Dataset Information

A high-throughput integrated microfluidics method enables tyrosine autophosphorylation discovery.

ABSTRACT: Autophosphorylation of receptor and non-receptor tyrosine kinases is a common molecular switch with broad implications for pathogeneses and therapy of cancer and other human diseases. Technologies for large-scale discovery and analysis of autophosphorylation are limited by the inherent difficulty to distinguish between phosphorylation and autophosphorylation in vivo and by the complexity associated with functional assays of receptors kinases in vitro. Here, we report a method for the direct detection and analysis of tyrosine autophosphorylation using integrated microfluidics and freshly synthesized protein arrays. We demonstrate the efficacy of our platform in detecting autophosphorylation activity of soluble and transmembrane tyrosine kinases, and the dependency of in vitro autophosphorylation assays on membranes. Our method, Integrated Microfluidics for Autophosphorylation Discovery (IMAD), is high-throughput, requires low reaction volumes and can be applied in basic and translational research settings. To our knowledge, it is the first demonstration of posttranslational modification analysis of membrane protein arrays.

SUBMITTER: Nevenzal H

PROVIDER: S-EPMC6353932 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

A high-throughput integrated microfluidics method enables tyrosine autophosphorylation discovery.

Nevenzal Hadas H Noach-Hirsh Meirav M Skornik-Bustan Or O Brio Lev L Barbiro-Michaely Efrat E Glick Yair Y Avrahami Dorit D Lahmi Roxane R Tzur Amit A Gerber Doron D

Communications biology 20190130

Autophosphorylation of receptor and non-receptor tyrosine kinases is a common molecular switch with broad implications for pathogeneses and therapy of cancer and other human diseases. Technologies for large-scale discovery and analysis of autophosphorylation are limited by the inherent difficulty to distinguish between phosphorylation and autophosphorylation in vivo and by the complexity associated with functional assays of receptors kinases in vitro. Here, we report a method for the direct dete ...[more]

PMID: 30729180

Similar Datasets

Project description:Polymeric excipients are crucial ingredients in modern pills, increasing the therapeutic bioavailability, safety, stability, and accessibility of lifesaving products to combat diseases in developed and developing countries worldwide. Because many early-pipeline drugs are clinically intractable due to hydrophobicity and crystallinity, new solubilizing excipients can reposition successful and even failed compounds to more effective and inexpensive oral formulations. With assistance from high-throughput controlled polymerization and screening tools, we employed a strategic, molecular evolution approach to systematically modulate designer excipients based on the cyclic imide chemical groups of an important (yet relatively insoluble) drug phenytoin. In these acrylamide- and methacrylate-containing polymers, a synthon approach was employed: one monomer served as a precipitation inhibitor for phenytoin recrystallization, while the comonomer provided hydrophilicity. Systems that maintained drug supersaturation in amorphous solid dispersions were identified with molecular-level understanding of noncovalent interactions using NOESY and DOSY NMR spectroscopy. Poly(N-isopropylacrylamide-co-N,N-dimethylacrylamide) (poly(NIPAm-co-DMA)) at 70 mol % NIPAm exhibited the highest drug solubilization, in which phenytoin associated with inhibiting NIPAm units only with lowered diffusivity in solution. In vitro dissolution tests of select spray-dried dispersions corroborated the screening trends between polymer chemical composition and solubilization performance, where the best NIPAm/DMA polymer elevated the mean area-under-the-dissolution-curve by 21 times its crystalline state at 10 wt % drug loading. When administered to rats for pharmacokinetic evaluation, the same leading poly(NIPAm-co-DMA) formulation tripled the oral bioavailability compared to a leading commercial excipient, HPMCAS, and translated to a remarkable 23-fold improvement over crystalline phenytoin.

Dataset Information

A high-throughput integrated microfluidics method enables tyrosine autophosphorylation discovery.

Publications

A high-throughput integrated microfluidics method enables tyrosine autophosphorylation discovery.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets