Specific Ligands Research Articles

Uncovering the role of GPR39 in the kidney

G protein-coupled receptors (GPCRs) constitute the largest class of proteins in the mammalian genome. Upon activation by a specific ligand, these receptors initiate downstream cellular responses that regulate various physiological processes. One such receptor under investigation in our lab is the ghrelin family receptor GPR39. Unlike its relatives, GPR39 does not respond to peptide hormones or neuropeptides. Initial studies suggested zinc as the endogenous ligand, but recent research indicates that zinc functions as an allosteric potentiator for another unidentified endogenous ligand. To date, synthetic agonists for GPR39 have revealed functions for GPR39 in the heart, bone, skin, pancreas, and gastrointestinal tract. However, GPR39’s role in renal physiology is currently unknown, despite relatively high kidney expression. To address this gap in knowledge, we first worked to localize GPR39 within the kidney. As a reliable antibody for GPR39 is not available, we utilized a combination of RNAScope (Gpr39) and immunofluorescence (AQP2). We find that GPR39 is expressed in the renal collecting duct, with the highest expression in AQP2-positive cells (principal cells) in the inner medullary collecting duct, and lesser expression in the cortical collecting duct. To determine if GPR39 is expressed apically or basolaterally, we then cloned GPR39 with a C-terminal EGFP tag and observed basolateral targeting of GPR39 in polarized MDCK cells grown on filters. In order to query the function of GPR39 in principal cells, we used murine principal kidney cortical collecting duct cells (mpkCCD). As others have shown, we find that mpkCCD express and traffc AQP2 to the apical plasma membrane only in the presence of the vasopressin analog dDAVP. Furthermore, we observe that treatment of mpkCCD with the GPR39-specific agonist cpd1324 induces internalization of AQP2 into cytoplasmic vesicles, even in the continued presence of dDAVP. Under vehicle conditions (dDAVP + vehicle), 80.9±7.7% (mean±SD) of AQP2 stain colocalized with an apical membrane marker (WGA staining). In contrast, when mpkCCD cells were co-treated ddAVP+cpd1324, only 58.3±6.02% of AQP2 stain colocalized with the apical membrane (p < 0.001, t-test). These results indicate that GPR39 activation may act to antagonize vasopressin-induced AQP2 traffcking in mpkCCD cells. NHLBIT32HL007534 (MK) and American Heart Association Established Investigator Award (JLP). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Tissue Factor Maintains Lung Epithelial Barrier Integrity in Acute Lung Injury Through Cell Adhesion

