A randomized phase III adjuvant study in high-risk cervical cancer: simultaneous radiochemotherapy with cisplatin (S-RC) versus systemic paclitaxel and carboplatin followed by percutaneous radiation (PC-R): a NOGGO-AGO Intergroup Study.

BACKGROUND: Simultaneous adjuvant platinum-based radiochemotherapy in high-risk cervical cancer (CC) is an established treatment strategy. Sequential paclitaxel (Taxol) and platinum followed by radiotherapy may offer further advantages regarding toxicity. PATIENTS AND METHODS: An open-labeled randomized phase III trial was conducted to compare paclitaxel (175 mg/m(2)) plus carboplatin (AUC5) followed by radiation (50.4 Gy) (experimental arm-A) versus simultaneous radiochemotherapy with cisplatin (40 mg/m(2)/week) (arm-B) in patients with stage IB-IIB CC after surgery. Primary objective was progression-free survival (PFS). RESULTS: Overall, 271 patients were randomized and 263 were eligible for evaluation; 132 in arm-A and 131 in arm-B appropriately balanced. The estimated 2-year PFS was 81.8% [95% confidence interval (CI) 74.4-89.1] in arm-B versus 87.2% (95% CI 81.2-93.3) in arm-A (P = 0.235) and the corresponding 5-year survival rates were 85.8% in arm-A and 78.9% in arm-B (P = 0.25). Hematological grade 3/4 toxicity was higher in arm-B. Alopecia (87.9% versus 4.1%; P < 0.001) and neurotoxicity (65.9% versus 15.6%; P < 0.001) were significantly higher in arm-A. Early treatment termination was significantly more frequent in arm-B than in arm-A (32.1% versus 12.9%; P = 0.001). CONCLUSIONS: Sequential chemotherapy and radiation in high-risk CC could not show any significant survival benefit; however, a different toxicity profile appeared. This sequential regime may constitute an alternative option when contraindications for immediate postoperative radiation are present.

Optical imaging in drug discovery and diagnostic applications.

Optical imaging combines a variety of different diagnostic modalities which have shown great promise for biomedical imaging and as a tool in drug discovery. Several different principles to identify and characterize fundamental processes at the organ, tissue, cellular and molecular level have been exploited, supported by the design of novel imaging agents and biomolecular reporter systems. New optical imaging procedures will contribute considerably to the improvement of the knowledge of disease processes and the more efficient evaluation of drug effects in living laboratory animals. They may also find new diagnostic and therapeutic applications in human clinical practices. These techniques can be used in the field of molecular imaging to allow both visualization and quantification of molecular events associated with disease in a non-invasive and radiation-free manner using relatively simple equipment. The different aspects of imaging instrumentation and methods; the achievements in synthesis and evaluation of novel imaging agents and biochemical reporters; as well as the opportunities of optical imaging in drug delivery, drug discovery and imaging diagnostics will be discussed in this review article.

Epinephrine potentiates calcium mobilization and activation of protein kinases in platelets stimulated by ADP through a mechanism unrelated to phospholipase C.

ADP, added to suspensions of aspirinized 32P-prelabelled washed platelets, induced reversible platelet aggregation, the rapid elevation of cytosolic Ca2+ (maximum at 2 s), 20 kDa myosin light chain phosphorylation (maximum faster than 3 s), 40 kDa protein phosphorylation (maximum at 3-10 s) and phosphatidic acid formation (maximum at 30 s). Prior addition of epinephrine potentiated platelet aggregation, cytosolic Ca2(+)-elevation, 20 and 40 kDa protein phosphorylation evoked by ADP, but it did not enhance phosphatidic acid formation induced by ADP. The potentiating effect of epinephrine on aggregation, cytosolic Ca2(+)-increase and 20 and 40 kDa protein phosphorylation induced by ADP was also observed in the presence of EGTA. Ethylisopropylamiloride, an inhibitor of Na+/H(+)-exchange, did not affect the potentiation of ADP-induced platelet aggregation by epinephrine. We conclude that epinephrine primes platelets to increase Ca2(+)-influx and Ca2(+)-mobilization in response to ADP. The potentiation of cytosolic Ca2(+)-elevation by epinephrine leads to further stimulation of myosin light chain phosphorylation and protein kinase C activation and ultimately to enhanced platelet aggregation. These effects of epinephrine do not seem to take place at the level of phospholipase C.

Epinephrine and the Ca2+ ionophore A23187 synergistically induce platelet aggregation without protein kinase C activation.

