The first clinical trial of an implantable microchip-based drug delivery device is discussed. Human parathyroid hormone fragment (1-34) hPTH(1-34) was delivered from the device in vivo. hPTH(1-34) is ...the only approved anabolic osteoporosis treatment, but requires daily injections, making patient compliance an obstacle to effective treatment. Furthermore, a net increase in bone mineral density requires intermittent or pulsatile hPTH(1-34) delivery, a challenge for implantable drug delivery products. The microchip-based devices, containing discrete doses of lyophilized hPTH(1-34), were implanted in eight osteoporotic postmenopausal women for 4 months and wirelessly programmed to release doses from the device once daily for up to 20 days. A computer-based programmer, operating in the Medical Implant Communications Service band, established a bidirectional wireless communication link with the implant to program the dosing schedule and receive implant status confirming proper operation. Each woman subsequently received hPTH(1-34) injections in escalating doses. The pharmacokinetics, safety, tolerability, and bioequivalence of hPTH(1-34) were assessed. Device dosing produced similar pharmacokinetics to multiple injections and had lower coefficients of variation. Bone marker evaluation indicated that daily release from the device increased bone formation. There were no toxic or adverse events due to the device or drug, and patients stated that the implant did not affect quality of life.
Display omitted
Small interfering RNAs (siRNAs) have the potential to be of therapeutic value for many human diseases. So far, however, a serious obstacle to their therapeutic use is represented by ...the absence of appropriate delivery systems able to protect them from degradation and to allow an efficient cellular uptake.
In this work we developed a siRNA delivery system based on inulin (Inu), an abundant and natural polysaccharide. Inu was functionalized via the conjugation with diethylenetriamine (DETA) residues to form the complex Inu-DETA. We studied the size, surface charge and the shape of the Inu-DETA/siRNA complexes; additionally, the cytotoxicity, the silencing efficacy and the cell uptake-mechanisms were studied in the human bronchial epithelial cells (16HBE) and in the hepatocellular carcinoma derived cells (JHH6).
The results presented here indicate that Inu-DETA copolymers can effectively bind siRNAs, are highly cytocompatible and, in JHH6, can effectively deliver functional siRNAs. Optimal delivery is observed using a weight ratio Inu-DETA/siRNA of 4 that corresponds to polyplexes with an average size of 600nm and a slightly negative surface charge. Moreover, the uptake and trafficking mechanisms, mainly based on micropinocytosis and clatrin mediated endocytosis, allow the homogeneous diffusion of siRNA within the cytoplasm of JHH6. Notably, in 16 HBE where the trafficking mechanism (caveolae mediated endocytosis) does not allow an even distribution of siRNA within the cell cytoplasm, no significant siRNA activity is observed.
In conclusion, we developed a novel inulin-based siRNA delivery system able to efficiently release siRNA in JHH6 with negligible cytotoxicity thus opening the way for further testing in more complex in vivo models.
Display omitted
Nucleic acid molecules such as small interfering RNAs (siRNAs) and plasmidic DNAs (pDNAs) have been shown to have the potential to be of therapeutic value in different human diseases. ...Their practical use is however compromised by the lack of appropriate release systems. Delivered as naked molecules, siRNAs/pDNAs are rapidly degraded by extracellular nucleases thus considerably reducing the amount of molecule which can reach the target cells. Additionally, the anionic charge of the phosphate groups present on the siRNAs/pDNAs backbone, disfavors the interaction with the negatively charged surface of the cell membrane.
In this paper we describe the generation of a novel polymer able to deliver both siRNAs and pDNAs. The combined release of these molecules is used in many different experimental settings such as the evaluation of the silencing efficiency of a given siRNA targeted against a given RNA, encoded by the pDNA. The possibility to use the same delivery system is very convenient from the technical point of view and it allows minimizing possible artifacts introduced by the use of different delivery agents for siRNAs and pDNA.
The copolymer described here is based on α,β-poly(N-2-hydroxyethyl)-d,l-aspartamide (PHEA) bearing positively chargeable side oligochains, with diethylamino ethyl methacrylate (DEAEMA) as monomer. Monomer polymerization has been obtained by atom transfer radical polymerization (ATRP), a technique which allows the precise polymerization of the monomer. In addition to the chemical–physical characterization of the polymer, we provide evidences of the polymer ability to delivery both siRNAs and pDNA to cultured cells. Whereas additional investigations are necessary to study the delivery mechanisms of this polyplex, the polymer generated represents a novel and convenient device for the delivery of both siRNAs and pDNA.
The thickening of the vessel wall (intimal hyperplasia) is a pathological process which often follows revascularization approaches such as transluminal angioplasty and artery bypass graft, procedures ...used to re-vascularize stenotic artery. Despite the significant improvements in the treatment of intimal hyperplasia obtained in the last years, the problem has not completely solved. Nucleic acid based-drugs (NABDs) represent an emergent class of molecules with potential therapeutic value for the treatment of intimal hyperplasia. NABDs of interest in the field of intimal hyperplasia are: ribozymes, DNAzymes, antisense oligonucleotides, decoy oligonucleotides, small interfering RNAs and micro interfering RNAs. These molecules can recognize, in a sequencespecific fashion, a target which, depending on the different NABDs, can be represented by a nucleic acid or a protein. Upon binding, NABDs can down-modulate the functions of the target (mRNA/proteins) and thus they are used to impair the functions of disease-causing biological molecules.In spite of the great therapeutic potential demonstrated by NABDs in many experimental model of intima hyperplasia, their practical use is hindered by the necessity to identify optimal delivery systems to the vasculature. In the first part of this review a brief description of the clinical problem related to intima hyperplasia formation after revascularization procedures is reported. In the second part, the attention is focused on the experimental evidences of NABD therapeutic potential in the prevention of intimal hyperplasia. Finally, in the third part, we will describe the strategies developed to optimize NABD delivery to the diseased vessel.
