Biomedical subjects

J Kreuter

Publications and source records attributed to J Kreuter.

At least 19 recordsLinked to original sources

PubMed2006

Pharmacology of intracellular cytosine-arabinoside-5'-triphosphate in malignant cells of pediatric patients with initial or relapsed leukemia and in normal lymphocytes.

PURPOSE: The prodrug cytosinearabinoside (ara-C) is widely used in the treatment of acute leukemias. The active drug is the intracellular metabolite cytosine-arabinoside-5'-triphosphate (ara-CTP). The purpose of the present study was to investigate the relation between sensitivity and pharmacokinetic parameters Cmax, t1/2 and AUC of ara-CTP. The obtained results were compared to previous studies. EXPERIMENTAL DESIGN: Cmax, t1/2 and AUC of ara-CTP were assessed in leukemic cells of 17 pediatric patients with acute lymphoblastic leukemia (ALL) and in 6 lymphoblastic cell lines and compared with normal lymphocytes of 9 healthy donors by high pressure liquid chromatography (HPLC). The sensitivity of the cells against ara-C was determined by the MTT assay. RESULTS: The intracellular accumulation of ara-CTP was significantly lower in normal lymphocytes (Cmax 47.7-60.9 pmol/10(6) cells) compared to leukemic cell lines (Cmax 11-1128 pmol/10(6) cells) and leukemic cells of our patients (Cmax 85.9-631 pmol/10(6) cells). Similar results were found for the AUC. There was no significant difference between initial and relapsed leukemias in our small cohort. A correlation between sensitivity in terms of IC50 values and the intracellular ara-CTP accumulation was observed in cell lines, but not in leukemic cells and normal lymphocytes from healthy donors. CONCLUSIONS: Pharmacokinetic parameters varied tremendously in leukemic cells in contrast to normal lymphocytes without a difference in sensitivity. It is worthwhile to compare literature data to assess an optimal dosage of ara-C in pediatric patients.

Adolescent↗

PubMed2006

Chemotherapy of brain tumour using doxorubicin bound to surfactant-coated poly(butyl cyanoacrylate) nanoparticles: revisiting the role of surfactants.

Poly(butyl cyanoacrylate) nanoparticles coated with poloxamer 188 (Pluronic) F68) and also, as shown previously, polysorbate 80 (Tween 80) considerably enhance the anti-tumour effect of doxorubicin against an intracranial glioblastoma in rats. The investigation of plasma protein adsorption on the surface of the drug-loaded nanoparticles by two-dimensional electrophoresis (2-D PAGE) revealed that both surfactants, besides other plasma components, induced a considerable adsorption of apolipoprotein A-I (ApoA-I). It is hypothesized that delivery of doxorubicin to the brain by means of nanoparticles may be augmented by the interaction of apolipoprotein A-I that is anchored on the surface of the nanoparticles with the scavenger receptor class B type I (SR-BI) located at the blood-brain barrier. This is the first study that shows a correlation between the adsorption of apolipoprotein A-I on the nanoparticle surface and the delivery of the drug across the blood-brain barrier.

Adsorption↗

PubMed2006

Covalent linkage of apolipoprotein e to albumin nanoparticles strongly enhances drug transport into the brain.

Drug delivery to the brain is becoming more and more important but is severely restricted by the blood-brain barrier. Nanoparticles coated with polysorbates have previously been shown to enable the transport of several drugs across the blood-brain barrier, which under normal circumstances is impermeable to these compounds. Apolipoprotein E was suggested to mediate this drug transport across the blood-brain barrier. In the present study, apolipoprotein E was coupled by chemical methods to nanoparticles made of human serum albumin (HSA-NP). Loperamide, which does not cross the blood-brain barrier but exerts antinociceptive effects after direct injection into the brain, was used as model drug. Apolipoprotein E was chemically bound via linkers to loperamide-loaded HSA-NP. This preparation induced antinociceptive effects in the tail-flick test in ICR mice after i.v. injection. In contrast, nanoparticles linked to apolipoprotein E variants that do not recognize lipoprotein receptors failed to induce these effects. These results indicate that apolipoprotein E attached to the surface of nanoparticles facilitates transport of drugs across the blood-brain barrier, probably after interaction with lipoprotein receptors on the brain capillary endothelial cell membranes.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2006

Dihydroxythiophenes are novel potent inhibitors of human immunodeficiency virus integrase with a diketo acid-like pharmacophore.

