Ballistic Energy Transport via Long Alkyl Chains: A New Initiation Mechanism.

In an effort to increase the speed and efficiency of ballistic energy transport via oligomeric chains, we performed measurements of the transport in compounds featuring long alkyl chains of up to 37 methylene units. Compounds of the N3-(CH2)n-COOMe type (denoted as aznME) were synthesized with n = 5, 10, 15, 19, 28, 37 and studied using relaxation-assisted two-dimensional infrared spectroscopy. The speed of the ballistic transport, initiated by the N3 tag excitation, increased ca. 3-fold for the longer chains (n = 19-37) compared to the shorter chains, from 14.7 to 48 Å/ps, in line with an earlier prediction (Nawagamuwage et al. 2021, J. Phys. Chem. B, 125, 7546). Modeling, based on solving numerically the Liouville equation, was capable of reproducing the experimental data only if three wavepackets are included, involving CH2 twisting (Tw), wagging (W), and rocking (Ro) chain bands. The approaches for designing molecular systems featuring a higher speed and efficiency of energy transport are discussed.

Structure-Activity Relationships in Supramolecular Hosts Targeting Bacterial Phosphatidylethanolamine (PE) Lipids.

The World Health Organization has described the antimicrobial resistance crisis as one of the top ten global public health threats. New antimicrobial agents that can fight infections caused by antimicrobial resistant pathogens are therefore needed. A potential strategy is the development of small molecules that can selectively interact with bacterial membranes (or membranes of other microbial pathogens), and thereby rapidly kill the bacteria. Here, we report the structure-activity relationship within a group of 22 compounds that were designed to bind the bacterial lipid phosphatidylethanolamine (PE). Liposome-based studies reveal that the lipophilicity of the compounds has the strongest effect on both the affinity and selectivity for PE. The best results were obtained for compounds with logP ≈ 3.75, which showed a 5x to 7x selectivity for bacterial PE lipids over human PC (phosphatidylcholine) lipids. Furthermore, these compounds also showed potent antibacterial activity against the Gram-positive bacterium B. cereus, with minimum inhibitory concentrations (MICs) below 10 µM, a concentration where they showed minimal hemolytic activity against human red blood cells. These results not only show the possibility of PE-binding small molecules to function as antibiotics, but also provide guidelines for the development of compounds targeting other types of biologically relevant membrane lipids.

NASC 2023: Showcasing Diversity in North American Supramolecular Chemistry.

Diversity in supramolecular chemistry can showcase itself in many ways. This includes the diversity of thought and topics covered in research (from fundamental science to applications in biology and materials), as well as the diversity of people (e.g., diversity in race, gender, sexual orientation, country of origin, type of higher education institute, career stage,…). At the North American Supramolecular Chemistry (NASC) meetings, we aim to bring together the best that supramolecular chemistry has to offer in North America, create a sense of community and provide a platform for researchers at any stage of their career to present their work. NASC 2023 was the successful second edition of the NASC meeting series, and this proceedings article highlights the research and impressions of some of the speakers at NASC 2023.

Managing research throughout COVID-19: Lived experiences of supramolecular chemists.

The international Women in Supramolecular Chemistry network believes that taking an area-specific approach effectively supports equality, diversity, and inclusion. Science lacks diversity, and this is intersectional. We share effects of coronavirus disease 2019 (COVID-19) by triangulating findings from an online survey, a collaborative autoethnography, and reflective group research meetings. We show how qualitative research with the community offers insights into challenges and supports individuals, and we demonstrate that research leaders have often taken responsibility for their teams' mental health and well-being at the cost of their own.

Improving Structural Stability and Anticoagulant Activity of a Thrombin Binding Aptamer by Aromatic Modifications.

The thrombin binding aptamer (TBA) is a 15-mer DNA oligonucleotide (5'-GGT TGG TGT GGT TGG-3'), that can form a stable intramolecular antiparallel chair-like G-quadruplex structure. This aptamer shows anticoagulant properties by interacting with one of the two anion binding sites of thrombin, namely the fibrinogen-recognition exosite. Here, we demonstrate that terminal modification of TBA with aromatic fragments such as coumarin, pyrene and perylene diimide (PDI), improves the G-quadruplex stability. The large aromatic surface of these dyes can π-π stack to the G-quadruplex or to each other, thereby stabilizing the aptamer. With respect to the original TBA, monoPDI-functionalized TBA exhibited the most remarkable improvement in melting temperature (ΔTm ≈+18 °C) and displayed enhanced anticoagulant activity.

A supramolecular host for phosphatidylglycerol (PG) lipids with antibacterial activity.

