Actively targeted delivery of anticancer drug to tumor cells by redox-responsive star-shaped micelles

In cancer therapy nanocargos based on star-shaped polymer exhibit unique features such as better stability, smaller size distribution and higher drug capacity in comparison to linear polymeric micelles. In this study, we developed a multifunctional star-shaped micellar system by combination of active targeting ability and redox-responsive behavior. The star-shaped micelles with good stability were self-assembled from four-arm poly(ε-caprolactone)-poly(ethylene glycol) copolymer. The redox-responsive behaviors of these micelles triggered by glutathione were evaluated from the changes of micellar size, morphology and molecular weight. In vitro drug release profiles exhibited that in a stimulated normal physiological environment, the redox-responsive star-shaped micelles could maintain good stability, whereas in a reducing and acid environment similar with that of tumor cells, the encapsulated agent was promptly released. In vitro cellular uptake and subcellular localization of these micelles were further studied with confocal laser scanning microscopy and flow cytometry against the human cervical cancer cell line HeLa. In vivo and ex vivo DOX fluorescence imaging displayed that these FA-functionalized star-shaped micelles possessed much better specificity to target solid tumor. Both the qualitative and quantitative results of the antitumor effect in 4T1 tumor-bearing BALB/c mice demonstrated that these redox-responsive star-shaped micelles have a high therapeutic efficiency to artificial solid tumor. Therefore, the multifunctional star-shaped micelles are a potential platform for targeted anticancer drug delivery.     

Thermosensitive hydrogels based on polypeptides for localized and sustained delivery of anticancer drugs

Thermosensitive hydrogels based on poly(γ-ethyl-l-glutamate)-poly(ethylene glycol)-poly(γ-ethyl-l-glutamate) triblock copolymers (PELG-PEG-PELG) were prepared for localized and sustained delivery of anticancer drugs. The polypeptide-based hydrogels showed much lower critical gelation concentration than the traditional polyester-based hydrogels. In vivo biocompatibility studies revealed that the in situ formed gels in the subcutaneous layer last for ∼21 days, and H&E staining study suggested acceptable biocompatibility of our materials in vivo. Then the hydrogels were tried as injectable implants to encapsulate antitumor drug, paclitaxel (PTX), to assess the in situ anti-tumoral activity using liver cancer xenograft model. The results demonstrated that the PTX-incorporated hydrogels could efficiently suppress the tumor growth, and did not result in obvious damage to normal organs. Therefore, the polypeptide-based thermosensitive hydrogels designed in the present study have great potential to serve as an effective platform for localized anti-cancer drug delivery.     

Matrix metalloproteinase 2-sensitive multifunctional polymeric micelles for tumor-specific co-delivery of siRNA and hydrophobic drugs

Co-delivery of hydrophilic siRNA and hydrophobic drugs is one of the major challenges for nanomaterial-based medicine. Here, we present a simple but multifunctional micellar platform constructed by a matrix metalloproteinase 2 (MMP2)-sensitive copolymer (PEG-pp-PEI-PE) via self-assembly for tumor-targeted siRNA and drug co-delivery. The micellar nanocarrier possesses several key features for siRNA and drug delivery, including (i) excellent stability; (ii) efficient siRNA condensation by PEI; (iii) hydrophobic drug solubilization in the lipid “core”; (iv) passive tumor targeting via the enhanced permeability and retention (EPR) effect; (v) tumor targeting triggered by the up-regulated tumoral MMP2; and (vi) enhanced cell internalization after MMP2-activated exposure of the previously hidden PEI. These cooperative functions ensure the improved tumor targetability, enhanced tumor cell internalization, and synergistic antitumor activity of co-loaded siRNA and drug.     

Effects of PEG tethering chain length of vitamin E TPGS with a Herceptin-functionalized nanoparticle formulation for targeted delivery of anticancer drugs

Drug formulation by ligand conjugated nanoparticles of biodegradable polymers has become one of the most important strategies in drug targeting. We have developed in our previous work nanoparticles of a mixture of two vitamin E TPGS based copolymers PLA-TPGS and TPGS-TOOH with the latter for Herceptin conjugation for targeted delivery of anticancer drugs such as docetaxel to the cancer cells of human epidermal growth factor receptor 2 (HER2) overexpression. In this research, we investigated the effects of the PEG chain length in TPGS, which is in fact a PEGylated vitamin E, on the cellular uptake and cytotoxicity of the drug formulated in the Herceptin-conjugated nanoparticles of PLA-TPGS/TPGS-COOH blend (NPs). Such NPs of PEG1000, PEG2000, PEG3350 and PEG5000, i.e. the PEG of molecule weight 1000, 2000, 3350 and 5000, were prepared by the nanoprecipitation method and characterized for their size and size distribution, drug loading, surface morphology, surface charge and surface chemistry as well as in vitro drug release profile, cellular uptake and cytotoxicity. We found among such nanoparticles, those of PEG1000, i.e. of the shortest PEG tethering chain length, could result in the best therapeutic effects, which are 24.1%, 37.3%, 38.1% more efficient in cellular uptake and 68.1%, 90%, 92.6% lower in IC₅₀ (thus higher in cytotoxicity) than the Herceptin-conjugated nanoparticles of PLA-TPGS/TPGS-COOH blend of PEG2000, PEG3350 and PEG5000 respectively in treatment of SK-BR-3 cancer cells which are of high HER2 overexpression. We provided a theoretical explanation from surface mechanics and thermodynamics for endocytosis of nanoparticles.     

Actively-targeted polyion complex micelles stabilized by cholesterol and disulfide cross-linking for systemic delivery of siRNA to solid tumors

For small interfering RNA (siRNA)-based cancer therapies, we report an actively-targeted and stabilized polyion complex micelle designed to improve tumor accumulation and cancer cell uptake of siRNA following systemic administration. Improvement in micelle stability was achieved using two stabilization mechanisms; covalent disulfide cross-linking and non-covalent hydrophobic interactions. The polymer component was designed to provide disulfide cross-linking and cancer cell-targeting cyclic RGD peptide ligands, while cholesterol-modified siRNA (Chol-siRNA) provided additional hydrophobic stabilization to the micelle structure. Dynamic light scattering confirmed formation of nano-sized disulfide cross-linked micelles (<50 nm in diameter) with a narrow size distribution. Improved stability of Chol-siRNA-loaded micelles (Chol-siRNA micelles) was demonstrated by resistance to both the dilution in serum-containing medium and counter polyion exchange with dextran sulfate, compared to control micelles prepared with Chol-free siRNA (Chol-free micelles). Improved stability resulted in prolonged blood circulation time of Chol-siRNA micelles compared to Chol-free micelles. Furthermore, introduction of cRGD ligands onto Chol-siRNA micelles significantly facilitated accumulation of siRNA in a subcutaneous cervical cancer model following systemic administration. Ultimately, systemically administered cRGD/Chol-siRNA micelles exhibited significant gene silencing activity in the tumor, presumably due to their active targeting ability combined with the enhanced stability through both hydrophobic interactions of cholesterol and disulfide cross-linking.     

Poly-DL-lactic acid: Polyethylene glycol block copolymers. The influence of polyethylene glycol on the degradation of poly-DL-lactic acid

ABA block copolymers of polyethylene glycol and poly-DL-lactic acid were prepared by ring-opening polymerization of DL-dilactide with α,ω-dihydroxy polyethylene glycol, _n, 1000 or 2000. The morphology of the resulting copolymers, with PEG : PLA ratios(mol/mol) of 1:2, 1: 3 and 1:4, was characterized by DSC and ESR spectroscopy. The rate of water uptake was biphasic, reflecting the contribution of two processes: rapid diffusion of water into the initially miscible PEG and PLA blocks; then a slower rate of hydration possibly due to phase separation and hydrolytic cleavage of the PLA blocks. The rate of hydrolytic degradation of the block copolymers in DI water at 37°C was measured by two methods: weight loss and colorimetric analysis of the carboxy end group concentration resulting from chain scission of PLA blocks. As a result of phase separation, the rate of scission of PLA blocks in the copolymers was similar to that of the PLA homopolymer. The more rapid onset of weight loss of the copolymers, relative to PLA, is attributed to the greater water solubility of PEG-PLA oligomers and their greater diffusivity in the more highly hydrated copolymers.     

The use of hollow polymer fibers for the delivery of bioactive molecules to the brain

Experimental studies have provided evidence that bioactive molecules can be delivered to the central nervous system through the use of hollow polymer fibers. These implantable fibers can be used for the delivery of peptide solutions or serve as a carrier device for encapsulating tissue preparations and cell suspensions. This commentary will address the use of these polymer fibers as a potential therapeutic strategy for treating neurodegenerative disorders such as Alzheimer's disease.     

最佳适形野边距的决定因素和求取方法

目的：探讨最佳适形野边距(block aperture margin，BAM)的决定因素和求取方法。方法：采用三维治疗计划系统(three-dimensional treatment planning system,3-D TPS)，测算出头部和胸部常用放疗条件下“建议射野边距”(P90／50)；根据“建议射野边距”，计算出采用不同BAM时，6MeV X线三野照射头部靶区的剂量分布及15MeV X线四野照射胸部靶区的剂量分布。确定其中符合临床剂量要求，治疗体积在大小和形状上与计划靶区适形程度最好者所采用的BAM为最佳BAM。结果：头部靶区6MeV X线三野照射的最佳BAM为5-7mm，胸部靶区15MeV X线四野照射的最佳BAM为7-10mm。结论：最佳BAM的决定因素复杂，最终求取需三维剂量分布计算。     

The use of lactoferrin as a ligand for targeting the polyamidoamine-based gene delivery system to the brain

Development of an efficient gene vector is a key-limiting factor of brain gene therapy. In this study, lactoferrin (Lf), for the first time, was investigated as a brain-targeting ligand in the design of polyamidoamine (PAMAM)-based non-viral gene vector to the brain. Using polyethyleneglycol (PEG) as a spacer, PAMAM-PEG-Lf was successfully synthesized. This vector showed a concentration-dependent manner in the uptake in brain capillary endothelial cells (BCECs). The brain uptake of PAMAM-PEG-Lf was 2.2-fold compared to that of PAMAM-PEG-transferrin (Tf) in vivo. The transfection efficiency of PAMAM-PEG-Lf/DNA complex was higher than that of PAMAM-PEG-Tf/DNA complex in vitro and in vivo. The results of frozen sections showed the widespread expression of an exogenous gene in mouse brain via intravenous administration. With a PAMAM/DNA weight ratio of 10:1, the brain gene expression of the PAMAM-PEG-Lf/DNA complex was about 2.3-fold when compared to that of the PAMAM-PEG-Tf/DNA complex. These results provide evidence that PAMAM-PEG-Lf can be exploited as a potential non-viral gene vector targeting to the brain via noninvasive administration. Lf is a promising ligand for the design of gene delivery systems targeting to the brain.     

The use of immunoliposomes for specific delivery of antimicrobial agents to oral bacteria immobilized on polystyrene

Antibacterial immunoliposomes have been prepared using covalently bound antibody, raised to the cell surface of the bacterium Streptococcus oralis (S. oralis), and incorporating the bactericides chlorhexidine and Triclosan?. A regrowth assay, in which the ability of a bacterial biofilm immobilised on polystyrene to grow after exposure to a test solution, was undertaken to study the action of the antibacterial immunoliposomes. The antibacterial anti-oralis immunoliposomes show enhanced growth inhibition of S. oralis, compared to free bactericide, using low bactericide concentrations. For short exposure times to the biofilms, antibacterial anti-oralis immunoliposomes can show several times enhanced growth inhibition of S. oralis compared to free bactericide. Antibacterial anti-oralis immunoliposomes inhibit the growth of S. oralis more than that of other oral bacteria. The extent of growth inhibition by antibacterial anti-oralis immunoliposomes is linearly related to the number of immunoliposomes targeted to the biofilm surface.     

