Hyperhomocysteinemia is a risk factor for cardiovascular disease, neurological disorders, and bone abnormalities. The key enzyme in homocysteine metabolism, cystathionine-β-synthase (CBS) is recognized as a target for new homocysteine-lowering therapies including enzyme replacement and gene therapy. Currently, there are no pharmacotherapies available that enhance CBS activity through its allosteric mechanism. The only known allosteric activator of CBS is S-adenosyl-L-methionine (SAM), which is available as a food supplement, but its effectiveness is limited by low membrane permeability and universal involvement in methylation reactions as a substrate. The discovery of CBS activators in high-throughput screening is challenging due to a lack of dedicated assays. Available HTS-compatible activity assays for CBS rely on measuring the product hydrogen sulfide or methanethiol where the signal increases with increased CBS activity. In the case of fluorescence-based assays, it is challenging to discern activators from autofluorescent compounds. In this study, we introduce a homocysteine consumption assay for isolated human CBS (HconCBS) based on the absorbance of Ellman's reagent. This assay leverages a decrease in signal upon CBS activation, with performance parameters exceeding the requirements for high-throughput screening. In a commercial library of 3010 compounds, the HconCBS assay identified 10 hit compounds as more active than SAM, whereas a fluorescence-based assay using 7-azido-4-methylcoumarin (AzMC) identified 141 hits. HconCBS identified 101 compounds with autoabsorbance which did not include hit compounds, while the fluorescence-based assay identified 383 autofluorescent compounds which included all hit compounds. While 4 out of 10 HconCBS hits were confirmed when purchased from a new source, the compounds affected homocysteine rather than CBS. Nevertheless, HconCBS consistently detected the CBS activator seleno-adenosyl-L-methionine (SeAM) added to 4 library plates and re-discovered the same library hits in 3 out of 4 re-screened plates. Taken together, HconCBS was designed to enable the discovery of allosteric CBS activators with greater reliability than fluorescence-based methods. Despite identifying some compounds that acted on homocysteine rather than CBS, the assay consistently identified the CBS activators SAM and SeAM and demonstrated reproducibility across two screening rounds. These findings establish HconCBS as a valuable tool for identifying potential therapeutic candidates for hyperhomocysteinemia, addressing a key gap in the development of CBS-targeted pharmacotherapies.
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News media in The Netherlands show great variety in the extent and ways, in which they realize media accountability online in terms of actor transparency, product transparency and feedback opportunities online. It is suggested that even those news rooms that seem to adhere to transparency and public accountability still need to explore the functionality and application of media accountability instruments (MAI). Both in terms of potentials and pitfalls, news rooms need to consider about what they want to be transparent and in what ways. To the extent that online innovations are visible, traditional news media seem to experiment, as is the case with newsroom blogs or the project of hyper local journalism Dichtbij.nl, part of the Telegraaf Company. Various news media have on-going projects on audience participation, online applications and distribution models. However, since many projects merely aim at finding new applications, processes, platforms and business models, it remains to be seen assess whether projects are indeed reasonably innovative and feasible at the same time. The development of an online and therefore immediate, archived, personalized and interactive context, offers practical and ethical challenges to Dutch journalism. These challenges bring shifts in its role and responsibility to society. It means that changes occur in what journalists are accountable for, as well as ways in how they are accountable. The Dutch media landscape lodges various professional accountability instruments like the press council and both profession-wide and news media specific codes of ethics, but some of these instruments receive only moderate support. Proactive openness is more an exception than the rule and may well be a distinctive indicator for quality journalism. Although news media often acknowledge the importance of media accountability offline and online, they often lack the resources or courage to use them or have different priorities. This ambiguous position may indicate that in relation to media accountability online, Dutch news media are between hope and fear: that it will either improve their relationship with the public and fuel professional quality, or ask too much of resources with too little benefit.
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