Measuring the action of CPP–siRNA conjugates in the lung

Two of the most promising and complex areas in biologics development, either as research tools or potential therapeutics, are cell-penetrating peptides (CPPs) and RNA interference (RNAi) modulators. Consequently, the combined application of these technologies in pursuit of improved delivery profiles for RNAi cargoes presents its own unique challenges. Direct access to the targeted tissue is luxury not always available to the researcher; however, the example of lung presents an excellent opportunity for presenting methodologies relevant to understanding the local impact of CPP-conjugated RNAi modulators. This chapter therefore expands upon updated protocols established on the study of the function of endogenous RNAi and the utility of CPPs in the delivery of short interfering RNA (siRNA) to therapeutically relevant cells in the lung. Methods for sample collection, preservation, and processing are provided with a view to facilitate qualitative and quantitative analysis of delivery. In addition, a protocol for mapping siRNA delivery by in situ hybridisation is provided

Relevance of systems pharmacology in drug discovery

The pharmaceutical industry is in the process of re-inventing its pipeline in an attempt to overcome its increasing phase II and III attrition rates. Here, we describe how systems pharmacology can be used as a risk assessment tool to alleviate this problem before bringing in larger investments. We propose that this translational research tool could provide a valuable, complementary addition to other emerging innovative approaches for target identification and validation in discovery and, ultimately, for aiding clinical trial optimization

Targeting the lung using siRNA and antisense based oligonucleotides

The accessibility to topical administration through inhalation, combined with its large surface area, has led to speculation that the lung might offer an ideal target for the application of oligonucleotide based therapeutics. In this review, we shall critically examine the challenges facing antisense and siRNA based approaches for target validation in vivo and as potential therapeutics. In particular, we shall discuss the antisense and siRNA based approaches in relation to factors such as delivery, distribution, stability, off-target effects, unwanted immune responses and the selection of the optimum mRNA targets

Representações socias em enfermagem: comentários sobre teses e dissertações Social representations: commentary about thesis and dissertations

O ponto de partida deste trabalho se constitui de um levantamento de teses e dissertações desenvolvidas por enfermeiros com o objetivo de verificar como a representação social foi utilizada enquanto referencial teórico nestas pesquisas. Através de uma leitura sistemática dos trabalhos destacou-se as temáticas, os principais resultados e as reflexões que estes propunham para as práticas profissionais de enfermagem. Constatou-se, a partir dos trabalhos analisados que este referencial teórico é utilizado pelos enfermeiros, e que alguns estudos desenvolvidos trazem resultados significativos favorecendo a mudança da visão desses profissionais.<br>The starting point of this work consists on a revision of thesis and dissertations developed by nurses aiming to check how the social representation was used as a theoretical referencial on these researches. Through a sistematic reading of these works some points were dettached, as thematic, main resultas and reflections proposed in order to atain professional practice nursing. It was ascertained from the analysed works that this theoretical referencial is used by nurses, and that some studies developed introduce significative results from these professionals

Genomic complexity of urothelial bladder cancer revealed in urinary cfDNA

Urothelial bladder cancers (UBCs) have heterogeneous clinical characteristics that are mirrored in their diverse genomic profiles. Genomic profiling of UBCs has the potential to benefit routine clinical practice by providing prognostic utility above and beyond conventional clinicopathological factors, and allowing for prediction and surveillance of treatment responses. Urinary DNAs representative of the tumour genome provide a promising resource as a liquid biopsy for non-invasive genomic profiling of UBCs. We compared the genomic profiles of urinary cellular DNA and cell-free DNA (cfDNA) from the urine with matched diagnostic formalin-fixed paraffin-embedded tumour DNAs for 23 well-characterised UBC patients. Our data show urinary DNAs to be highly representative of patient tumours, allowing for detection of recurrent clinically actionable genomic aberrations. Furthermore, a greater aberrant load (indicative of tumour genome) was observed in cfDNA over cellular DNA (P<0.001), resulting in a higher analytical sensitivity for detection of clinically actionable genomic aberrations (P<0.04) when using cfDNA. Thus, cfDNA extracted from the urine of UBC patients has a higher tumour genome burden and allows greater detection of key genomic biomarkers (90%) than cellular DNA from urine (61%) and provides a promising resource for robust whole-genome tumour profiling of UBC with potential to influence clinical decisions without invasive patient interventions.European Journal of Human Genetics advance online publication, 13 January 2016; doi:10.1038/ejhg.2015.281