We have identified dihydroxythiophenes (DHT) as a novel series of human immunodeficiency virus type 1 (HIV-1) integrase inhibitors with broad antiviral activities against different HIV isolates in vitro. DHT were discovered in a biochemical integrase high-throughput screen searching for inhibitors of the strand transfer reaction of HIV-1 integrase. DHT are selective inhibitors of integrase that do not interfere with virus entry, as shown by the inhibition of a vesicular stomatitis virus G-pseudotyped retroviral system. Moreover, in quantitative real-time PCR experiments, no effect on the synthesis of viral cDNA could be detected but rather an increase in the accumulation of 2-long-terminal-repeat cycles was detected. This suggests that the integration of viral cDNA is blocked. Molecular modeling and the structure activity relationship of DHT demonstrate that our compound fits into a two-metal-binding motif that has been suggested as the essential pharmacophore for diketo acid (DKA)-like strand transfer inhibitors (Grobler et al., Proc. Natl. Acad. Sci. USA 99:6661-6666, 2002.). This notion is supported by the profiling of DHT on retroviral vectors carrying published resistance mutations for DKA-like inhibitors where DHT showed partial cross-resistance. This suggests that DHT bind to a common site in the catalytic center of integrase, albeit with an altered binding mode. Taken together, our findings indicate that DHT are novel selective strand transfer inhibitors of integrase with a pharmacophore homologous to DKA-like inhibitors.

Amino Acid Motifs↗

PubMed2004

Physicochemical characterization of protamine-phosphorothioate nanoparticles.

Protamine-oligonucleotide nanoparticles represent effective colloidal drug carriers for antisense phosphorothioate oligonucleotides (PTO). This study describes improvements in particle preparation and the physicochemical properties of the complexes prepared. The influence of component concentrations, length of the PTO chain and the PTO/protamine weight ratio on particle formation and size, shape and surface charge of the particles were studied in detail. Nanoparticles with diameters of 90-200nm were obtained, using protamine free base (PFB) and phosphorothioate in water. The chemical composition of the nanoparticles was analysed. More than 90% of the PTO could be assembled in the particle matrix using a > or = 1:2 ratio (w/w) of PTO and PFB. About 53-68% of the PFB was incorporated in the particle matrix. The complexes had a zetapotential of -19 up to +32 mV, depending on the PTO/PFB ratio. The kinetics of the assembly of this binary system were observed by dynamic light scattering (DLS) measurements and by sedimentation velocity analysis in the analytical ultracentrifuge (AUC). In addition, scanning electron microscopy (SEM) and atomic force microscopy (AFM) were applied to verify the results of DLS and the ultracentrifuge measurements. According to sedimentation velocity analysis, the particles were only moderately stable in water and unstable in salt solutions. However, the colloidal solution in water could be stabilized by polyethylenglycol 20000 (PEG), which also led to an increase of stability in cell medium.

Chemical Phenomena↗

PubMed2002

Toxicological studies of doxorubicin bound to polysorbate 80-coated poly(butyl cyanoacrylate) nanoparticles in healthy rats and rats with intracranial glioblastoma.