Background. Acute Respiratory Distress Syndrome (ARDS) is a common cause of acute respiratory failure. While loss of lung epithelial barrier integrity is a pathologic mechanism of ARDS, little is known about the factors that regulate epithelial barrier integrity in the lung. Our previously published mouse studies showed that deletion of alveolar type II epithelial cell Tissue Factor (TF) caused impaired lung barrier integrity in models of Acute Lung Injury (ALI). TF is an integral membrane protein that both initiates the extrinsic coagulation cascade and serves several non-coagulant functions. Notably, TF promotes cell adhesion through interactions with cell surface integrin heterodimers, comprised of individual α and β integrin subunits. As such, we hypothesize that epithelial TF is necessary for maintaining lung barrier integrity and that its overexpression is protective in ALI through increased cell adhesion. Methods. To determine whether supraphysiologic overexpression of TF in the lung epithelium can enhance barrier integrity we created a novel transgenic mouse in which TF was inducibly overexpressed in the lung epithelium (TFEpi+) using a CMV-TetO promoter and crossed with SPC-rtTA59 mice. High alveolar epithelial TF expression compared to wild-type littermates (WT) was confirmed through immunohistochemistry and western blot analysis after one week of doxycycline in drinking water. To induce ALI, mice were intranasally infected with 2000 colony forming units of Klebsiella pneumoniae (KP) or PBS control. At 24-hours post infection, mice were euthanized, lung tissue was collected, and a bronchoalveolar lavage (BAL) was performed. BAL protein, clot time, and leukocyte influx were measured. Lung wet-to-dry weight ratios and bacterial burden were measured. To better understand TF regulation of β1 integrin function, TF-knockout A549 cells were created using CRISPR and cultured on specific β1 integrin ligands (Collagen I, IV, Laminin-511, and Fibronectin). On-cell western blots and crystal violet were used to assess cell surface β1 integrin expression and cell adhesion. Results. TFEpi+ mice showed higher lung TF protein expression (median 38031 [IQR 25033, 57948] vs 1801 [IQR 804.3, 2083] expression normalized to total protein; p=0.0016) compared to WT. TFEpi+ mice infected with KP showed lower BAL protein (mean 248.51 [SD 98.03] vs 366.3 [SD 159.8] μg/ml; p=0.0035) and lung wet-to-dry weight ratios (mean 4.27 [SD 0.37] vs 5.29 [SD 0.75]; p=0.0152) compared to WT. Bacterial burden and BAL inflammatory cell counts were higher, while BAL clot time was lower in infected WT mice compared to control. However, these outcomes did not differ between infected TFEpi+ and WT mice. Loss of TF in alveolar epithelial cells reduced cell surface β1 integrin expression and cell adhesion to Laminin-511. Conclusion: Alveolar epithelial TF overexpression is protective for maintaining lung barrier integrity independent of coagulation and inflammation. Further, cell adhesion studies suggest that this is potentially mediated through increased a6β1 epithelial cell adhesion. AHA Postdoctoral Fellowship, National Institute of Health Grants (HL150783, I01BX002288). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Ligand Profiling as a Diagnostic Tool to Differentiate Patient-Derived α-Synuclein Polymorphs.

Amyloid fibrils are characteristic of many neurodegenerative diseases, including Alzheimer's and Parkinson's diseases. While different diseases may have fibrils formed of the same protein, the supramolecular morphology of these fibrils is disease-specific. Here, a method is reported to distinguish eight morphologically distinct amyloid fibrils based on differences in ligand binding properties. Eight fibrillar polymorphs of α-synuclein (αSyn) were investigated: five generated de novo using recombinant αSyn and three generated using protein misfolding cyclic amplification (PMCA) of recombinant αSyn seeded with brain homogenates from deceased patients diagnosed with Parkinson's disease (PD), multiple system atrophy (MSA), and dementia with Lewy bodies (DLB). Fluorescence binding assays were carried out for each fibril using a toolkit of six different ligands. The fibril samples were separated into five categories based on a binary classification of whether they bound specific ligands or not. Quantitative binding measurements then allowed every fibrillar polymorph to be uniquely identified, and the PMCA fibrils derived from PD, MSA, and DLB patients could be unambiguously distinguished. This approach constitutes a novel and operationally simple method to differentiate amyloid fibril morphologies and to identify disease states using PMCA fibrils obtained by seeding with patient samples.

Ouabain modulates collective cell migration via a Na,K-ATPase activated Ca2+ signal pathway