Osteossíntese de úmero em pombos domésticos (Columba livia) associando-se pinos metálicos e polimetilmetacrilato intramedulares após osteotomia diafisária Humerus osteosynthesis using intramedullary pins and polymethylmethacrylate in domestic pigeons (Columba livia)

Foram utilizadas 28 aves adultas, separadas aleatoriamente em quatro grupos. Os pombos foram anestesiados com isoflurano para a realização da osteotomia diafisária transversa do úmero direito. No grupo I, a osteossíntese foi realizada associando-se dois pinos de Kirschner e polimetilmetacrilato, intramedulares; no grupo II, os pinos de Kirschner foram substituídos por pinos de Schanz; no grupo III, foram utilizados apenas dois pinos de Kirschner; e, no grupo IV, apenas dois pinos de Shanz. Os tempos médios para a consolidação óssea foram de 29±4,04 dias no grupo I; 24±5,29 dias no grupo II; 33±3,74 dias no grupo III; e 32,9±5,21 dias no grupo IV. Foi observada migração dos pinos em 42,9% dos animais do grupo I, em 0% nos do grupo II, em 85,7% nos do grupo III, e em 28,6% nos do grupo IV. Em duas aves dos grupos I, III e IV notou-se incapacidade de voar. Os resultados demonstram que a associação de dois pinos de Schanz e polimetilmetacrilato, ambos intramedulares, é um método efetivo para osteossíntese de úmero em pombos domésticos (Columba livia), proporcionando rápida consolidação óssea e mínimas complicações.<br>Twenty-eight adult domestic pigeons (Columba livia) were randomly divided into four groups of seven birds each. Anesthesia was performed with isoflurane and oxygen, and an osteotomy of the right humerus midshaft was performed with an electric cutter. On the sequence, one of the following treatments was chosen: group I, two Kirschner pins and polymethylmethacrylate intramedullary; group II, two Schanz pins and polymethylmethacrylate intramedullary; group III, two Kirschner pins only; and group IV, two Schanz pins only. The mean time ± standard deviation for fracture healing was 29±4.04 days in group I; 24±5.29 days in group II; 33±3.74 days in group III; 32.9±5.21 days in group IV. Pin migration was observed in 42.9% of the group I animals, 0% of group II, 85.7% of group III, and 28.6% of group IV. Two pigeons of groups I, III, and IV presented flight incapability. The results suggest that two Schanz pins and polymethylmethacrylate intramedullary are an effective method of humeral ostheosynthesis in domestic pigeons (Columba livia), resulting in faster fracture healing with minimal complications

Uptake, efficacy, and systemic distribution of naked, inhaled short interfering RNA (siRNA) and locked nucleic acid (LNA) antisense

Antisense oligonucleotides (ASOs) and small interfering RNA (siRNA) promise specific correction of disease-causing gene expression. Therapeutic implementation, however, has been forestalled by poor delivery to the appropriate tissue, cell type, and subcellular compartment. Topical administration is considered to circumvent these issues. The availability of inhalation devices and unmet medical need in lung disease has focused efforts in this tissue. We report the development of a novel cell sorting method for quantitative, cell type-specific analysis of siRNA, and locked nucleic acid (LNA) ASO uptake and efficacy after intratracheal (i.t.) administration in mice. Through fluorescent dye labeling, we compare the utility of this approach to whole animal and whole tissue analysis, and examine the extent of tissue distribution. We detail rapid systemic access and renal clearance for both therapeutic classes and lack of efficacy at the protein level in lung macrophages, epithelia, or other cell types. We nevertheless observe efficient redirection of i.t. administered phosphorothioate (PS) LNA ASO to the liver and kidney leading to targeted gene knockdown. These data suggest delivery remains a key obstacle to topically administered, naked oligonucleotide efficacy in the lung and introduce inhalation as a potentially viable alternative to injection for antisense administration to the liver and kidneys