percent of the fishes harbored 3-6 parasite species. Four parasite genera new to the genus Sebastes were found in S. capensis, which also shares several parasite genera with its congeneric species from other geographic areas. However, in contrast to its congeners, S. capensis exhibits a lower helminth species richness, although when all the metazoan fauna is considered the species number and diversity are similar. When compared with other demersal fishes of the Chilean coast, S. capensis shows a high number of species and high parasite abundance. Diphtherostomum sp. and Gnathia sp. are new generic records for the parasite fauna of Chilean coast fishes and the finding of Paramicrocotyle sp., Neobenedenia melleni, and Interniloculus chilensis in this study represents a new geographical record for these parasites.     

Induction of senescence in human malignant glioma cells by p16INK4A

In a previous study, we reported that protein intake at the level of dietary protein allowance for Japanese adults, i.e. 1.08 g/kg per day, was enough for recommended daily exercise. However, whether or not recommended daily exercise increases the protein requirement for young adults has not been examined. In this study, we investigated the effect of recommended daily exercise on the protein requirement under an isoenergetic state by a nitrogen balance method. After an adaptation period of 3 days, 12 healthy college students exercised for 10 days with a non-exercise control period of 10 days before or after the exercise period. They were given a maintenance level of energy and 0.64 g/kg per day of high-quality mixed proteins, estimated as the average protein requirement for adults by the Ministry of Health and Welfare of Japan, throughout the experimental period. They performed treadmill running during the exercise period at about 65% of VO2 max for 25 or 40 min/d, which expended 200 or 300 kcal of extra energy, respectively. Although the exercise increased the dermal nitrogen loss, a compensatory decrease in urinary nitrogen excretion was observed. Consequently, the exercises (200 and 300 kcal/d) did not significantly affect the nitrogen balance. These findings indicate that the recommended amount of daily exercise does not change the protein requirement.     

Clinical response to local delivery of tetracycline in relation to overall and local periodontal conditions

The purpose of this study was to determine the clinical response to local delivery of tetracycline in relation to clinical and microbiological conditions of the other teeth. 4 deep pockets were monitored in 19 subjects with multiple deep periodontal lesions and high counts of P. gingivalis. In 9 patients (LT) only 2 of the selected lesions were treated by placement of tetracycline fibers (Actisite), while the rest of the dentition was left untreated. In the other 10 patients, all teeth were supragingivally scaled and then treated by application of polymeric tetracycline HCl containing fibers, the whole dentition was subject to full mouth scaling and root planing, and the patients rinsed with 0.2% chlorhexidine (FT). A significant reduction in mean PPD was observed in all treated sites after two months. This reduction was maintained over the following 4 months. The magnitude of the effect was significantly greater in the FT group (1.74 mm) than in the LT group (0.88 mm). The mean attachment level changes were similar after 2 months in locally and fully treated subjects. A tendency of relapse was noted for treated sites in LT patients from month 2 to 6. A level of statistical significance was not reached for this effect. Data from measurements recorded at 6 sites around all teeth in the full mouth treated patients were analyzed using multiple linear regression. This analysis showed local changes in PPD and AL were significantly and strongly correlated with the baseline value of the respective parameter at the same site. In addition, more pocket depth reduction was noted if a site was not bleeding on probing at 6 months, if the location of a site was not approximal and if the tooth was not a second molar. Sites located on second molars showed also less AL gain than sites located on other teeth. Smokers showed significantly less reduction in PPD and significantly less AL gain. Furthermore, if subjects had a high % of pockets deeper than 4 mm at baseline they showed significantly less attachment gain.     

