A novel screening system improves genetic correction by internal exon replacement

Trans-splicing is a powerful approach to reprogram the genome. It can be used to replace 5′, 3′ or internal exons. The latter approach has been characterized by low efficiency, as the requirements to promote internal trans-splicing are largely uncharacterized. The trans-splicing process is induced by engineered ‘RNA trans-splicing molecules’ (RTMs), which target a selected pre-mRNA to be reprogrammed via two complementary binding domains. To facilitate the development of more efficient RTMs for therapeutic applications we constructed a novel fluorescence based screening system. We incorporated exon 52 of the COL17A1 gene into a GFP-based cassette system as the target exon. This exon is mutated in many patients with the devastating skin blistering disease epidermolysis bullosa. In a double transfection assay we were able to rapidly identify optimal binding domains targeted to sequences in the surrounding introns 51 and 52. The ability to replace exon 52 was then evaluated in a more endogenous context using a target containing COL17A1 exon 51–intron 51–exon 52–intron 52–exon 53. Two selected RTMs produced significantly higher levels of GFP expression in up to 61% assayed cells. This novel approach allows for rapid identification of efficient RTMs for internal exon replacement

Detection of large molecular weight cytokeratin 8 as carrier protein of CA19–9 in non-small-cell lung cancer cell lines

It has been reported that cytokeratin 8 (CK8) is expressed in all non-small-cell lung cancers (NSCLC). We hypothesized that antigenic changes of CK8 may occur in some NSCLC cell lines. To prove this, Western immunoblot analysis using anti-human CK8 monoclonal antibodies as well as immunohistological staining of CK8 were performed in NSCLC cell lines. As a result, CK8 which had a higher molecular weight than recombinant CK8 was demonstrated in two of eight NSCLC cell lines. In addition, this CK8 contained antigenic epitopes of CA19–9. This CK8 with higher molecular weight, may have played a role in the process of invasion or metastasis of NSCLC. © 1999 Cancer Research Campaig

Improved homology-driven computational validation of protein-protein interactions motivated by the evolutionary gene duplication and divergence hypothesis

<p>Abstract</p> <p>Background</p> <p>Protein-protein interaction (PPI) data sets generated by high-throughput experiments are contaminated by large numbers of erroneous PPIs. Therefore, computational methods for PPI validation are necessary to improve the quality of such data sets. Against the background of the theory that most extant PPIs arose as a consequence of gene duplication, the sensitive search for homologous PPIs, i.e. for PPIs descending from a common ancestral PPI, should be a successful strategy for PPI validation.</p> <p>Results</p> <p>To validate an experimentally observed PPI, we combine FASTA and PSI-BLAST to perform a sensitive sequence-based search for pairs of interacting homologous proteins within a large, integrated PPI database. A novel scoring scheme that incorporates both quality and quantity of all observed matches allows us (1) to consider also tentative paralogs and orthologs in this analysis and (2) to combine search results from more than one homology detection method. ROC curves illustrate the high efficacy of this approach and its improvement over other homology-based validation methods.</p> <p>Conclusion</p> <p>New PPIs are primarily derived from preexisting PPIs and not invented <it>de novo</it>. Thus, the hallmark of true PPIs is the existence of homologous PPIs. The sensitive search for homologous PPIs within a large body of known PPIs is an efficient strategy to separate biologically relevant PPIs from the many spurious PPIs reported by high-throughput experiments.</p

Acute mountain sickness.

Acute mountain sickness (AMS) is a clinical syndrome occurring in otherwise healthy normal individuals who ascend rapidly to high altitude. Symptoms develop over a period ofa few hours or days. The usual symptoms include headache, anorexia, nausea, vomiting, lethargy, unsteadiness of gait, undue dyspnoea on moderate exertion and interrupted sleep. AMS is unrelated to physical fitness, sex or age except that young children over two years of age are unduly susceptible. One of the striking features ofAMS is the wide variation in individual susceptibility which is to some extent consistent. Some subjects never experience symptoms at any altitude while others have repeated attacks on ascending to quite modest altitudes. Rapid ascent to altitudes of 2500 to 3000m will produce symptoms in some subjects while after ascent over 23 days to 5000m most subjects will be affected, some to a marked degree. In general, the more rapid the ascent, the higher the altitude reached and the greater the physical exertion involved, the more severe AMS will be. Ifthe subjects stay at the altitude reached there is a tendency for acclimatization to occur and symptoms to remit over 1-7 days

8th European Conference on Rare Diseases & Orphan Products (ECRD 2016)

MEETING ABSTRACTS: Open Access 8th European Conference on Rare Diseases & Orphan Products (ECRD 2016), Edinburgh, UK. 26-28 May 201

Over-the-Counter Monocyclic Non-Steroidal Anti-Inflammatory Drugs in Environment—Sources, Risks, Biodegradation

Recently, the increased use of monocyclic non-steroidal anti-inflammatory drugs has resulted in their presence in the environment. This may have potential negative effects on living organisms. The biotransformation mechanisms of monocyclic nonsteroidal anti-inflammatory drugs in the human body and in other mammals occur by hydroxylation and conjugation with glycine or glucuronic acid. Biotransformation/biodegradation of monocyclic non-steroidal anti-inflammatory drugs in the environment may be caused by fungal or bacterial microorganisms. Salicylic acid derivatives are degraded by catechol or gentisate as intermediates which are cleaved by dioxygenases. The key intermediate of the paracetamol degradation pathways is hydroquinone. Sometimes, after hydrolysis of this drug, 4- aminophenol is formed, which is a dead-end metabolite. Ibuprofen is metabolized by hydroxylation or activation with CoA, resulting in the formation of isobutylocatechol. The aim of this work is to attempt to summarize the knowledge about environmental risk connected with the presence of over-the-counter antiinflammatory drugs, their sources and the biotransformation and/or biodegradation pathways of these drugs

Epidermolysis bullosa House Austria as a role model for the care of a rare disease

The evolution of the Epidermolysis bullosa (EB) House Austria in Salzburg has demonstrated from its beginning in 2005 in an exceptional way the establishment of an optimized health care for a hitherto neglected group of patients, suffering from a rare but devastating skin disease: Epidermolysis bullosa. Patients with this hereditary mechanobullous skin disease, characterized by a heterogenous clinical course, multisystemic manifestations and increased morbidity and mortality, find in the EB House Austria a multidisciplinary, medical and psychosocial, family-centered support, optimally customized to this condition and individualized to each patient. Its unique structure of four divisions (Outpatient Unit, Research Laboratory, Academy, Clinical Research and Study Center) has set the basis for the delivery of best medical practice and state-of-the-art care as well as the establishment/ performance of high quality and patient centered research and translational medicine. Initially the (ongoing) close collaboration with the powerful patient group and medical research charity “DEBRA Austria” that is dedicated to a multidimensional support of EB patients and their relatives living in Austria and neighboring countries, has enabled the construction of the EB House Austria. The acknowledgement of this institution as a successful model has been officially obtained in 2017 by its designation as a national Center of Expertise for Genodermatoses with special focus on EB and its inclusion into the European Reference Network (ERN) for Rare Skin Disorders in September 2018. Therefore, the history of the EB house is worth reviewing since it can be regarded as a role model for the care of other rare and multisystemic diseases