Aspirin-pretreated, 32P-prelabeled, washed human platelets resuspended in a buffer containing apyrase and 2% plasma were exposed to epinephrine and the Ca2+ ionophore A23187. Epinephrine potentiated platelet aggregation (not secretion), the production of [32P]phosphatidic acid and myosin light chain phosphorylation induced by A23187. No phosphorylation of the 40 kDa protein, the substrate of protein kinase C, was observed. We conclude that G1-protein activation evoked by epinephrine and Ca2+ mobilization caused by A23187 represents a novel synergism for platelet aggregation and that protein kinase C activation, under these conditions is not needed for platelet aggregation.

Cationic solid-lipid nanoparticles can efficiently bind and transfect plasmid DNA.

The suitability of cationically modified solid-lipid nanoparticles (SLN) as a novel transfection agent was investigated. SLN were produced by hot homogenisation using either Compritol ATO 888 or paraffin as matrix lipid, a mixture of Tween 80 and Span 85 as tenside and either EQ1 (N,N-di-(beta-steaorylethyl)-N,N-dimethylammonium chloride) or cetylpyridinium chloride as charge carrier. The resulting particles were approximately 100 nm in size and showed zeta potentials around +40 mV at pH 7.4. DNA binding was tested by agarose gel electrophoresis. The resulting SLN-DNA complexes were further characterised by AFM and zeta potential measurements. Only the SLN batch SII-13, composed of 4% Compritol, 4% Tween/Span and 1% EQ1, was able to form stable complexes with DNA. Typical complexes were 300 to 800 nm in size. Cytotoxicity and transfection efficiency was tested in vitro on Cos-1 cells. Cationic SLN produced by modification with EQ1 were well tolerated, with LD50 values >3 mg/ml in the LDH release assay and >0.6 mg/ml in the WST-1 assay. Further, SLN-DNA complexes containing between 10 and 200 weight equivalents of SII-13 (matrix lipid) efficiently transfected the galactosidase expression plasmid pCMVbeta in the absence and presence of the endosomolytic agent chloroquine.

Enzymatic degradation of SLN-effect of surfactant and surfactant mixtures.

Solid lipid nanoparticles (SLN) show different degradation velocities by the lipolytic enzyme pancreatic lipase as a function of their composition (lipid matrix, stabilizing surfactant). In combination with pancreatic colipase a degradation assay has been developed for studying the degradation behavior. As a measure to follow the degradation the formed free fatty acids have been analyzed using an enzymatic test. In the studies SLN degradation showed dependencies in relation to the length of the fatty acid chains in the triglycerides and the surfactants used for SLN production. The longer the fatty acid chains in the glycerides, the slower the degradation. The influence of surfactants can be degradation accelerating (e.g. cholic acid sodium salt) or a hindering, degradation slowing down effect due to steric stabilization (e.g. Poloxamer 407). As a second steric stabilizer, Tween 80 has been used and the results showed a less pronounced effect on hindering the degradation process than for Poloxamer 407. This result seems to be correlated to the number of ethyleneoxide chains in the molecule. The longer the ethyleneoxide chains are in the molecule, the more hindered is the anchoring of the lipase/colipase complex and consequently the degradation of the SLN. The result can be used to adjust degradation of SLN and consequently drug release in a controlled way.

The role of plasma proteins in brain targeting: species dependent protein adsorption patterns on brain-specific lipid drug conjugate (LDC) nanoparticles.

The in vivo organ distribution of particulate drug carriers is decisively influenced by the interaction with plasma proteins after i.v. administration. Serum protein adsorption on lipid drug conjugate nanoparticles, a new carrier system for i.v. application, was investigated by 2-dimensional electrophoresis (2-DE). The particles were surface-modified to target them to the brain. To assess the protein adsorption pattern after i.v. injection in mice prior to in vivo studies, the particles were incubated in mouse serum. Incubation in human serum was carried out in parallel to investigate similarities or differences in the protein patterns obtained from men and mice. Distinct differences were found. Particles incubated in human serum showed preferential adsorption of apolipoproteins A-I, A-IV and E. Previously, preferential adsorption of ApoE was reported as one important factor for targeting of Tween(R)80 modified polybutylcyanoacrylate nanoparticles to the brain. Preferential adsorption of ApoA-I and A-IV took place after incubation in mouse serum, adsorption of ApoE could not be clearly confirmed. In vivo localization of the LDC nanoparticles at the blood-brain barrier and diffusion of the marker Nile Red into the brain could be shown by confocal laser-scanning microscopy. Differences of the obtained adsorption patterns are discussed with regard to their relevance for correlations of in vitro and in vivo data obtained from different species.

Formulation of amphotericin B as nanosuspension for oral administration.