Lipids fulfill a variety of important physiological functions, such as energy storage, providing a hydrophobic barrier, and signal transduction. Despite this plethora of biological roles, lipids are rarely considered a potential target for medical applications. Here, we report a set of neutral small molecules that contain boronic acid and urea functionalities to selectively recognize the bacterial lipid phosphatidylglycerol (PG). The affinity and selectivity was determined using 1H NMR titrations and a liposome-based Alizarin Red S assay. Minimum inhibitory concentrations (MIC) were determined to assess antibacterial activity. The most potent compounds display an association constant with PG in liposomes of at least 5 × 103 M-1, function as antibacterial agents against Gram-positive bacteria (MIC = 12.5-25 µM), and show little hemolytic activity. Mode of action studies suggest that the boronic acids bind to the headgroup of the PG lipids, which leads to a change in membrane fluidity and ultimately causes membrane depolarization and cell death.

Increasing membrane permeability of carboxylic acid-containing drugs using synthetic transmembrane anion transporters.

In this manuscript, we show that small-molecule-based anion transporters can significantly increase the permeability of carboxylic acid containing drugs across lipid bilayers of model vesicles. Due to the drug-like characteristics of the transporters, this finding could not only have implications for drug delivery, but also hints towards potential drug-drug and drug-food interactions.

Bactericidal urea crown ethers target phosphatidylethanolamine membrane lipids.

An increasing number of people are infected with antibiotic-resistant bacteria each year, sometimes with fatal consequences. In this manuscript, we report a novel urea-functionalized crown ether that can bind to the bacterial lipid phosphatidylethanolamine (PE), facilitate PE flip-flop and displays antibacterial activity against the Gram-positive bacterium Bacillus cereus with a minimum inhibitory concentration comparable to that of the known PE-targeting lantibiotic duramycin.

Enhancing the selectivity of optical sensors using synthetic transmembrane ion transporters.

Herein, we introduce a new method to optimize the properties of optical sensors, coined the transporter-liposome-fluorophore (TLF) approach. It is shown that this approach can greatly improve the selectivity of the sensor, increase the dynamic range and maintain the sensitivity of the original fluorophore.

Author Correction: Squaramide-based synthetic chloride transporters activate TFEB but block autophagic flux.

In the version of this article originally submitted, it was stated that the first three authors (Shaoyi_ Than, Yan Wang, Wei Xie) had contributed equally. However, in the published version this information was missing.

Fluorinated synthetic anion carriers: experimental and computational insights into transmembrane chloride transport.

A series of fluorinated tripodal tris-thioureas function as highly active anion transporters across lipid bilayers and cell membranes. Here, we investigate their mechanism of action using anion transport assays in cells and synthetic vesicles and molecular modelling of transporter-lipid interactions. When compared with non-fluorinated analogues, fluorinated compounds demonstrate a different mechanism of membrane transport because the free transporter cannot effectively diffuse through the membrane. As a result, in H+/Cl- cotransport assays, fluorinated transporters require the presence of oleic acid to form anionic oleate complexes for recycling of the transporter, whereas non-fluorinated analogues readily diffuse through the membrane as free transporters and show synergistic transport with the proton transporter gramicidin. Molecular dynamics simulations revealed markedly stronger transporter-lipid interactions for fluorinated compounds compared with non-fluorinated analogues and hence, higher energy barriers for fluorinated compounds to cross the membrane as free transporters. With use of appropriate proton transporters to ensure measurement of the correct rate-limiting steps, the transport rates determined in synthetic vesicle assays show excellent agreement with the anion transport rates determined in cell-based assays. We conclude that integration of computational and experimental methods provides a strategy to optimise transmembrane anion transporter design for biomedical applications.

Squaramide-based synthetic chloride transporters activate TFEB but block autophagic flux.

Cystic fibrosis is a disease caused by defective function of a chloride channel coupled to a blockade of autophagic flux. It has been proposed to use synthetic chloride transporters as pharmacological agents to compensate insufficient chloride fluxes. Here, we report that such chloride anionophores block autophagic flux in spite of the fact that they activate the pro-autophagic transcription factor EB (TFEB) coupled to the inhibition of the autophagy-suppressive mTORC1 kinase activity. Two synthetic chloride transporters (SQ1 and SQ2) caused a partially TFEB-dependent relocation of the autophagic marker LC3 to the Golgi apparatus. Inhibition of TFEB activation using a calcium chelator or calcineurin inhibitors reduced the formation of LC3 puncta in cells, yet did not affect the cytotoxic action of SQ1 and SQ2 that could be observed after prolonged incubation. In conclusion, the squaramide-based synthetic chloride transporters studied in this work (which can also dissipate pH gradients) are probably not appropriate for the treatment of cystic fibrosis yet might be used for other indications such as cancer.

Determinants of Ion-Transporter Cancer Cell Death.