Collective cell migration is important for many physiological and pathological processes. Ouabain, a cardiotonic steroid and specific ligand to Na,K-ATPase, has been reported to modulate collective cell migration, inhibiting it in cancer cells, and increasing it in normal cells. However, it is not clear what the mechanism is behind this difference in response. It is well-documented that ouabain blocks the pump function of Na,K-ATPase. At low concentrations this activates on the cellular level a Ca2+ mediated signaling chain with multiple downstream effects, including adhesion and migration pathways. At higher concentrations the inhibition of Na+ and K+ transport dominates the effect on the cellular level. We hypothesize that the difference in migration response of cancer and normal cells to ouabain is due to a difference in the Ca2+ signaling response. We further hypothesize that the spread and amplification of Ca2+signals between cells via gap junctions is upregulated by ouabain. To test the hypothesis, we made scratch assays in cultures of MDCK cells and studied migration rate and Ca2+signaling. 10 nM ouabain increases the migration rate, an effect that is inhibited by 2-APB, that blocks the IP3R Ca2+ release from ER. The effect on migration is also blocked by KN-93 that inhibits Ca2+ activated CAMKII. Interestingly, ouabain at 200 nM concentration decreased the migration rate compared to untreated cells. To analyze the strength of intercellular communication we analyzed the spread and synchronicity of spontaneous Ca2+ signals between neighboring cells. For this we used MDCK cells stable transfected with the Ca2+ sensitive fluorescent protein GCamp6f. Cells were treated with vehicle, ouabain 10 nM, or heptanol (blocks gap junctions). Timelapse Ca2+ imaging for 10-30 minutes was done. Machine learning based image analysis with Cellpose was then used to segment cells, followed by cross correlation of the integrated Ca2+signals from neighboring cells. Despite findings from previous phosphoproteomic studies indicating that there should be an upregulation of gap junctions, no conclusive differences in intercellular signaling could be identified for the different treatments. Finally, we made high resolution imaging of cells immunolabeled for vinculin and FAK, two proteins that are involved in migration and adhesion. We found ouabain correlated effects on the localization of vinculin and FAK with a polarized and concentrated localization after high dose ouabain, compared to a diffuse and unpolarized localization after low dose ouabain. Taken together we suggest that low dose ouabain via Na,K-ATPase triggers a Ca2+ signal that destabilize cell adhesions and that this is an important mechanism contributing to the observed upregulation of cell migration. Marta and Gunnar V Philipson's foundation, and VR 2020-05347 (Swedish Research Council). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Development of a synthetic relaxin-3/INSL5 chimeric peptide ligand for NanoBiT complementation binding assays

INSL5 and relaxin-3 are relaxin family peptides with important roles in gut and brain function, respectively. They mediate their actions through the class A GPCRs RXFP4 and RXFP3. RXFP4 has been proposed to be a therapeutic target for colon motility disorders whereas RXFP3 targeting could be effective for neurological conditions such as anxiety. Validation of these targets has been limited by the lack of specific ligands and the availability of robust ligand-binding assays for their development. In this study, we have utilized NanoBiT complementation to develop a SmBiT-conjugated tracer for use with LgBiT-fused RXFP3 and RXFP4. The low affinity between LgBiT:SmBiT should result in a low non-specific luminescence signal and enable the quantification of binding without the tedious separation of non-bound ligands. We used solid-phase peptide synthesis to produce a SmBiT-labelled RXFP3/4 agonist, R3/I5, where SmBiT was conjugated to the B-chain N-terminus via a PEG12 linker. Both SmBiT-R3/I5 and R3/I5 were synthesized and purified in high purity and yield. Stable HEK293T cell lines expressing LgBiT-RXFP3 and LgBiT-RXFP4 were produced and demonstrated normal signaling in response to the synthetic R3/I5 peptide. Binding was first characterized in whole-cell binding kinetic assays validating that the SmBiT-R3/I5 bound to both cell lines with nanomolar affinity with minimal non-specific binding without bound and free SmBiT-R3/I5 separation. We then optimized membrane binding assays, demonstrating easy and robust analysis of both saturation and competition binding from frozen membranes. These assays therefore provide an appropriate rigorous binding assay for the high-throughput analysis of RXFP3 and RXFP4 ligands.

Higher-order G-quadruplex structures and porphyrin ligands: Towards a non-ambiguous relationship

Herein, we evaluated the interaction of the tetracationic porphyrin H2TCPPSpm4 with three distinct DNA G-quadruplex (G4) models, i.e., the tetramolecular G4 d(TGGGGT)4 (Q1), the 5′-5′ stacked G4-dimer [d(CGGAGGT)4]2 (Q2), and a mixture of 5′-5′ stacked G-wires [d(5′-CGGT-3′-3′-GGC-5′)4]n (Qn). The combined data obtained from UV–Vis, CD, fluorescence, PAGE, RLS, AFM, NMR, and HPLC-SEC experiments allowed us to shed light on the binding mode of H2TCPPSpm4 with the three G4 models differing for the type and the number of available G4 ending faces, the length of the G4 units, and the number of stacked G4 building blocks. Specifically, we found that H2TCPPSpm4 interacted with the shortest Q1 as an end-stacking ligand, whereas the groove binding mode was ascertained in the case of the Q2 and Qn G4 models. In the case of the interaction with Q1 and Qn, we found that H2TCPPSpm4 induces the formation of supramolecular aggregates at porphyrin/G4 ratios higher than 2:1, whereas no significant aggregation was observed for the interaction with Q2 up to the 5:1 ratio. These results unambiguously demonstrated the suitability of porphyrins for the development of specific G4 ligands or G4-targeting diagnostic probes, being H2TCPPSpm4 capable to distinguish between different G4s.

