ABSTRACT
LIN28 is an RNA-binding protein that regulates the maturation of the let-7 family of microRNAs by bipartite interactions with let-7 precursors through its two distinct cold shock and zinc-knuckle domains. Through inhibition of let-7 biogenesis, LIN28 functions as a pluripotency factor, as well as a driver of tumorigenesis. Here, we report a fluorescence polarization assay to identify small-molecule inhibitors for both domains of LIN28 involved in let-7 interactions. Of 101,017 compounds screened, six inhibit LIN28:let-7 binding and impair LIN28-mediated let-7 oligouridylation. Upon further characterization, we demonstrate that the LIN28 inhibitor TPEN destabilizes the zinc-knuckle domain of LIN28, while LI71 binds the cold shock domain to suppress LIN28's activity against let-7 in leukemia cells and embryonic stem cells. Our results demonstrate selective pharmacologic inhibition of individual domains of LIN28 and provide a foundation for therapeutic inhibition of the let-7 biogenesis pathway in LIN28-driven diseases.
Subject(s)
MicroRNAs/metabolism , RNA Processing, Post-Transcriptional , RNA-Binding Proteins/metabolism , Small Molecule Libraries/pharmacology , Uridine/metabolism , Binding Sites , Cell Line, Tumor , Embryonic Stem Cells/drug effects , Embryonic Stem Cells/metabolism , Fluorescence Polarization , High-Throughput Screening Assays , Humans , MicroRNAs/genetics , Models, Molecular , Niacin/chemistry , Small Molecule Libraries/chemistryABSTRACT
Screening is a methodology widely used in biological and biomedical research. There are numerous visualization methods to validate screening data quality but very few visualization applications capable of hit selection. Here, we present MightyScreen ( mightyscreen.net ), a novel web-based application designed for visual data evaluation as well as visual hit selection. We believe MightyScreen is an intuitive and interactive addition to conventional hit selection methods. We also provide study cases showing how MightyScreen is used to visually explore screening data and make hit selections.
Subject(s)
High-Throughput Screening Assays/methods , Computers , Data Analysis , Internet , Software , User-Computer InterfaceABSTRACT
The presence of organochlorine pesticides (OCPs) in biological and environmental samples has been studied for decades in many countries. Nonetheless, studies in Latin American countries like Colombia have been scarce. Determining the presence of OCPs in breast milk will be of relevance to assess exposures, potential health risks, and for surveillance among Latin American populations. Thirty-two breast-feeding mothers were selected to voluntarily participate in the study. Breast milk samples were analyzed for 10 OCPs (α-, ß-, γ-, δ-HCH, Heptachlor, α-, γ-Chlordane, 4,4' DDT, 4,4' DDE, 4,4' DDD). Milk samples were analyzed using liquid-liquid extraction, followed by sulfuric acid clean-up, and quantified using GC/µECD. Results were confirmed by GC/MS. OCPs concentrations were normalized using fat content. In all but one sample, 4,4' DDE was quantified in concentrations ranging between<17 and 14948 ng g(-1) (ng of OCP per g of lipids), with a mean value of 203 ng g(-1). One woman had 4,4' DDE concentrations that were orders of magnitude above the average concentrations observed worldwide. Concentrations of 4,4' DDE in a second breast milk sample collected in a different time period of lactation from a sub-group of 13 women from the original participants, showed no statistically significant difference with the concentrations found in the first sample. Based on the results obtained from the Persistent Organic Pollutants Global Monitoring Plan report of 2009 of the Stockholm Convention, Colombia ranks fourth from bottom to top in terms of 4,4' DDE average concentrations.
Subject(s)
Environmental Monitoring , Environmental Pollutants/analysis , Hydrocarbons, Chlorinated/analysis , Milk, Human/chemistry , Pesticides/analysis , Adolescent , Adult , Colombia , Environmental Pollutants/chemistry , Environmental Pollutants/isolation & purification , Female , Humans , Hydrocarbons, Chlorinated/chemistry , Hydrocarbons, Chlorinated/isolation & purification , Pesticides/chemistry , Pesticides/isolation & purification , Pregnancy , Young AdultABSTRACT
BACKGROUND: Despite the widespread acceptance of the 23-valent pneumococcal capsular polysaccharide vaccine (PPV), its protective effect continues to be disputed. We describe a novel approach to examine the protective effect of this vaccine. METHODS: We recorded the vaccination status of every patient for whom a culture yielded Streptococcus pneumoniae during a 4.5-year period, comparing rates of prior PPV administration in patients with (1) bacteremic pneumococcal pneumonia, (2) all-invasive pneumococcal disease, (3) nonbacteremic pneumococcal pneumonia, (4) acute exacerbation of chronic bronchitis (AECB) due to S. pneumoniae, and (5) pneumococcal colonization. The principal comparisons were with patients who had bacteremic pneumonia or any invasive pneumococcal disease and those with nonbacteremic pneumococcal pneumonia. We also compared vaccination rates in patients who had nonbacteremic pneumonia with vaccination rates in patients with AECB or pneumococcal colonization. RESULTS: The rate of prior PPV vaccination was lower among patients with bacteremic pneumococcal pneumonia (39.7%) or any invasive pneumococcal disease (38.0%) than among patients with nonbacteremic pneumonia (57.6%), AECB (60.0%), or pneumococcal colonization (57.8%). PPV conferred a 54% protection rate against bacteremic versus nonbacteremic pneumococcal pneumonia. There was no apparent protection against nonbacteremic pneumonia compared, for example, with colonized persons or with those who had AECB. CONCLUSIONS: PPV provides moderate protection against invasive pneumococcal disease but does not protect against nonbacteremic pneumococcal pneumonia. These findings suggest the importance of a continued search for a better pneumococcal vaccine.
