Live-cell microscopy or fluorescence anisotropy with budded baculoviruses - which way to go with measuring ligand binding to M4 muscarinic receptors?

M4 muscarinic receptor is a G protein-coupled receptor that has been associated with alcohol and cocaine abuse, Alzheimer's disease and schizophrenia which makes it an interesting drug target. For many G protein-coupled receptors, the development of high-affinity fluorescence ligands has expanded the options for high throughput screening of drug candidates and serve as useful tools in fundamental receptor research. So far, the lack of suitable fluorescence ligands has limited studying M4 receptor ligand binding. Here, we explored the possibilities of using fluorescence-based methods for studying binding affinity and kinetics to M4 receptor of both labeled and unlabeled ligands. We used two TAMRA-labeled fluorescence ligands, UR-MK342 and UR-CG072, for assay development. Using budded baculovirus particles as M4 receptor preparation and fluorescence anisotropy method, we determined the affinities and binding kinetics of both fluorescence ligands. The fluorescence ligands could also be used as reported probes for determining binding affinities of a set of unlabeled ligands. Based on these results, we took a step further towards a more natural signaling system and developed a method using live CHO-K1-hM4R cells and automated fluorescence microscopy suitable for routine determination of unlabeled ligand affinities. For quantitative image analysis, we developed random forest and deep learning-based pipelines for cell segmentation. The pipelines were integrated into the user-friendly open-source Aparecium software. All developed assays were suitable for measuring fluorescence ligand saturation binding, association and dissociation kinetics as well as for screening binding affinities of unlabeled ligands.

International Union of Basic and Clinical Pharmacology. Guidelines for GPCR ligand bias

G protein-coupled receptors modulate a plethora of physiological processes and mediate the effects of one-third of FDA-approved drugs. Notably, depending on which ligand has activated a particular receptor, it can engage different intracellular transducers. This paradigm of ligand-dependent ‘biased signaling’ dictates a need to advance beyond the level of receptors to consider the combined ligand-receptor pair in order to understand physiological signaling. Bias signaling also has the potential to improve medicines by reducing adverse effects. However, this is challenged by inconsistent interpretation of results and lack of commonly agreed guidelines. Here, we present recommended terminology and guidelines to conduct, report and quantify bias in a comparable and reproducible fashion. We expect these recommendations will facilitate a common understanding of experiments and findings across basic receptor research and drug discovery, while the area and the analytical methods to measure bias are still evolving, especially in complex cellular, tissue and organismal systems.

International Union of Basic and Clinical Pharmacology. Recommendations for GPCR ligand bias

G protein-coupled receptors modulate a plethora of physiological processes and mediate the effects of one-third of FDA-approved drugs. Notably, depending on which ligand has activated a particular receptor, it can engage different intracellular transducers. This paradigm of ligand-dependent ‘biased signaling’ dictates a need to advance beyond the level of receptors to consider the combined ligand-receptor pair in order to understand physiological signaling. Bias signaling also has the potential to improve medicines by reducing adverse effects. However, this is challenged by inconsistent interpretation of results and lack of commonly agreed guidelines. Here, we present recommended terminology and guidelines to conduct, report and quantify bias in a comparable and reproducible fashion. We expect these recommendations will facilitate a common understanding of experiments and findings across basic receptor research and drug discovery, while the area and the analytical methods to measure bias are still evolving, especially in complex cellular, tissue and organismal systems.

Receptor research for the development of new drug therapies for stimulant addiction

Drug addiction is associated with a range of medical and social problems, not least the negative side effects experienced by those who take drugs. The long-term effects are particularly concerning and can be difficult to treat. Changes in the behaviour of long-term drug users can include impulsivity and irrationality, which can negatively impact on society. Dr Kyoji Okita, Integrative Brain Imaging Center (IBIC), National Center of Neurology and Psychiatry, Japan, is working to shed light on the neurological biochemistry behind drug addiction, as this aspect is little understood. In his work, he is collaborating with experts and he intends to apply the knowledge he and his collaborators uncover to real-world treatments. A particular focus for Okita is the use of positron emission tomography (PET) to explore the dynamics behind psychological and addiction biology. Currently, he is investigating the potential of a drug called Istradefylline as a treatment for methamphetamine use disorder. The drug is currently used as part of the treatment for Parkinson's disease and Okita wants to decipher if administering Istradefylline to people addicted to methamphetamines can help increase their dopamine sensitivity and, in the process, reduce their reliance on the drug, as well as the psychological side effects. If found to be effective, Istradefylline could form a key part of a treatment programme for addiction.