Extracellular domains of CARs reprogramme T cell metabolism without antigen stimulation.

Metabolism is an indispensable part of T cell proliferation, activation and exhaustion, yet the metabolism of chimeric antigen receptor (CAR)-T cells remains incompletely understood. CARs are composed of extracellular domains-often single-chain variable fragments (scFvs)-that determine ligand specificity and intracellular domains that trigger signalling following antigen binding. Here, we show that CARs differing only in the scFv variously reprogramme T cell metabolism. Even without exposure to antigens, some CARs increase proliferation and nutrient uptake in T cells. Using stable isotope tracers and mass spectrometry, we observed basal metabolic fluxes through glycolysis doubling and amino acid uptake overtaking anaplerosis in CAR-T cells harbouring a rituximab scFv, unlike other similar anti-CD20 scFvs. Disparate rituximab and 14G2a-based anti-GD2 CAR-T cells are similarly hypermetabolic and channel excess nutrients to nitrogen overflow metabolism. Modest overflow metabolism of CAR-T cells and metabolic compatibility between cancer cells and CAR-T cells are identified as features of efficacious CAR-T cell therapy.

NLRP3 Inflammasome in Acute and Chronic Liver Diseases.

NLRP3 (NOD-, LRR-, and pyrin domain-containing protein 3) is an intracellular complex that upon external stimuli or contact with specific ligands, recruits other components, forming the NLRP3 inflammasome. The NLRP3 inflammasome mainly mediates pyroptosis, a highly inflammatory mode of regulated cell death, as well as IL-18 and IL-1β production. Acute and chronic liver diseases are characterized by a massive influx of pro-inflammatory stimuli enriched in reactive oxygen species (ROS) and damage-associated molecular patterns (DAMPs) that promote the assemblage and activation of the NLRP3 inflammasome. As the major cause of inflammatory cytokine storm, the NLRP3 inflammasome exacerbates liver diseases, even though it might exert protective effects in regards to hepatitis C and B virus infection (HCV and HBV). Here, we summarize the current knowledge concerning NLRP3 inflammasome function in both acute and chronic liver disease and in the post liver transplant setting, focusing on the molecular mechanisms involved in NLRP3 activity.

Ethylenediamine-functionalized metal-organic frameworks for applications of electrochemical supercapacitors as an additive

In this study, the impact of incorporating ethylenediamine (ED)-functionalized metal-organic frameworks (MOFs) on enhancing the electrochemical performance of polypyrrole(PPy)-based conducting polymers as supercapacitor electrode materials was investigated. The stability of terephthalic acid-containing MOFs in different environments, including exposure to acids, relies on the specific metal ions and ligands utilized in their construction. In this research, ED treatment was chosen for its resistance to acid attack. Unlike the post-synthetic modifications observed in previous literature regarding ED-modified MOF5, this study introduces a unique approach where ED is directly integrated into the framework in a single step. The resulting materials, referred to as ED-MOF, were employed in creating a novel composite material concurrently with polypyrrole, designed for use as electrode materials in supercapacitor applications. The optimal loading in the electrochemical preparation of polypyrrole-based conducting polymers was determined using cyclic voltammetry and electrochemical impedance spectroscopy techniques. Areal capacitance for the resulting electrodes was evaluated through cyclic charge-discharge experiments. The areal capacitance of the ED-ZnMOF/1@/PPy/PGE electrode was found to be 575.2 mF.cm−2 at a charge/discharge current density of 1 mA.cm−2. This research underscores the potential advantages of combining conducting polymers and metal-organic frameworks in supercapacitors, as well as other electrochemical systems.