The effect of iron and iron chelators on the in-vitro block to development of the mouse preimplantation embryo: BAT6 a new medium for improved culture of mouse embryos in vitro

The effect of iron and iron chelators on the development of the mouse embryo in vitro from the 1-cell stage to the blastocyst has been investigated. An adverse effect of iron was found. The high affinity iron chelator, desferal, also blocked development, whilst transferrin (whether as apoprotein or saturated with iron), DETAPAC and EDTA promoted development. The addition of transferrin permitted development to the blastocyst stage of embryos from stains normally exhibiting the 2-cell block. Under such circumstances both the rate of embryonic development and the proportion of embryos reaching the blastocyst stage approached levels found in vivo. Based on these results, a new medium, BAT6, is described for the optimal in-vitro culture of mouse embryos.     

A randomized, double blind, placebo-controlled study to investigate the use of conscious sedation in conjunction with paracervical block for reducing pain in termination of first trimester pregnancy by suction evacuation

BACKGROUND: This study evaluated the role of conscious sedation in pain relief during termination of first trimester pregnancy by suction evacuation (SE) under local anaesthesia. METHODS: A hundred women undergoing SE before 12 weeks gestation were randomized by computer using the sealed envelope method to receive placebo (saline) or conscious sedation (2 mg midazolam and 25 microg fentanyl) i.v. 5 min before cervical dilatation. Paracervical block was given to all patients, 2 min later. Pain scores during and after SE, post-operative side-effects and satisfaction level were compared. RESULTS: No statistically significant differences in pain scores were found between the two groups. Post-operative side-effects such as dizziness (P = 0.015) and drowsiness (P < 0.001) were significantly more severe in the conscious sedation group. However, patients in the conscious sedation group reported better satisfaction levels than the control group (P = 0.003). CONCLUSION: The use of conscious sedation significantly improved patient satisfaction during termination of first trimester pregnancy by SE under local anaesthesia, despite a lack of improvement in pain relief and the presence of increased severe dizziness/drowsiness in the post-operative period.     

Antibodies to amino acid 200-239 (p200) of Ro52 as serological markers for the risk of developing congenital heart block

Maternal autoantibodies to the p200-epitope of Ro52 have been suggested to correlate with development of congenital heart block. The aim of the present study was to evaluate the clinical relevance and predictive value of p200-antibodies in high-risk pregnancies. Sera from 515 Finnish, Swedish and American women were included in the study. Sera originated from 202 mothers with an infant affected by second- or third-degree atrioventricular block (AVB), 177 mothers with rheumatic disease having infants with normal heart rate and female blood donors (n = 136). A novel serological assay for Ro52 p200-antibodies with intra- and inter-assay variability of 3% and 3.8% respectively was developed. Mothers of children affected by AVB II-III had significantly higher p200-antibody levels than mothers with rheumatic disease having children with normal heart rate (P < 0.001). In the Swedish cohort, a distinction between foetuses with normal conduction, AVB I, AVB II and III was possible. A significant difference in anti-p200 levels between AVB I and AVB II-III groups compared with foetuses with normal conduction (P < 0.05 and P < 0.01) was observed. Using p200-antibodies as a second step analysis in Ro52-positive pregnancies increased the positive predictive value for foetal cardiac involvement (AVB I, II or III) from 0.39 (0.27-0.51) to 0.53 (0.37-0.68). In conclusion, Ro52 p200-antibodies may occur in women with unaffected children, but levels are significantly higher in mothers of children with congenital heart block and are suggested as a relevant marker in evaluating the risk for foetal AV block.     