Polysorbate 80-coated poly(butyl cyanoacrylate) nanoparticles (NP) were shown to enable the transport of a number of drugs including the anti-tumour antibiotic doxorubicin (DOX) across the blood-brain barrier (BBB) to the brain after intravenous administration and to considerably reduce the growth of brain tumours in rats. The objective of the present study was to evaluate the acute toxicity of DOX associated with polysorbate 80-coated NP in healthy rats and to establish a therapeutic dose range for this formulation in rats with intracranially implanted 101/8 glioblastoma. Single intravenous administration of empty poly(butyl cyanoacrylate) NP in the dose range 100-400 mg/kg did not cause mortality within the period of observation. NP also did not affect body weight or weight of internal organs. Association of DOX with poly(butyl cyanoacrylate) NP did not produce significant changes of quantitative parameters of acute toxicity of the anti-tumour agent. Likewise, the presence of polysorbate 80 in the formulations was not associated with changes in toxicity compared with free or nanoparticulate drug. Dose regimen of 3x1.5 mg/kg on days 2, 5, 8 after tumour implantation did not cause drug-induced mortality. The results in tumour-bearing rats were similar to those in healthy rats. These results demonstrate that the toxicity of DOX bound to NP was similar or even lower than that of free DOX.

Animals↗

PubMed2002

Antiviral and immunomodulatory activity of the metal chelator ethylenediaminedisuccinic acid against cytomegalovirus in vitro and in vivo.

Antiviral activity of the metal chelator ethylenediaminedisuccinic acid (EDDS) was examined in vitro against human cytomegalovirus (HCMV) wild type strains and strains that are resistant against ganciclovir (GCV) and cidofovir (HPMPC). EDDS inhibited the replication of wild-type as well as GCV- and HPMPC-resistant strains with a 50% effective concentration of 7.4-12 microg/ml. At concentrations of 100 microg/ml EDDS, unlike GCV or HPMPC, suppressed HCMV-induced up-regulation of intercellular adhesion molecule-1 (ICAM-1) and reduced T-cell adhesion to HCMV-infected cells in a monolayer adhesion model. In vitro EDDS inhibited murine cytomegalovirus (MCMV) replication (EC50 8.6 microg/ml) and caused in mice some protection against MCMV induced mortality at a non-toxic dose. Since immunopathological factors may play a significant role in HCMV disease it will be of interest to further study whether EDDS is effective in terms of modulation of inflammatory responses to HCMV infections.

Adjuvants, Immunologic↗

PubMed2002

DNA from bovine papillomavirus type 2 induces warts in a xenograft model.

Bovine papillomavirus (BPV) type 2 DNA was inoculated into calf scrotal skin before grafting onto severe combined immunodeficient mice. Inoculation with viral DNA isolated from a bovine wart induced fibropapillomas that exhibited all the morphological features of a BPV infection in cattle. The production of capsid protein and infectious BPV2 particles was demonstrated by immunohistochemistry and a transformed cell focus assay. In contrast, the injection of molecularly cloned viral genomic DNA led to the induction of papilloma-like lesions in the epidermis, but a fibroma was not formed. In addition, only early genes were expressed and infectious virus particles could not be detected. A restriction enzyme accessibility assay suggested that the methylation status of the molecularly cloned BPV2 DNA was different from that of native viral DNA. A possible correlation between methylation status and tumour phenotype is discussed.

Animals↗

PubMed2001

Nanoparticulate systems for brain delivery of drugs.

The blood--brain barrier (BBB) represents an insurmountable obstacle for a large number of drugs, including antibiotics, antineoplastic agents, and a variety of central nervous system (CNS)-active drugs, especially neuropeptides. One of the possibilities to overcome this barrier is a drug delivery to the brain using nanoparticles. Drugs that have successfully been transported into the brain using this carrier include the hexapeptide dalargin, the dipeptide kytorphin, loperamide, tubocurarine, the NMDA receptor antagonist MRZ 2/576, and doxorubicin. The nanoparticles may be especially helpful for the treatment of the disseminated and very aggressive brain tumors. Intravenously injected doxorubicin-loaded polysorbate 80-coated nanoparticles were able to lead to a 40% cure in rats with intracranially transplanted glioblastomas 101/8. The mechanism of the nanoparticle-mediated transport of the drugs across the blood-brain barrier at present is not fully elucidated. The most likely mechanism is endocytosis by the endothelial cells lining the brain blood capillaries. Nanoparticle-mediated drug transport to the brain depends on the overcoating of the particles with polysorbates, especially polysorbate 80. Overcoating with these materials seems to lead to the adsorption of apolipoprotein E from blood plasma onto the nanoparticle surface. The particles then seem to mimic low density lipoprotein (LDL) particles and could interact with the LDL receptor leading to their uptake by the endothelial cells. After this the drug may be released in these cells and diffuse into the brain interior or the particles may be transcytosed. Other processes such as tight junction modulation or P-glycoprotein (Pgp) inhibition also may occur. Moreover, these mechanisms may run in parallel or may be cooperative thus enabling a drug delivery to the brain.