Mincle as a potential intervention target for the prevention of inflammation and fibrosis (Review).

Macrophage‑inducible C‑type lectin receptor (Mincle) is predominantly found on antigen‑presenting cells. It can recognize specific ligands when stimulated by certain pathogens such as fungi and Mycobacterium tuberculosis. This recognition triggers the activation of the nuclear factor‑κB pathway, leading to the production of inflammatory factors and contributing to the innate immune response of the host. Moreover, Mincle identifies lipid damage‑related molecules discharged by injured cells, such as Sin3‑associated protein 130, which triggers aseptic inflammation and ultimately hastens the advancement of renal damage, autoimmune disorders and malignancies by fostering tissue inflammation. Presently, research on the functioning of the Mincle receptor in different inflammatory and fibrosis‑associated conditions has emerged as a popular topic. Nevertheless, there remains a lack of research on the impact of Mincle in promoting long‑lasting inflammatory reactions and fibrosis. Additional investigation is required into the function of Mincle receptors in chronological inflammatory reactions and fibrosis of organ systems, including the progression from inflammation to fibrosis. Hence, the present study showed an overview of the primary roles and potential mechanism of Mincle in inflammation, fibrosis, as well as the progression of inflammation to fibrosis. The aim of the present study was to clarify the potential mechanism of Mincle in inflammation and fibrosis and to offer perspectives for the development of drugs that target Mincle.

A proteomics-based survey reveals thrombospondin-4 as a ligand regulated by the mannose receptor in the injured lung

Receptor-mediated cellular uptake of specific ligands constitutes an important step in the dynamic regulation of individual protein levels in extracellular fluids. With a focus on the inflammatory lung, we here performed a proteomics-based search for novel ligands regulated by the mannose receptor (MR), a macrophage-expressed endocytic receptor. Wildtype and MR-deficient mice were exposed to lipopolysachharide (LPS), after which the protein content in their lung epithelial lining fluid (ELF) was compared by tandem mass tag-based mass spectrometry. More than 1200 proteins were identified in the ELF using this unbiased approach, but only six showed a statistically different abundance. Among these, an unexpected potential new ligand, thrombospondin-4 (TSP-4), displayed a striking 17-fold increased abundance in the MR-deficient mice. Experiments using exogenous addition of TSP-4 to MR-transfected CHO cells or MR-positive alveolar macrophages confirmed that TSP-4 is a ligand for MR-dependent endocytosis. Similar studies revealed that the molecular interaction with TSP-4 depends on both the lectin activity and the fibronectin type-II domain of MR and that a closely related member of the thrombospondin family, TSP-5, is also efficiently internalized by the receptor. This was unlike the other members of this protein family, including TSPs -1 and -2, which are ligands for a close MR homologue known as uPARAP. Our study shows that MR takes part in the regulation of TSP-4, an important inflammatory component in the injured lung, and that two closely related endocytic receptors, expressed on different cell types, undertake the selective endocytosis of distinct members of the thrombospondin family.

Activin E is a transforming growth factor β ligand that signals specifically through activin receptor-like kinase 7.

Activins are one of the three distinct subclasses within the greater Transforming growth factor β (TGFβ) superfamily. First discovered for their critical roles in reproductive biology, activins have since been shown to alter cellular differentiation and proliferation. At present, members of the activin subclass include activin A (ActA), ActB, ActC, ActE, and the more distant members myostatin and GDF11. While the biological roles and signaling mechanisms of most activins class members have been well-studied, the signaling potential of ActE has remained largely unknown. Here, we characterized the signaling capacity of homodimeric ActE. Molecular modeling of the ligand:receptor complexes showed that ActC and ActE shared high similarity in both the type I and type II receptor binding epitopes. ActE signaled specifically through ALK7, utilized the canonical activin type II receptors, ActRIIA and ActRIIB, and was resistant to the extracellular antagonists follistatin and WFIKKN. In mature murine adipocytes, ActE invoked a SMAD2/3 response via ALK7, like ActC. Collectively, our results establish ActE as a specific signaling ligand which activates the type I receptor, ALK7.