Recently, we showed that synthetic anion transporters DSC4P-1 and SA-3 had activity related to cancer cell death. They were found to increase intracellular chloride and sodium ion concentrations. They were also found to induce apoptosis (DSC4P-1) and both induce apoptosis and inhibit autophagy (SA-3). However, determinants underlying these phenomenological findings were not elucidated. The absence of mechanistic understanding has limited the development of yet-improved systems. Here, we show that three synthetic anion transporters, DSC4P-1, SA-3, and 8FC4P, induce osmotic stress in cells by increasing intracellular ion concentrations. This triggers the generation of reactive oxygen species via a sequential process and promotes caspase-dependent apoptosis. In addition, two of the transporters, SA-3 and 8FC4P, induce autophagy by increasing the cytosolic calcium ion concentration promoted by osmotic stress. However, they eventually inhibit the autophagy process as a result of their ability to disrupt lysosome function through a transporter-mediated decrease in a lysosomal chloride ion concentration and an increase in the lysosomal pH.

Anion carriers as potential treatments for cystic fibrosis: transport in cystic fibrosis cells, and additivity to channel-targeting drugs.

Defective anion transport is a hallmark of the genetic disease cystic fibrosis (CF). One approach to restore anion transport to CF cells utilises alternative pathways for transmembrane anion transport, including artificial anion carriers (anionophores). Here, we screened 22 anionophores for biological activity using fluorescence emission from the halide-sensitive yellow fluorescent protein. Three compounds possessed anion transport activity similar to or greater than that of a bis-(p-nitrophenyl)ureidodecalin previously shown to have promising biological activity. Anion transport by these anionophores was concentration-dependent and persistent. All four anionophores mediated anion transport in CF cells, and their activity was additive to rescue of the predominant disease-causing variant F508del-CFTR using the clinically-licensed drugs lumacaftor and ivacaftor. Toxicity was variable but minimal at the lower end. The results provide further evidence that anionophores, by themselves or together with other treatments that restore anion transport, offer a potential therapeutic strategy for CF.

Full elucidation of the transmembrane anion transport mechanism of squaramides using in silico investigations.

A comprehensive experimental and theoretical investigation of the transmembrane chloride transport promoted by four series of squaramide derivatives, with different degrees of fluorination, number of convergent N-H binding units and conformational shapes, is reported. The experimental chloride binding and transport abilities of these small synthetic molecules in liposomes were rationalised with quantum descriptors and molecular dynamics simulations in POPC bilayers. The tripodal tren-based compounds, with three squaramide binding motifs, have high chloride affinity, isolating the anion from water molecules within the membrane model and preventing its release to the aqueous phase, in agreement with the absence of experimental transport activity. In contrast, the symmetrical mono-squaramides, with moderate chloride binding affinity, are able to bind and release chloride either in the aqueous phase or at the membrane interface level, in line with experimentally observed high transport activity. The PMF profiles associated with the diffusion of these free transporters and their chloride complexes across phospholipid bilayers show that the assisted chloride translocation is thermodynamically favoured.

New Insights into the Anion Transport Selectivity and Mechanism of Tren-based Tris-(thio)ureas.

The anion transport properties of a series of previously reported tren-based anionophores have been revisited using new assays designed to measure anion uniport. This study provides new insights into the transport mechanism and selectivity of this important class of transporters. Specifically, we report the chloride and nitrate transport selectivity of these systems and quantify sulfate transport to determine EC50 values for sulfate transport for the first time. Two new assays were developed to study bicarbonate transport allowing accurate quantification of chloride/bicarbonate exchange.

Dissecting the chloride-nitrate anion transport assay.

A systematic study of chloride vs. nitrate selectivity across six anion transporters has revealed a good correlation between the selectivities of their anion binding and membrane transport properties. This work reveals the limitations of the chloride-nitrate exchange assay and shows how new approaches can be used to measure anion uniport.

A synthetic ion transporter that disrupts autophagy and induces apoptosis by perturbing cellular chloride concentrations.

Perturbations in cellular chloride concentrations can affect cellular pH and autophagy and lead to the onset of apoptosis. With this in mind, synthetic ion transporters have been used to disturb cellular ion homeostasis and thereby induce cell death; however, it is not clear whether synthetic ion transporters can also be used to disrupt autophagy. Here, we show that squaramide-based ion transporters enhance the transport of chloride anions in liposomal models and promote sodium chloride influx into the cytosol. Liposomal and cellular transport activity of the squaramides is shown to correlate with cell death activity, which is attributed to caspase-dependent apoptosis. One ion transporter was also shown to cause additional changes in lysosomal pH, which leads to impairment of lysosomal enzyme activity and disruption of autophagic processes. This disruption is independent of the initiation of apoptosis by the ion transporter. This study provides the first experimental evidence that synthetic ion transporters can disrupt both autophagy and induce apoptosis.