The susceptibility of Candida albicans strains to selected anticancer drugs and flucytosine,relevance of the presence of self-splicing intron in 25S rDNA

Background

The presence of intron 25S allows to divide the Candida albicans species into three subclasses (A, B, C). Intronless and intron harboring strains were reported to have different susceptibility to some drugs, for example to flucytosine and bleomycin.

The use of collagen-based scaffolds to simulate prostate cancer bone metastases with potential for evaluating delivery of nanoparticulate gene therapeutics

Prostate cancer bone metastases are a leading cause of cancer-related death in men with current treatments offering only marginally improved rates of survival. Advances in the understanding of the genetic basis of prostate cancer provide the opportunity to develop gene-based medicines capable of treating metastatic disease. The aim of this work was to establish a 3D cell culture model of prostate cancer bone metastasis using collagen-based scaffolds, to characterise this model, and to assess the potential of the model to evaluate delivery of gene therapeutics designed to target bone metastases. Two prostate cancer cell lines (PC3 and LNCaP) were cultured in 2D standard culture and compared to 3D cell growth on three different collagen-based scaffolds (collagen and composites of collagen containing either glycosaminoglycan or nanohydroxyapatite). The 3D model was characterised for cell proliferation, viability and for matrix metalloproteinase (MMP) enzyme and Prostate Specific Antigen (PSA) secretion. Chemosensitivity to docetaxel treatment was assessed in 2D in comparison to 3D. Nanoparticles (NPs) containing siRNA formulated using a modified cyclodextrin were delivered to the cells on the scaffolds and gene silencing was quantified. Both prostate cancer cell lines actively infiltrated and proliferated on the scaffolds. Cell culture in 3D resulted in reduced levels of MMP1 and MMP9 secretion in PC3 cells. In contrast, LNCaP cells grown in 3D secreted elevated levels of PSA, particularly on the scaffold composed of collagen and glycosaminoglycans. Both cell lines grown in 3D displayed increased resistance to docetaxel treatment. The cyclodextrin.siRNA nanoparticles achieved cellular uptake and knocked down the endogenous GAPDH gene in the 3D model. In conclusion, development of a novel 3D cell culture model of prostate cancer bone metastasis has been initiated resulting, for the first time, in the successful delivery of gene therapeutics in a 3D in vitro model. Further enhancement of this model will help elucidate the pathogenesis of prostate cancer and also accelerate the design of effective therapies which can penetrate into the bone microenvironment for prostate cancer therapy.     

Applying the health belief model to the rational use of drugs for hemodialysis patients: A randomized controlled trial

BackgroundNon-rational use of drugs is a common problem among people with chronic disease. The Health Belief Model (HBM) can develop beliefs and behaviors related to rational drug use.ObjectivesTo investigate the effect of HBM-based training on the rational use of drugs in hemodialysis patients.MethodsThis randomized controlled trial was conducted in 132 people receiving hemodialysis treatment. The data were collected using a Demographic Survey and the Rational Use of Drugs Scale (RUDS). Patients in the study groups were provided with HBM-based rational use of drugs training. The first training session took an average of 30–35 min for each patient, and the second (summary) took an average of 15–20 minResultsThe mean RUDS pretest score was 60.29 ± 10.17 in the intervention group and 62.85 ± 9.94 in the control group. The mean RUDS posttest scores were 78.80 ± 8.16 in the intervention group and 63.48 ± 9.77 in the control group. The difference between the pretest scores in these groups was not statistically significant (p > 0.05), whereas the difference between the posttest scores was found to be significant (p < 0.001).ConclusionIt was observed that training based on the HBM increased the RUD scores. Thus, HBM may be recommended for use as a guide for rational drug use training, especially for patients undergoing hemodialysis.Practice implicationsThe HBM can be an effective and cost-efficient strategy for standardized rational drug use training and supporting hemodialysis patients.     