Correction to: A Bibliometric analysis of folate receptor research

An amendment to this paper has been published and can be accessed via the original article.

A Bibliometric analysis of folate receptor research

Background The objective of this study was to conduct a bibliometric analysis of the entire field of folate receptor research. Folate receptor is expressed on a wide variety of cancers and certain immune cells. Methods A Web of Science search was performed on folate receptor or folate binding protein (1969-to June 28, 2019). The following information was examined: publications per year, overall citations, top 10 authors, top 10 institutions, top 10 cited articles, top 10 countries, co-author collaborations and key areas of research. Results In total, 3248 documents for folate receptor or folate binding protein were retrieved for the study years outlined in the methods section search query. The range was 1 per year in 1969 to 264 for the last full year studied (2018). A total of 123,720 citations for the 3248 documents retrieved represented a mean citation rate per article of 38.09 and range of 1667 citations (range 0 to 1667). Researchers in 71 countries authored publications analyzed in this study. The US was the leader in publications and had the highest ranking institution. The top 10 articles have been cited 7270 times during the time frame of this study. The top cited article had an average citation rate of 110 citations per year. Network maps revealed considerable co-authorship among several of the top 10 authors. Conclusion Our study presents several important insights into the features and impact of folate receptor research. To our knowledge, this is the first bibliometric analysis of folate receptor.

A Bibliometric Analysis of Folate Receptor Research

Background The objective of this study is to conduct a comprehensible bibliometric analysis of the entire field of folate receptor research. Methods A Web of Science search was performed on folate receptor or folate binding protein (1969-to June 28, 2019). The following information was examined: publications per year, overall citations, top 10 authors, top 10 institutions, top 10 cited articles, top 10 countries, cc-author collaborations and key areas of research. Results In total, 3248 documents for folate receptor or folate binding protein were retrieved for the study years outlined in the methods section search query. The range was 1 per year in 1969 to 264 for the last full year studied (2018). A total of 123,720 citations for the 3,248 documents retrieved represented a mean citation rate per article of 38.09 and range of 1667 citations (range 0 to 1667). Researchers in 71 countries authored publications analyzed in this study. The US was the leader in publications and had the highest ranking institution. The top 10 articles have been cited 7270 times during the time frame of this study. The top cited article had an average citation rate of 110 citations per year. Network maps revealed considerable co-authorship among several of the top 10 authors. Conclusion Our study presents several important insights into the features and impact of folate receptor research. To our knowledge, this is the first bibliometric analysis of folate receptor.

A Bibliometric Analysis of Folate Receptor Research: A comprehensive bibliometric analysis of the entire field of Folate Receptor Research

Background The objective of this study is to conduct a comprehensible bibliometric analysis of the entire field of folate receptor research. Methods A Web of Science search was performed on folate receptor or folate binding protein (1969-to June 28, 2019). The following information was examined: publications per year, overall citations, top 10 authors, top 10 institutions, top 10 cited articles, top 10 countries, cc-author collaborations and key areas of research. Results In total, 3248 documents for folate receptor or folate binding protein were retrieved for the study years outlined in the methods section search query. The range was 1 per year in 1969 to 264 for the last full year studied (2018). A total of 123,720 citations for the 3,248 documents retrieved represented a mean citation rate per article of 38.09 and range of 1667 citations (range 0 to 1667). Researchers in 71 countries authored publications analyzed in this study. The US was the leader in publications and had the highest ranking institution. The top 10 articles have been cited 7270 times during the time frame of this study. The top cited article had an average citation rate of 110 citations per year. Network maps revealed considerable co-authorship among several of the top 10 authors. Conclusion Our study presents several important insights into the features and impact of folate receptor research. To our knowledge, this is the first bibliometric analysis of folate receptor.