Synthesis and Pharmacological Characterization of 2-Aminoethyl Diphenylborinate (2-APB) Derivatives for Inhibition of Store-Operated Calcium Entry (SOCE) in MDA-MB-231 Breast Cancer Cells.

Calcium ions regulate a wide array of physiological functions including cell differentiation, proliferation, muscle contraction, neurotransmission, and fertilization. The endoplasmic reticulum (ER) is the major intracellular Ca2+ store and cellular events that induce ER store depletion (e.g., activation of inositol 1,4,5-triphosphate (IP3) receptors) trigger a refilling process known as store-operated calcium entry (SOCE). It requires the intricate interaction between the Ca2+ sensing stromal interaction molecules (STIM) located in the ER membrane and the channel forming Orai proteins in the plasma membrane (PM). The resulting active STIM/Orai complexes form highly selective Ca2+ channels that facilitate a measurable Ca2+ influx into the cytosol followed by successive refilling of the ER by the sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA). STIM and Orai have attracted significant therapeutic interest, as enhanced SOCE has been associated with several cancers, and mutations in STIM and Orai have been linked to immunodeficiency, autoimmune, and muscular diseases. 2-Aminoethyl diphenylborinate (2-APB) is a known modulator and depending on its concentration can inhibit or enhance SOCE. We have synthesized several novel derivatives of 2-APB, introducing halogen and other small substituents systematically on each position of one of the phenyl rings. Using a fluorometric imaging plate reader (FLIPR) Tetra-based calcium imaging assay we have studied how these structural changes of 2-APB affect the SOCE modulation activity at different compound concentrations in MDA-MB-231 breast cancer cells. We have discovered 2-APB derivatives that block SOCE at low concentrations, at which 2-APB usually enhances SOCE.

Development of chemical tools based on GSK-7975A to study store-operated calcium entry in cells.

Many physiological functions, such as cell differentiation, proliferation, muscle contraction, neurotransmission and fertilisation, are regulated by changes of Ca2+ levels. The major Ca2+ store in cells is the endoplasmic reticulum (ER). Certain cellular processes induce ER store depletion, e.g. by activating IP3 receptors, that in turn induces a store refilling process known as store-operated calcium entry (SOCE). This refilling process entails protein-protein interactions between Ca2+ sensing stromal interaction molecules (STIM) in the ER membrane and Orai proteins in the plasma membrane. Fully assembled STIM/Orai complexes then form highly selective Ca2+ channels called Ca2+ release-activated Ca2+ Channels (CRAC) through which Ca2+ ions flow into the cytosol and subsequently are pumped into the ER by the sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA). Abnormal SOCE has been associated with numerous human diseases and cancers, and therefore key players STIM and Orai have attracted significant therapeutic interest. Several potent experimental and clinical candidate compounds have been developed and have helped to study SOCE in various cell types. We have synthesized multiple novel small-molecule probes based on the known SOCE inhibitor GSK-7975A. Here we present GSK-7975A derivatives, which feature photo-caging, photo-crosslinking, biotin and clickable moieties, and also contain deuterium labels. Evaluation of these GSK-7975A probes using a fluorometric imaging plate reader (FLIPR)-Tetra-based Ca2+ imaging assay showed that most synthetic modifications did not have a detrimental impact on the SOCE inhibitory activity. The photo-caged GSK-7975A was also used in patch-clamp electrophysiology experiments. In summary, we have developed a number of active, GSK-7975A-based molecular probes that have interesting properties and therefore are useful experimental tools to study SOCE in various cells and settings.

Evaluation of fiber and debris release from protective COVID-19 mask textiles and in vitro acute cytotoxicity effects.

Since the start of the current COVID-19 pandemic, for the first time a significant fraction of the world's population cover their respiratory system for an extended period with mostly medical facemasks and textile masks. This new situation raises questions about the extent of mask related debris (fibers and particles) being released and inhaled and possible adverse effects on human health. This study aimed to quantify the debris release from a textile-based facemask in comparison to a surgical mask and a reference cotton textile using both liquid and air extraction. Under liquid extractions, cotton-based textiles released up to 29'452 ± 1'996 fibers g-1 textile while synthetic textiles released up to 1'030 ± 115 fibers g-1 textile. However, when the masks were subjected to air-based extraction scenarios, only a fraction (0.1-1.1%) of this fiber amount was released. Several metals including copper (up to 40.8 ± 0.9 µg g-1) and iron (up to 7.0 ± 0.3 µg g-1) were detected in acid dissolved textiles. Additionally the acute in vitro toxicity of size-fractionated liquid extracts (below and above 0.4 µm) were assessed on human alveolar basal epithelial cells. The current study shows no acute cytotoxicity response for all the analyzed facemasks.