Animals↗

PubMed2001

Phosphodiester and phosphorothioate oligonucleotide condensation and preparation of antisense nanoparticles.

Protamine is a cationic peptide with a molecular mass of approx. 4000 Da that is able to condense DNA. In the present study it was used to complex antisense oligonucleotides (ODNs) and to form solid particles with initial diameters of 90-150 nm. The reaction was very rapid and occurred by simple mixing of diluted solutions of the polycation with the oligonucleotide. The aggregation was dependent on the oligonucleotide chain length and the protamine/ODN mass ratio. Particle formation required a minimal chain length of nine nucleotides and a mass ratio of 0.5:1. The particle surface charge and the number of particles depended on the mass ratio. With increasing amounts of the peptide, the number of particles and the zeta potential increased. Both negatively and positively charged particles improved the stability of oligonucleotides against DNase I digestion. Above a mass ratio of 2.5:1 no degradation was found. The uptake of unbound rhodamine-labelled ODNs and its complexes with protamine was determined with Vero cells under in vitro cell culture conditions at 37 degrees C and 4 degrees C. At 37 degrees C the cellular uptake increased with increasing mass ratio. The internalized oligonucleotides were localized in the cytoplasm and in the nucleus of the cells. When Vero cells were treated with these samples at 4 degrees C for 4 h, no fluorescence could be detected inside the cells. Therefore, our data indicate an energy dependent endocytotic uptake mechanism. In contrast, spermine and spermidine, which are also known condensation agents, did not aggregate with oligonucleotides into nanoparticles under the same conditions.

Animals↗

PubMed2001

Influence of surface-modifying surfactants on the pharmacokinetic behavior of 14C-poly (methylmethacrylate) nanoparticles in experimental tumor models.

PURPOSE: The aim of this study was to investigate the different pharmacokinetic behavior of surface-modified poly(methylmethacrylate) (PMMA) nanoparticles. METHODS: The particles were 14C-labeled and coated with polysorbate 80, poloxamer 407, and poloxamine 908. Plain particles served as control particles. In vivo studies were performed in three tumor models differing in growth, localization, and origin. Particle suspensions were administered via the tail vein, and at given time animals were killed and organs were dissected for determination of PMMA concentration. RESULTS: For the PMMA nanoparticles coated with poloxamer 407 or poloxamine 908, high and long-lasting concentrations were observed in the melanoma and at a lower level in the breast cancer model. In an intracerebrally growing glioma xenograft, the lowest concentrations that did not differ between the tumor-loaded and tumor-free hemispheres were measured. Organ distribution of the four investigated batches differed significantly. For instance, poloxamer 407- and poloxamine 908-coated particles circulated over a longer period of time in the blood, leading additionally to a higher tumor accumulation. In contrast, plain and polysorbate 80-coated particles accumulated mainly in the liver. The strong expression of vascular endothelial growth factor and Flk-1 in the melanoma correlated with high concentrations of PMMA in this tumor. CONCLUSION: The degree of accumulation of PMMA nanoparticles in tumors depended on the particle surface properties and the specific growth differences of tumors.

Animals↗

PubMed2001

Ablation of pituitary pro-opiomelanocortin (POMC) cells produces alterations in hypothalamic POMC mRNA levels and midbrain mu opioid receptor binding in a conditional transgenic mouse model.