Generation of fiber-mutant recombinant adenoviruses for gene therapy of malignant glioma

Recombinant adenovirus (Adv)-mediated gene transduction is a powerful technology for cancer gene therapy. In this article, we report the generation of a fiber-mutant Adv vector, using the Adv genomic DNA-terminal protein complex (DNA-TPC) cotransfection method. First, a fiber-mutant construct in a plasmid carrying the right-side two-thirds of the human adenovirus type 5 (Ad5) genome (pTR) was cotransfected with Ad5 DNA-TPC, yielding the recombinant Adv with the desired fiber mutation. The DNA-TPC from the mutant Adv was then utilized to produce a second-step recombinant Adv with an expression cassette in the place of E1. By this procedure, we generated a fiber mutant, F/K20, that has a linker and a stretch of 20 lysine residues added at the C terminus of the fiber. By using Adv carrying a reporter lacZ gene (AxCAZ2) with either F/K20 or wild-type fiber (F/wt), we examined the transduction efficiency of F/K20-Adv. No significant difference in the transduction efficiency between F/K20 and F/wt-Adv was observed for a human fibroblast line, WI-38, or various tumor cell lines, including melanoma, prostate, esophageal, and pancreatic cancer lines. In clear contrast, F/K20-Adv showed a remarkably enhanced efficiency in genetic transduction of human glioma cells. In all four human glioma lines tested, the multiplicities of infection (MOIs) for transduction of 50% of the population (ED50) were decreased with F/K20-Adv compared with F/wt-Adv: 7-fold for T98G, 14-fold for U251, 9-fold for U373, and 42-fold for U87 cells. Therefore, we attempted to apply F/K20-Adv for gene therapy of malignant glioma. Glioma cells infected with F/K20-Adv carrying genes for interleukin 2 or interleukin 12 produced a high level of each cytokine at a much lower MOI than did cells infected with F/wt-Adv. Infection with F/K20-Adv carrying the wild-type p53 tumor suppressor gene resulted in an enhanced level of p53 protein expression and an increased incidence of F/K20-Adv in transduction efficiency for malignant glioma, providing promising tools for gene therapy.     

A simple method for local delivery of various substances to the rat neuromuscular system

Rabies enzootics in southern Africa are associated with two genetically distinct groups of viruses, thought to be adapted to two different sets of host species. The virus groups are referred to as the canid biotype (infecting carnivores of the family Canidae) and the viverrid biotype (infecting carnivores of the subfamily Viverrinae). Cross- or spillover infections of one biotype into the host range of the other are thought to occur from time to time. However, very little is known about this phenomenon and its role in the epidemiology of rabies in southern Africa. We have investigated spillover by monoclonal antibody and nucleic acid sequence analysis of a wide range of virus isolates. Although the inverse had been documented, this report constitutes the first evidence of spillover of canid biotype viruses into viverrid hosts. Our genetic analysis was focused specifically on the G-L intergenic region of the virus genome, thought to be a remnant or pseudogene and it was indicated that, with respect to this region of the genome, spillover does not influence the phylogeny of virus isolates.     

An experimental model for infiltration of malignant lymphoma to the eye and brain

For troponin T a characteristic biphasic change in the plasma time-concentration curve has been described, especially in patients with early reperfusion after thrombolytic therapy. As troponin T is bound to myofibrillar structures, treatment strategy or treatment outcome could influence the cumulative plasma release of this protein in a different way compared to the cumulative release of free cytoplasmic cardiac enzymes. The present study is the first study comparing the total quantity of troponin T released by the heart during the first 168 hours after acute myocardial infarction, both in patients treated with thrombolytic therapy (n = 16) and in patients not treated with thrombolytic therapy (n = 7). On the basis of clinical symptoms and coronary arteriogram within 24 hours, the patients treated with thrombolytic therapy were divided into two groups, reperfused (n = 9) and non-reperfused (n = 7). In the patients not treated with thrombolytic therapy, absence of spontaneous early reperfusion was judged only from clinical symptoms. Cumulative troponin T release into plasma was compared to the cumulative release of the cytoplasmic cardiac enzymes creatine kinase (EC 2.7.3.2) and hydroxybutyrate dehydrogenase (EC 1.1.1.27). Cumulative release, i. e., infarct size, was calculated using a two-compartment model for circulating proteins. Mean tissue contents, per gram wet weight, of 156 U/g for hydroxybutyrate dehydrogenase, 2.163 U/g for creatine kinase and 234 microg/g for troponin T, were used to express infarct size in gram-equivalents of healthy myocardium per litre plasma (g-eq/l). Release rates were represented by the ratio of cumulative quantities released in 10 hours and 72 hours for creatine kinase and hydroxybutyrate dehydrogenase and in 10 hours and 168 hours for troponin T. CONCLUSIONS: - Plasma time-concentration curves and release rates of troponin T in patients treated with thrombolytic therapy showing reperfusion differ significantly from those of patients not treated with thrombolytic therapy, showing no reperfusion. - Creatine kinase and hydroxybutyrate dehydrogenase release is completed within 72-100 hours in all patients, whereas troponin T release still continues after 168 hours. - Cumulative troponin T release at 168 hours is only a fraction (around 8%) of cumulative cytoplasmic enzyme release and the percentage released is not influenced by the treatment strategy or outcome, i. e., vessel patency. - Although troponin T release is only a fraction of the cumulative enzyme release (infarct size) there is a highly significant correlation between both, independent of the treatment strategy or treatment outcome.     