A hierarchical cerium metal organic framework-graphene oxide heterostructure composite for rapid and efficient scavenging of fluoride

The rational design of functional heretrostructures with integrated advantages for the efficient adsorption and removal of fluoride is important for water pollution control. In this study, a cerium metal organic framework-graphene oxide heterostructure composite (CeMOF@GO) is successfully prepared through ultrasonic irradiation and vapor diffusion-assisted interfacial growth. CeMOF@GO with hierarchical structure boosts the mass transport of fluoride with adsorption time reduced to 25 min. By virtue of the specific ligand exchange between adsorbed H2O/OH− at metal defect sites and fluoride ions, such unique CeMOF@GO can be utilized for fluoride removal, showing satisfactory adsorption capacity (151.5 mg g−1), excellent specific affinity (with a distribution coefficient of up to 21.865 L g−1), glorious salt tolerance (8.0 mol L−1 of nitrate), wide pH applicability and eminent recycling ability. The practical application of CeMOF@GO is well demonstrated by the selective scavenging of fluoride from diverse environmental water samples. This work is expected to provide a reliable defluorination approach, and bridge the gap between academic MOF-based adsorbent research and practical environmental application.

Robust Approach for Quantifying Glucocorticoid Binding to the Anti-Cortisol Fab Fragment via Native Mass Spectrometry.

In the development of proteins, aptamers, and molecular imprints for diagnostic purposes, a major goal is to obtain a molecule with both a high binding affinity and specificity for the target ligand. Cushing syndrome or Addison's disease can be diagnosed by cortisol level tests. We have previously characterized and solved the crystal structure of an anti-cortisol (17) Fab fragment having a high affinity to cortisol but also significant cross-reactivity to other glucocorticoids, especially the glucocorticoid drug prednisolone. We used native mass spectrometry (MS) to determine the binding affinities of nine steroid hormones to anti-cortisol (17) Fab, including steroidogenic precursors of cortisol. Based on the results, the number of hydroxyl groups in the structure of a steroid ligand plays a key role in the antigen recognition by the Fab fragment as the ligands with three hydroxyl groups, cortisol and prednisolone, had the highest affinities. The antibody affinity toward steroid hormones often decreases with a decrease in the number of hydroxyl groups in the structure. The presence of the hydroxyl group at position C11 increased the affinity more than did the other hydroxyl groups at positions C17 or C21. The binding affinities obtained by native MS were compared to the values determined by surface plasmon resonance (SPR), and the affinities were found to correlate well between these two techniques. Our study demonstrates that native MS with a large dynamic range and high sensitivity is a versatile tool for ligand binding studies of proteins.

Functionalizing Yeast Lipid Droplets as Versatile Biomaterials.

Lipid droplets (LD) are dynamic cellular organelles of ≈1µm diameter in yeast where a neutral lipid core is surrounded by a phospholipid monolayer and attendant proteins. Beyond the storage of lipids, opportunities for LD engineering remain underdeveloped but they show excellent potential as new biomaterials. In this research, LD from yeast Saccharomyces cerevisiae is engineered to display mCherry fluorescent protein, Halotag ligand binding protein, plasma membrane binding v-SNARE protein, and carbonic anhydrase enzyme via linkage to oleosin, an LD anchoring protein. Each protein-oleosin fusion is coded via a single gene construct. The expressed fusion proteins are specifically displayed on LD and their functions can be assessed within cells by fluorescence confocal microscopy, TEM, and as isolated materials via AFM, flow cytometry, spectrophotometry, and by enzyme activity assay. LD isolated from the cell are shown to be robust and stabilize proteins anchored into them. These engineered LD function as reporters, bind specific ligands, guide LD and their attendant proteins into union with the plasma membrane, and catalyze reactions. Here, engineered LD functions are extended well beyond traditional lipid storage toward new material applications aided by a versatile oleosin platform anchored into LD and displaying linked proteins.

