Erythritol as a Saccharide Multifunctional Electrolyte Additive for Highly Reversible Zinc Anode.

Dendrite formation and water-triggered side reactions on the surface of Zn metal anodes severely restrict the commercial viability of aqueous zinc-ion batteries (AZIBs). In this work, we introduce erythritol (Et) as an electrolyte additive to enhance the reversibility of zinc anodes, given its cost-effectiveness, mature technology, and extensive utilization in various domains such as food, medicine, and other industries. By combining multiscale theoretical simulation and experimental characterization, it was demonstrated that Et molecules can partially replace the coordination H2O molecules to reshape the Zn2+ solvation sheath and destroy the hydrogen bond network of the aqueous electrolyte. More importantly, Et molecules tend to adsorb on the zinc anode surface, simultaneously inhibit water-triggered side reactions by isolating water and promote uniform and dense deposition by accelerating the Zn2+ diffusion and regulating the nucleation size of the Zn grain. Thanks to this synergistic mechanism, the Zn anode can achieve a cycle life of more than 3900 h at 1 mA cm-2 and an average Coulombic efficiency of 99.77%. Coupling with δ-MnO2 cathodes, the full battery delivers a high specific capacity of 228.1 mAh g-1 with a capacity retention of 76% over 1000 cycles at 1 A g-1.

[Research progress on HLA-G in non-small cell lung cancer].

Human leukocyte antigen G (HLA-G) belongs to the non-classic major histocompatibility complex class Ib (MHC Ib) molecule, including membrane and soluble isoforms. HLA-G regulates the function of various immune cells through corresponding receptors. It is one of the important immunological mechanisms of maternal-fetal tolerance and tumor immune escape. Non-small cell lung cancer (NSCLC) patients account for the highest proportion of lung cancer patients and have poor prognoses. Studies have shown that the gene polymorphism and expression level of HLA-G is closely related to the occurrence and development of NSCLC. It is suggested that HLA-G can be used as a potential biomarker for the early diagnosis, subtype differentiation, treatment, and prognosis of NSCLC patients. HLA-G has clinical value as an auxiliary diagnostic basis. An in-depth study of its mechanism can provide new strategies for the diagnosis and treatment of NSCLC patients.

Effect of Lymphocyte Subsets on Bone Density in Senile Osteoporosis: A Retrospective Study.

Background: Several studies have shown that lymphocyte subsets can mediate the occurrence of osteoporosis (OP); however, the predictive ability of lymphocyte subsets in senile OP has not been elucidated. Purpose: To investigate the ability of lymphocyte subsets to predict senile osteoporosis (OP). Methods and Materials: This study included 44 patients with senile OP and 44 without OP. Dual-energy X-ray absorptiometry (DEXA) was used to determine bone mineral density (BMD). Flow cytometry was used to analyze the absolute counts of the lymphocyte subsets and cytokine levels. Finally, the correlation between BMD and lymphocyte subset counts in the two groups was analyzed. Results: There were no significant differences in age, sex, or weight between the OP and non-OP groups. The absolute counts of total T lymphocytes and CD8+ T lymphocytes in the OP group were significantly lower than those in the non-OP group. The levels of IFN-γ or TNF-α in the OP group were significantly higher or lower, respectively, than those in the non-OP group. PCA showed that age, BMI, total T lymphocytes, CD4+ T lymphocytes, CD8+ T lymphocytes, and B lymphocytes were the principal components of senile OP. The linear regression equation showed that BMD of the right femoral neck significantly decreased with a decline in CD8+ T lymphocyte counts. Conclusion: BMD decreased with a decrease in CD8+ T lymphocytes. The mechanism by which lower lymphocyte subsets lead to lower BMD may be related to abnormal bone metabolism caused by immune aging. Therefore, we considered that CD8+ T lymphocytes could be used to predict the incidence of senile OP.

Bim transfer between Bcl-2-like protein and Hsp70 underlines Bcl-2/Hsp70 crosstalk to regulate apoptosis.