The hypothalamic-pituitary-adrenal (HPA) axis is regulated by stress-related excitatory inputs, and various inhibitory and negative-feedback controls by glucocorticoids and opioids, including pro-opiomelanocortin (POMC)-derived peptides. The role of POMC-derived peptides of pituitary origin in the modulation of brain POMC mRNA expression and opioid receptor binding was investigated using a line of transgenic mice that express a fusion gene composed of the pituitary expression-specific promoter region of the POMC gene driving the herpes simplex viral-1 thymidine kinase (TK). Male adult mice were treated with the antiherpes agent ganciclovir that selectively ablates cells expressing TK. Following treatment, POMC mRNA levels, measured by quantitative solution hybridization/RNase protection assays, were decreased by 48% in the pituitary of the TK+/+ mice, reflecting an expected loss of the pituitary corticotrope POMC cells. This treatment also significantly lowered pituitary beta-endorphin immunoreactivity content and plasma concentrations of corticosterone. In contrast, POMC mRNA levels were increased by 79% in the hypothalamus of the TK+/+ mice with pituitary POMC cell ablation. Binding of [(3)H]DAMGO to mu opioid receptors, as measured by quantitative autoradiography, was significantly reduced in several brain regions including the central grey, median raphe and superficial grey layer of the superior colliculus. These regions are innervated by hypothalamic POMC neurones. No significant differences in binding to either kappa or delta opioid receptors were found in the brain regions studied. These results suggest that POMC-derived peptides of pituitary origin may exert a tonic negative-feedback effect on hypothalamic POMC neurones. In turn, the downregulation of central mu opioid receptors in this model may be mediated through a mechanism related to hypothalamic POMC overexpression.

Amygdala↗

PubMed2001

Xenograft model for identifying chemotherapeutic agents against papillomaviruses.

The report describes the establishment and characterization of a mouse xenograft transplantation model for the study of papillomavirus infection of bovine skin. Calf scrotal skin was inoculated with bovine papillomavirus type 2 before grafting it to the dorsum of severe combined immunodeficient mice. The grafted skin contained epidermis, dermis, and a thin layer of fat. After 5 months the induced warts not only showed histological features of papillomavirus infections but also tested positive for viral DNA and papillomavirus capsid antigen. The formation of infectious virions was demonstrated by inoculation of new transplants with crude extract from the induced warts as well as in a cell culture focus assay. Topical application of bromovinyl-2'-deoxyuridine led to a reduction in viral DNA content in the developing wart. This small-animal xenograft model should be useful for characterizing antiviral compounds and providing an understanding of the regulation of papillomavirus infections.

Animals↗

PubMed2001

Effect of carbomer concentration and degree of neutralization on the mucoadhesive properties of polymer films.

Mucoadhesive drug delivery systems may enable a prolonged and localized drug release at various sites of the gastrointestinal tract. In the present study, the mucoadhesive properties of flexible polymeric films based on PVP or PVA as film-forming polymers were assessed by measuring the detachment force from excised porcine duodenal mucosa using a tensile strength tester. The mucoadhesive films were comprised of an impermeable backing layer of cellulose acetate butyrate. Carbopol 934P, Carbopol 974NF, and Noveon AA1 were incorporated as mucoadhesive excipients in concentrations of 0-22 wt% relative to the dry mass of the mucoadhesive layer and with various degrees of neutralization corresponding to pH 4.8, 5.5, 6.8, or 7.5. Films based on PVP were generally more mucoadhesive than corresponding formulations based on PVA. Maximum adhesion of PVP-films was observed at pH 5.5 and 6.8 depending on the type of the mucoadhesive polymer and its concentration. An optimal mucoadhesive polymer concentration range was found to be between 2 and 10 wt%. Higher polymer concentrations did not further enhance the mucoadhesive properties, and in some cases even decreased mucoadhesion. Film formulations based on PVA demonstrated no satisfactory mucoadhesive strength.

Acrylic Resins↗

PubMed2001

Interaction of poly(butylcyanoacrylate) nanoparticles with the blood-brain barrier in vivo and in vitro.