Amphotherin B was formulated in a nanosuspension as a new oral drug delivery system for the treatment of experimental visceral leishmaniasis. Amphotericin B (AmB) nanosuspensions were produced by high pressure homogenisation obtaining particles with a PCS diameter of 528 nm. Environmental stability was determined in artificial gastrointestinal fluids at different pH and electrolyte concentrations. In vivo efficacy was determined in a mouse model of visceral leishmaniasis. Following oral administration (5 mg kg(-1)), micronised amphotericin B did not show any curative effect. However, administrations of amphotericin B nanosuspension, reduced liver parasite load by 28.6% compared to untreated controls.

Surfactant, but not the size of solid lipid nanoparticles (SLN) influences viability and cytokine production of macrophages.

After intravenous (i.v.) injection, solid lipid nanoparticles (SLN) interact with mononuclear cells. Murine peritoneal macrophages were incubated with SLN formulations consisting of Dynasan 114 coated with different surfactants. The present study was performed to examine the impact of surfactants, which are important surface defining components of SLN, on viability and cytokine production by macrophages. Cytotoxicity, as assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) test, was strongly influenced by the surfactant used being marked with cetylpyridinium chloride- (CPC-) coated SLN at a concentration of 0.001% and further increased at SLN concentrations of 0.01 and 0.1%. All other SLN formulations -- containing Poloxamine 908 (P908), Poloxamer 407 (P407), Poloxamer 188 (P188), Solutol HS15 (HS15), Tween 80 (T80), Lipoid S75 (S75), sodium cholate (SC), or sodium dodecylsulfate (SDS) -- when used at the same concentrations reduced cell viability only slightly. None of the SLN formulations tested induced cytokine production but a concentration-dependent decrease of IL-6 production was observed, which appeared to be associated with cytotoxic effects. IL-12 and TNF-alpha were detected neither in supernatants of macrophages treated with SLN at any concentration nor in those of untreated cells. In contrast to the type of surfactant, the size of SLN was found neither to affect cytotoxicity of SLN nor to result in induction or digression of cytokine production by macrophages. In conclusion, testing the effects of surfactants on SLN on activity of macrophages is a prerequisite prior to in vivo use of SLN.

Adsorption of PTCDA on a partially KBr covered Ag(111) substrate.

Ordered growth of 3,4,9,10-perylene-tetracarboxylic-dianhydride (PTCDA) on Ag(111) partially covered by one or two monolayers of KBr was investigated by non-contact AFM with molecular resolution. Different adsorption patterns are found on the pure substrate, the one covered by a single monolayer, and the one covered by two monolayers KBr. Simulations with an extended Ising-type model reproduce these experimental patterns very well. The adsorbate-adsorbate and the adsorbate-substrate interaction parameters obtained from the simulation are discussed with respect to the interactions at the Ag(111)|KBr interface. As a result, alkali halide covered metals can be used for tuning the interactions and designing adsorption systems, which opens up new possibilities in the control of self-assembled nanostructures.

[Occurrence of suspicious changes in cervix cytology in women after liver transplantation]. / Zum Auftreten suspekter Veränderungen der Zervixzytologie bei Frauen nach Lebertransplantation.

After transplantation the necessary immunosuppressive therapy predisposes to the development of de novo cancers. From January 1989 to April 1994 we collected PAR smears of 98 women before and repeatedly after liver transplantation. After surgery all women received as immunosuppressive agents either Cyclosporin A or FK 506 as long-term medication. In seven patients (8.5%) who had a normal cervical cytology before transplantation a suspicious PAP smear was found on average 11 months after transplantation. In five cases a histological verification (exploratory excision, coning of the cervix) was made. In two cases we found a CIN III. Possible causes for the higher incidence of dysplasia of the cervix observed in the transplant patients are discussed. Similar to kidney transplant recipients female liver transplant recipients are at higher risk of developing cervical dysplasias and neoplasias. Accelerated development of malignancy seems likely. The influence on the cervical cells of both immunosuppressive drugs Cyclosporin A and FK 506 seems to be of a similar nature. Recommendations for the gynaecological care of female liver transplant recipients treated with immunosuppressive therapy are given.

Formulation and biopharmaceutical issues in the development of drug delivery systems for antiparasitic drugs.

The development of really new antiparasitic drugs to market level is a very rare event. A large number of lead structures have already been screened and discarded, the market is large but poor, and the administrative barriers are increasingly high and costly. Novel antiparasitics must not only be better, they must also be substantially safer than the existing repertoire. There are two major aspects to drug development. One is the strategy of pathogen-specific biochemical intervention, the other the strategy of optimal formulation and application. This review focuses on the latter. In finding and adapting innovative and "intelligent", i.e. parasite- and disease-specific formulations and delivery systems, established but deficient drugs might be optimised, enhancing their efficiency and reducing negative side effects at relatively low cost. Further, many promising new ideas are severely hampered by the low water solubility of the antiparasitic drug. Here as well, some of the innovative drug formulation and delivery systems discussed below might offer highly efficient, while technologically simple, solutions.