Biodegradable self-assembled nanoparticles of poly (d,l-lactide-co-glycolide)/hyaluronic acid block copolymers for target delivery of docetaxel to breast cancer

To develop biodegradable docetaxel-loaded self-assembled nanoparticles of poly (d,l-lactide-co-glycolide)/hyaluronic acid block copolymers were successfully synthesized. These copolymers could form nanoparticles with small size (<200 nm), an acceptable CMC (∼7.9 mg/L), typical core/shell structure and superior stability in one week. DTX-loaded PLGA_502H-b-HA_5.6k nanoparticles (DTX/SANPs) showed a biphasic release pattern within 120 h, and exhibited enhanced cytotoxicity toward CD44-overexpressing MDA-MB-231 cells. Cellular uptake study indicated that PLGA_502H-b-HA_5.6k nanoparticles (SANPs) were taken up in MDA-MB-231 cells by CD44-mediated endocytosis. Pharmacokinetics study revealed DTX/SANPs could prolong the circulation of DTX in the blood. In vivo studies demonstrated that SANPs exhibited enhanced tumor targeting and antitumor activity with lower systemic toxicity. In conclusion, DTX/SANPs have great potential for targeted chemotherapy for CD44-overexpressing breast cancer.     

The use of local anaesthetic microinjections to identify central pathways: a quantitative evaluation of the time course and extent of the neuronal block

Summary The time course and extent of local anaesthetic blocks within the spinal cord of cats were evaluated. A monopolar stimulation electrode with the tip lowered into the dorsal columns (DC) 1000 m below cord surface was used to activate antidromically DC fibers at the T13 level and evoke cord dorsum potentials at the level of the lumbar spinal cord. The amplitude of the negative deflection, the N-wave, was determined for various stimulation intensities (stimulation-response-function, SRF). Lidocaine (1%) was microinjected in volumes of 0.5 or 1.0 l into the DC from a glass micropipette 1 mm caudal to the stimulation site. Conduction block was characterized by a reversible shift of the SRFs to higher stimulation intensities. The diameter of the blocked area in the transverse plane was evaluated from threshold intensities and was found to be 0.9±0.1 mm 4 to 30 min after the injection of 0.5 l lidocaine and 1.6±0.36 mm 10 to 45 min after the injection of 1.0 l lidocaine. In the sagittal plane, the diameter of the blocked area following 1.0 l lidocaine was found to be up to 2.8 mm. The DC-block was reversible within 92 min following injection of 1.0 l and 69 min after the injection of 0.5 l lidocaine. The application of the present findings for blocks in other CNS structures is discussed.     

The cholesterol recognition/interaction amino acid consensus motif of the influenza A virus M2 protein is not required for virus replication but contributes to virulence

Influenza A virus particles assemble and bud from plasma membrane domains enriched with the viral glycoproteins but only a small fraction of the total M2 protein is incorporated into virus particles when compared to the other viral glycoproteins. A membrane proximal cholesterol recognition/interaction amino acid consensus (CRAC) motif was previously identified in M2 and suggested to play a role in protein function. We investigated the importance of the CRAC motif on virus replication by generating recombinant proteins and viruses containing amino acid substitutions in this motif. Alteration or completion of the M2 CRAC motif in two different virus strains caused no changes in virus replication in vitro. Viruses lacking an M2 CRAC motif had decreased morbidity and mortality in the mouse model of infection, suggesting that this motif is a virulence determinant which may facilitate virus replication in vivo but is not required for basic virus replication in tissue culture.     

The use of micronucleus test for detection of genotoxic damage to the thyroid gland

We studied the possibility of using the micronucleus test in in vivo experiments on the model of rat follicular thyrocytes prestimulated to cell division (hemithyroidectomy). Single administration of N-nitroso-N-methylurea produced a significant dose-dependent effect on micronucleus formation in thyrocytes and polychromatophilic erythrocytes of the bone marrow. The test system allowed us to reveal a cumulative effect of 2-fold and 4-fold treatment with the mitogen in low or subthreshold doses on the thyroid gland. Our results indicate that the micronucleus test is an informative method for the analysis of the effect of genotoxic agents on the thyroid parenchyma. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 141, No. 1, pp. 99–102, January, 2006