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.
Our growing ability to tailor healthcare to the needs of individuals has the potential to transform clinical treatment. However, the measurement of multiple biomarkers to inform clinical decisions ...requires rapid, effective, and affordable diagnostics. Chronic diseases and rapidly evolving pathogens in a larger population have also escalated the need for improved diagnostic capabilities. Current chemical diagnostics are often performed in centralized facilities and are still dependent on multiple steps, molecular labeling, and detailed analysis, causing the result turnaround time to be over hours and days. Rapid diagnostic kits based on lateral flow devices can return results quickly but are only capable of detecting a handful of pathogens or markers. Herein, we present the use of disposable plasmonics with chiroptical nanostructures as a platform for low-cost, label-free optical biosensing with multiplexing and without the need for flow systems often required in current optical biosensors. We showcase the detection of SARS-CoV-2 in complex media as well as an assay for the Norovirus and Zika virus as an early developmental milestone toward high-throughput, single-step diagnostic kits for differential diagnosis of multiple respiratory viruses and any other emerging diagnostic needs. Diagnostics based on this platform, which we term “disposable plasmonics assays,” would be suitable for low-cost screening of multiple pathogens or biomarkers in a near-point-of-care setting.
The structure of the type II DHQase from
Streptomyces coelicolor has been solved and refined to high resolution in complexes with a number of ligands, including dehydroshikimate and a rationally ...designed transition state analogue, 2,3-anhydro-quinic acid. These structures define the active site of the enzyme and the role of key amino acid residues and provide snap shots of the catalytic cycle. The resolution of the flexible lid domain (residues 21–31) shows that the invariant residues Arg23 and Tyr28 close over the active site cleft. The tyrosine acts as the base in the initial proton abstraction, and evidence is provided that the reaction proceeds via an enol intermediate. The active site of the structure of DHQase in complex with the transition state analog also includes molecules of tartrate and glycerol, which provide a basis for further inhibitor design.
Most rheumatic heart disease (RHD) registries are static and centralized, collecting epidemiological and clinical data without providing tools to improve care. We developed a dynamic cloud-based RHD ...case management application with the goal of improving care for patients with RHD in Uganda.
The Active Community Case Management Tool (ACT) was designed to improve community-based case management for chronic disease, with RHD as the first test case. Global and local partner consultation informed selection of critical data fields and prioritization of application functionality. Multiple stages of review and revision culminated in user testing of the application at the Uganda Heart Institute.
Global and local partners provided feedback of the application via survey and interview. The application was well received, and top considerations included avenues to import existing patient data, considering a minimum data entry form, and performing a situation assessment to tailor ACT to the health system setup for each new country. Test users completed a postuse survey. Responses were favorable regarding ease of use, desire to use the application in regular practice, and ability of the application to improve RHD care in Uganda. Concerns included appropriate technical skills and supports and potential disruption of workflow.
Creating the ACT application was a dynamic process, incorporating iterative feedback from local and global partners. Results of the user testing will help refine and optimize the application. The ACT application showed potential for utility and integration into existing care models in Uganda.
The fluoro analogue of the enolate intermediate in the reaction catalyzed by type II dehydroquinases has been prepared from naturally occurring (-)-quinic acid over seven steps and has been shown to ...be the most potent inhibitor reported to date of the type II enzyme from Mycobacterium tuberculosis.
Selective inhibitors of type II dehydroquinase were rationally designed to explore a second binding-pocket in the active-site. The molecular modelling, synthesis, inhibition studies and crystal ...structure determination are described.
Our growing ability to tailor healthcare to the needs of individuals has the potential to transform clinical treatment. However, the measurement of multiple biomarkers to inform clinical decisions ...requires rapid, effective, and affordable diagnostics. Chronic diseases and rapidly evolving pathogens in a larger population have also escalated the need for improved diagnostic capabilities. Current chemical diagnostics are often performed in centralised facilities and are still dependent on multiple steps, molecular labelling, and detailed analysis causing the result turnaround time to be over hours and days. Rapid diagnostic kits based on lateral flow devices can return results quickly but are only capable of detecting a handful of pathogens or markers. Herein we present the use of disposable plasmonics as a platform for low-cost label-free optical biosensing with multiplexing capabilities, eliminating the need for flow systems often required in current optical biosensors. We demonstrate the biosensing capabilities of a system that uses chiroptical measurements as well as plasmonic enhanced fluorescence from metafilms with shuriken shaped nanoindentations. We also perform the detection of SARS-CoV-2 in complex solutions as an early developmental milestone towards high-throughput, single-step, diagnostic kits for differential diagnosis of multiple respiratory viruses and any other emerging diagnostic needs. Diagnostics based on this platform which we term Disposable Plasmonics Assays, would be suitable for low-cost screening of multiple pathogens or biomarkers in a near point of care setting.
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