Poly(butylcyanoacrylate) nanoparticles were produced by emulsion polymerisation and used either uncoated or overcoated with polysorbate 80 (Tween 80). [3H]-dalargin bound to nanoparticles overcoated with polysorbate 80 or in the form of saline solution was injected into mice and the brain concentrations of radioactivity determined. Statistically significant, three-fold higher brain concentrations with the nanoparticle preparations were obtained after 45 minutes, the time of greatest pharmacological response assessed as analgesia in previous experiments. In addition the brain inulin spaces in rats and the uptake of fluoresceine isothiocyanate labelled nanoparticles in immortalised rat cerebral endothelial cells, (RBE4) were measured. The inulin spaces after i.v. injection of polysorbate 80-coated nanoparticles were significantly increased by 1% compared to controls. This is interpreted as indicating that there is no large scale opening of the tight junctions of the brain endothelium by the polysorbate 80-coated nanoparticles. In in vitro experiments endocytic uptake of fluorescent nanoparticles by RBE4 cells was only observed after polysorbate 80-overcoating, not with uncoated particles. These results further support the hypothesis that the mechanism of blood-brain barrier transport of drugs by polysorbate 80-coated nanoparticles is one of endocytosis followed by possible transcytosis. The experiments were conducted in several laboratories as part of an EEC/INTAS collaborative program. For various procedural and regulatory reasons this necessitated the use of both rats and mice as experimental animals. The brain endothelial cell line used for the in vitro studies is the rat RBE4.

Analgesics↗

PubMed2001

[Optimization of "Photosens" pharmakokinetics by biodegradable nanospheres].

The present study is dedicated to investigation of pharmacokinetics of the colloidal delivery system based on polybutylcyanoacrylate nanoparticles for the II generation photosensitizer Photosense. Free or nanoparticle-bound Photosense was injected intravenously in healthy rats in the dose 15 mg/kg. It was shown that pharmacokinetic curve of the free drug was characterized by peak concentration while plasma concentrations of nanoparticulate Photosense were relatively steady. Elimination of nanoparticulate Photosense was more rapid comparing to the free drug. It is noteworthy that nanoparticles did not enhance liver uptake of the drug. Lung level of nanoparticulate drug was found to be lower and spleen uptake was enhanced. More important is the fact that nanoparticles provided two-fold decrease of Photosense skin concentration which is potentially important for decrease of drug-related skin phototoxicity. The above data provide evidence that optimization of Photosense pharmacokinetic parameters could be achieved by the use of nanoparticles.

Animals↗

PubMed2000

Antisense delivery using protamine-oligonucleotide particles.

Protamine, a polycationic peptide (mol. wt 4000-4500), was evaluated as a potential penetration enhancer for phosphodiester antisense oligonucleotides (ODNs). Unique complexes in the form of nanoparticles were spontaneously formed, which we call 'proticles'. The stability of the particles and the ODNs bound into the proticles was examined in foetal calf serum and cell culture medium. FITC-labelled ODNs bound to protamine showed an increased cellular uptake into human histiocytic lymphoma U 937 cells compared to free ODNs. Proticles significantly decreased cellular growth in a cell proliferation assay using ODNs against the c- myc proto-oncogene.

Cell Division↗

PubMed2000

Preparation of avidin-labelled gelatin nanoparticles as carriers for biotinylated peptide nucleic acid (PNA).

The possibility of preparing uniform nanoparticles consisting of proteins such as gelatin followed by covalent linkage of avidin was investigated. Gelatin nanoparticles were prepared by two step desolvation. Functional groups at the surface of the particulate system were quantified with site-specific reagents. The surface of the nanoparticles was thiolated and avidin was covalently attached to the nanoparticles via a bifunctional spacer at high levels. Biotinylated peptide nucleic acid (PNA) was effectively complexed by the avidin-conjugated nanoparticles. Avidin-conjugated protein nanoparticles should prove as potential carrier system for biotinylated drug derivatives in antisense therapy.

Avidin↗