Phase I trial of O6-benzylguanine for patients undergoing surgery for malignant glioma

PURPOSE: The major mechanism of resistance to alkylnitrosourea therapy is the DNA repair protein O6-alkylguanine-DNA alkyltransferase (AGT), which removes chlorethylation or methylation damage from the O6-position of guanine. O6-benzylguanine (O6-BG) is an AGT substrate that inhibits AGT by suicide inactivation. We conducted a phase I trial to define the presurgical dose required for depletion of tumor AGT activity in patients with malignant glioma. MATERIALS AND METHODS: Patients were to be treated 18 hours before craniotomy with intravenous doses that ranged between 40 and 100 mg/m2 given over 1 hour. Resected tumor was snap-frozen in liquid nitrogen and AGT activity analyzed by high-pressure liquid chromatography (HPLC). Up to 13 patients were treated at a specific dose of O6-BG, with a target end point of > or = 11 of 13 patients with undetectable tumor AGT levels (< 10 fmol/mg protein). RESULTS: Thirty patients with malignant gliomas were enrolled, with 11 of 11 patients treated at 100 mg/m2 O6-BG demonstrating tumor AGT levels less than 10 fmol/mg protein. No toxicity was noted in any patient treated. CONCLUSION: These results indicate that 100 mg/m2 of O6-BG can maintain tumor AGT levels less than 10 fmol/mg protein for at least 18 hours after treatment, a time interval in which bis(2-chloroethyl)nitrosourea (BCNU)-induced chloroethyl adducts are fully converted into interstrand cross-links. A 100-mg/m2 dose of O6-BG will be used in combination with BCNU in another phase I trial designed to determine the maximal-tolerated dose of BCNU.     

Response of human uterine arteries to local anesthetics

The in vitro effects of local anesthetics and norepinephrine upon strips of early gestation and term pregnancy uterine arteries were studied in eight cases. In another case, the effect upon uterine veins was studied with a standard organ bath used to record isometric contractions. Histologic preparations were made to verify the type of vessel studied. An artery obtained from an eight-week gestation did not respond to either lidocaine or mepivacaine. All other arterial specimens (radial and helicoidal strips) responded with slow, rising, strong contractions to diluted concentrations of both of these substances. Likewise they responded with rapid contractions when exposed to norepinephrine. Alpha blockers were unable to prevent the contractions triggered by the local anesthetics. The vein specimens did not respond to local anesthetics but contracted when stimulated by norepinephrine. Based on these observations and after brief review of some hypotheses advanced to explain post-paracervical anesthesia fetal bradycardia, it is postulated that this bradycardia is probably due to uterine artery spasm, causing decreased intervillous space blood flow and fetal hypoxia.     

Ocular-specific chemical delivery systems of betaxolol for safe local treatment of glaucoma

This paper examines the social origins of the rise in adult mortality in Russia and selected Eastern European countries. Three explanations for this trend are considered: (1) Soviet health policy, (2) social stress, and (3) health lifestyles. The socialist states were generally characterized by a persistently poor mortality performance as part of a long-term process of deterioration, with particularly negative outcomes for the life expectancy of middle-aged, male manual workers. Soviet-style health policy was ineffective in dealing with the crisis, and stress per se does not seem to be the primary cause of the rise in mortality. Although more research is needed, the suggestion is made that poor health lifestyles--reflected especially in heavy alcohol consumption, and also in smoking, lack of exercise, and high-fat diets--are the major social determinant of the upturn in deaths.