Structural basis of ligand specificity and channel activation in an insect gustatory receptor

Gustatory receptors (GRs) are critical for insect chemosensation and are potential targets for controlling pests and disease vectors, making their structural investigation a vital step toward such applications. We present structures of Bombyx mori Gr9 (BmGr9), a fructose-gated cation channel, in agonist-free and fructose-bound states. BmGr9 forms a tetramer similar to distantly related insect odorant receptors (ORs). Upon fructose binding, BmGr9's channel gate opens through helix S7b movements. In contrast to ORs, BmGr9's ligand-binding pocket, shaped by a kinked helix S4 and a shorter extracellular S3-S4 loop, is larger and solvent accessible in both agonist-free and fructose-bound states. Also, unlike ORs, fructose binding by BmGr9 involves helix S5 and a pocket lined with aromatic and polar residues. Structure-based sequence alignments reveal distinct patterns of ligand-binding pocket residue conservation in GR subfamilies associated with different ligand classes. These data provide insight into the molecular basis of GR ligand specificity and function.

Supramolecular Coordination Complexes for Synergistic Cancer Therapy.

Supramolecular coordination complexes (SCCs) are predictable and size-tunable supramolecular self-assemblies constructed through directional coordination bonds between readily available organic ligands and metallic receptors. Based on planar and 3D structures, SCCs can be mainly divided into two categories: metallacycles (e.g., rhomboidal, triangular, rectangular, and hexagonal) and metallacages (e.g., tetrahedral, hexahedral, and dodecahedral). The directional coordination bonds enable the efficient formation of metallacycles and metallacages with well-defined architectures and geometries. SCCs exhibit several advantages, including good directionality, strong interaction force, tunable modularity, and good solution processability, making them highly attractive for biomedical applications, especially in cellular imaging and cancer therapy. Compared with their molecular precursors, SCCs demonstrate enhanced cellular uptake and a strengthened tumor accumulation effect, owing to their inherently charged structures. These properties and the chemotherapeutic potential inherent to organic platinum complexes have promoted their widespread application in antitumor therapy. Furthermore, the defined structures of SCCs, achieved via the design modification of assembly elements and introduction of different functional groups, enable them to combat malignant tumors through multipronged treatment modalities. Because the development of cancer-treatment methodologies integrated in clinics has evolved from single-modality chemotherapy to synergistic multimodal therapy, the development of functional SCCs for synergistic cancer therapy is crucial. While some pioneering reviews have explored the bioapplications of SCCs, often categorized by a specific function or focusing on the specific metal or ligand types, a comprehensive exploration of their synergistic multifunctionality is a critical gap in the current literature.In this Account, we focus on platinum-based SCCs and their applications in cancer therapy. While other metals, such as Pd-, Rh-, Ru-, and Ir-based SCCs, have been explored for cancer therapy by Therrien and Casini et al., platinum-based SCCs have garnered significant interest, owing to their unique advantages in antitumor therapy. These platinum-based SCCs, which enhance antitumor efficacy, are considered prominent candidates for cancer therapies owing to their desirable properties, such as potent antitumor activity, exceptionally low systemic toxicity, active tumor-targeting ability, and enhanced cellular uptake. Furthermore, diverse diagnostic and therapeutic modalities (e.g., chemotherapy, photothermal therapy, and photodynamic therapy) can be integrated into a single platform based on platinum-based SCCs for cancer therapy. Consequently, herein, we summarize our recent research on platinum-based SCCs for synergistic cancer therapy with particular emphasis on the cooperative interplay between different therapeutic methods. In the Conclusions section, we present the key advancements achieved on the basis of our research findings and propose future directions that may significantly impact the field.

Biomimetic Chiral Nanomaterials with Selective Catalysis Activity.