In prostate adenocarcinoma, the dissection of the expression behaviour of the eukaryotic elongation factors (eEF1A1/2) has not yet fully elucidated.
The EEF1A1/A2 expressions were investigated by ...real-time PCR, western blotting (cytoplasmic and cytoskeletal/nuclear-enriched fractions) and immunofluorescence in the androgen-responsive LNCaP and the non-responsive DU-145 and PC-3 cells, displaying a low, moderate and high aggressive phenotype, respectively. Targeted experiments were also conducted in the androgen-responsive 22Rv1, a cell line marking the progression towards androgen-refractory tumour. The non-tumourigenic prostate PZHPV-7 cell line was the control.
Compared with PZHPV-7, cancer cells showed no major variations in EEF1A1 mRNA; eEF1A1 protein increased only in cytoskeletal/nuclear fraction. On the contrary, a significant rise of EEF1A2 mRNA and protein were found, with the highest levels detected in LNCaP. Eukaryotic elongation factor 1A2 immunostaining confirmed the western blotting results. Pilot evaluation in archive prostate tissues showed the presence of EEF1A2 mRNA in near all neoplastic and perineoplastic but not in normal samples or in benign adenoma; in contrast, EEF1A1 mRNA was everywhere detectable.
Eukaryotic elongation factor 1A2 switch-on, observed in cultured tumour prostate cells and in human prostate tumour samples, may represent a feature of prostate cancer; in contrast, a minor involvement is assigned to EEF1A1. These observations suggest to consider EEF1A2 as a marker for prostate cell transformation and/or possibly as a hallmark of cancer progression.
Guar gum (GG) and Guar gum/borax (GGb) hydrogels are studied by means of rheology, Low Field Nuclear Magnetic Resonance (LF NMR) and model drug release tests. These three approaches are used to ...estimate the mesh size (ζ) of the polymeric network. A comparison with similar Scleroglucan systems is carried out. In the case of GGb, the rheological and Low Field NMR estimations of ζ lead to comparable results, while the drug release approach seems to underestimate ζ. Such discrepancy is attributed to the viscous effect of some polymeric chains that, although bound to the network to one end, can freely fluctuate among meshes. The viscous drag exerted by these chains slows down drug diffusion through the polymeric network. A proof for this hypothesis is given by the case of Scleroglucan gel, where the viscous contribution is not so significant and a good agreement between the rheological and release test approaches was found.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Display omitted
This paper focuses on the micro- and nano-topological organization of a hydrogel, constituted by a mixture of bacterial cellulose and acrylic acid, and intended for biomedical ...applications. The presence of acrylic acid promotes the formation of two interpenetrated continuous phases: the primary “pores phase” (PP) containing only water and the secondary “polymeric network phase” (PNP) constituted by the polymeric network swollen by the water. Low field Nuclear Magnetic Resonance (LF NMR), rheology, Scanning Electron Microscopy (SEM) and release tests were used to determine the characteristics of the two phases. In particular, we found that this system is a strong hydrogel constituted by 81% (v/v) of PP phase the remaining part being occupied by the PNP phase. Pores diameters span in the range 10–100μm, the majority of them (85%) falling in the range 30–90μm. The high PP phase tortuosity indicates that big pores are not directly connected to each other, but their connection is realized by a series of interconnected small pores that rend the drug path tortuous. The PNP is characterized by a polymer volume fraction around 0.73 while mesh size is around 3nm.
The theoretical interpretation of the experimental data coming from the techniques panel adopted, yielded to the micro- and nano-organization of our hydrogel.
Despite the involvement of the elongation factors eEF1A (eEF1A1 and eEF1A2) in the development of different cancers no information is available on their possible contribution to the biology of ...hepatocellular carcinoma (HCC). We investigated the expression of both forms of eEF1A in HepG2 and JHH6 cell lines considered to be a good
in vitro model of HCC at different stage of differentiation. Our data indicate that the mRNA amount of eEF1A1 is increased in both cell lines as compared to normal liver tissue, but eEF1A2 mRNA level is markedly increased only in JHH6. Moreover, the less differentiated cell line JHH6 displays higher EEF1A1 and EEF1A2 mRNAs levels and an higher nuclear-enriched/cytoplasm ratio of EEF1A protein compared to the better differentiated HepG2 cell line. Over-expression depends only partially on gene amplification. The more abundant mRNA levels and the higher nuclear-enriched/cytoplasm ratio of eEF1A in JHH6 neither correlate with apoptosis resistance nor with proliferation rate in sub-confluent cells. However, in confluent cells, a clear tendency to maintain JHH6 into the cell cycle was observed.
In conclusion, we document the increased mRNA levels of EEF1A genes in HCC cell lines compared to normal liver. Additionally, we show the increased nuclear-enriched/cytoplasmic protein ratio of eEF1A and the marked raise of the expression of both eEF1A forms in JHH6 compared to HepG2, suggesting the possibility that eEF1A forms might become a relevant markers related to HCC tumor phenotype.
PDF