Chiral nanomaterials with unique chiral configurations and biocompatible ligands have been booming over the past decade for their interesting chiroptical effect, unique catalytical activity, and related bioapplications. The catalytic activity and selectivity of chiral nanomaterials have emerged as important topics, that can be potentially controlled and optimized by the rational biochemical design of nanomaterials. In this review, chiral nanomaterials synthesis, composition, and catalytic performances of different biohybrid chiral nanomaterials are discussed. The construction of chiral nanomaterials with multiscale chiral geometries along with the underlying principles for enhancing chiroptical responses are highlighted. Various biochemical approaches to regulate the selectivity and catalytic activity of chiral nanomaterials for biocatalysis are also summarized. Furthermore, attention is paid to specific chiral ligands, materials compositions, structure characteristics, and so on for introducing selective catalytic activities of representative chiral nanomaterials, with emphasis on substrates including small molecules, biological macromolecule, and in-site catalysis in living systems. Promising progress has also been emphasized in chiral nanomaterials featuring structural versatility and improved chiral responses that gave rise to unprecedented chances to utilize light for biocatalytic applications. In summary, the challenges, future trends, and prospects associated with chiral nanomaterials for catalysis are comprehensively proposed.

Development of an aptamer capable of multidrug resistance reversal for tumor combination chemotherapy

Multidrug resistance (MDR) is a major factor in the failure of many forms of tumor chemotherapy. Development of a specific ligand for MDR-reversal would enhance the intracellular accumulation of therapeutic agents and effectively improve the tumor treatments. Here, an aptamer was screened against a doxorubicin (DOX)-resistant human hepatocellular carcinoma cell line (HepG2/DOX) via cell-based systematic evolution of ligands by exponential enrichment. A 50 nt truncated sequence termed d3 was obtained with high affinity and specificity for HepG2/DOX cells. Multidrug resistance protein 1 (MDR1) is determined to be a possible recognition target of the selected aptamer. Aptamer d3 binding was revealed to block the MDR of the tumor cells and increase the accumulation of intracellular anticancer drugs, including DOX, vincristine, and paclitaxel, which led to a boost to the cell killing of the anticancer drugs and lowering their survival of the tumor cells. The aptamer d3-mediated MDR-reversal for effective chemotherapy was further verified in an in vivo animal model, and combination of aptamer d3 with DOX significantly improved the suppression of tumor growth by treating a xenograft HepG2/DOX tumor in vivo. This work demonstrates the feasibility of a therapeutic DNA aptamer as a tumor MDR-reversal agent, and combination of the selected aptamer with chemotherapeutic drugs shows great potential for liver cancer treatments.

In situ polymeric nanomicelle-generating dissolving microneedle patch for enhanced transdermal methotrexate delivery in rheumatoid arthritis treatment

Rheumatoid arthritis (RA) presents a chronic autoimmune pathology. Methotrexate (MTX), a primary therapeutic agent for RA, often administered orally or via injection, has shown dose-dependent systemic toxicity upon prolonged use. Consequently, a pressing need exists for a secure and facile MTX delivery system in RA management. In the current study, the methoxy poly (ethylene glycol)-poly (lactide) polymer (mPEG-PLA) and hyaluronic acid (HA) were used as materials for both nanomicelles and dissolving microneedles (MNs) to facilitate the effective transdermal delivery of MTX in RA therapy. HA, a specific ligand for the CD44 receptor, could deposit on the outer surface of nanomicelles by interacting with mPEG-PLA, which serve as a targeting ligand for macrophages activated at inflamed joints. Upon MTX-loaded polymeric nanomicelles MNs (MTX@PMs MNs) application to the skin, MNs could effectively puncture the stratum corneum and in situ generate nanomicelles with the dissolution of MNs. In vivo therapeutic efficiency evaluations suggested that the MTX@PMs MNs demonstrated enhanced efficacy in mitigating the progression of RA without inducing treatment-related toxicity. Our findings suggest that the utilization of in situ polymeric nanomicelle-generating dissolving microneedle patches holds promise as a potential strategy for the clinical treatment of RA.