The chaperone heat shock protein 70 (Hsp70) is crucial for avoiding protein misfolding under stress, but it is also upregulated in many kinds of cancers, where its ability to buffer cellular stress prevents apoptosis. Previous research has suggested that Bim, a BH3-only member of the Bcl-2 family proteins, also serves as a cochaperone for Hsp70, which modulates the folding and stabilization of many Hsp70 oncogenic substrates in tumor cells. However, a definitive demonstration of crosstalk between Bcl-2 and Hsp70 family proteins and molecular mechanism remain unclear. Herein, we examined the effects of pan-Bcl-2 inhibitor S1, Hsp70 inhibitor S1g-6 on the K562, U937, H23, HL-60 cell lines and these inhibitors synergistically induce mitochondrial apoptosis in cancer cell lines. Moreover, we identified that Bim transfer between Bcl-2-like protein and Hsp70 underlines Bcl-2/Hsp70 crosstalk in mitochondrial apoptosis pathway. Thus, the synergy of S1 and S1g-6 to induce a panel of cancer cell lines apoptosis by inhibiting free Bim and facilitating oncogenic client AKT folding and activation. Together, our results demonstrated the combination of Bcl-2 inhibitor and Hsp70 inhibitor showed synergistic effect in cancer cells and the potential to decrease tumor regression.

Fabrication of self-healing pectin/chitosan hybrid hydrogel via Diels-Alder reactions for drug delivery with high swelling property, pH-responsiveness, and cytocompatibility.

Self-healing hydrogels with pH-responsiveness could protect loaded drugs from being destroyed till it arrives to the target. The pectin-based hydrogel is a candidate due to the health benefit, anti-inflammation, antineoplastic activity, nontoxicity, and biospecific degradation, et al. However, the abundant existence of water-soluble branched heteropolysaccharide chains influenced its performance resulting in limitation of the potential. In the present study, we prepared a series of self-healing pectin/chitosan hydrogels via the Diels-Alder reaction. Moreover, pectin/chitosan composite hydrogel was prepared as a contrast. By comparison, it can be seen that the Diels-Alder reaction greatly improved the cross-linking density of hydrogels. The self-healing experiments showed excellent self-healing performance. In different swelling mediums, significant transformation in the swelling ratio was shown, indicating well-swelling property, pH- and thermo-responsiveness. The drug loading and release studies presented high loading efficiency and sustained release performance. The cytotoxicity assay that showed a high cell proliferation ratio manifested great cytocompatibility.

Small-molecule inhibitor targeting the Hsp70-Bim protein-protein interaction in CML cells overcomes BCR-ABL-independent TKI resistance.

Herein, we screened a novel inhibitor of the Hsp70-Bim protein-protein interaction (PPI), S1g-2, from a Bcl-2 inhibitor library; this compound specifically disrupted the Hsp70-Bim PPI by direct binding to an unknown site adjacent to that of an allosteric Hsp70 inhibitor MKT-077, showing binding affinity in sub-µM concentration range. S1g-2 exhibited overall 5-10-fold higher apoptosis-inducing activity in CML cells, primary CML blasts, and BCR-ABL-transformed BaF3 cells than other cancer cells, normal lymphocytes, and BaF3 cells, illustrating Hsp70-Bim PPI driven by BCR-ABL protects CML through oncoclient proteins that enriched in three pathways: eIF2 signaling, the regulation of eIF4E and p70S6K signaling, and the mTOR signaling pathways. Moreover, S1g-2 progressively enhanced lethality along with the increase in BCR-ABL-independent TKI resistance in the K562 cell lines and is more effective in primary samples from BCR-ABL-independent TKI-resistant patients than those from TKI-sensitive patients. By comparing the underlying mechanisms of S1g-2, MKT-077, and an ATP-competitive Hsp70 inhibitor VER-155008, the Hsp70-Bim PPI was identified to be a CML-specific target to protect from TKIs through the above three oncogenic signaling pathways. The in vivo activity against CML and low toxicity endows S1g-2 a first-in-class promising drug candidate for both TKI-sensitive and resistant CML.

A novel Hsp70 inhibitor specifically targeting the cancer-related Hsp70-Bim protein-protein interaction.

Targeting cancer-related Hsp70-Bim protein-protein interactions (PPIs) offers a new strategy for the design of Hsp70 inhibitors. Herein, we discovered a novel Hsp70 inhibitor, S1g-6, based on the established BH3 mimetics. S1g-6 exhibited sub-µM binding affinity toward Hsp70 and selectively disrupted Hsp70-Bim PPI. The target specificity of S1g-6in situ was validated by affinity-based protein profiling, co-immunoprecipitation, and cell-based shRNA assays. S1g-6 specifically antagonized the ATPase activity of Hsp70 upon recruiting Bim and showed selective apoptosis induction in some cancer cell lines over normal ones through suppression of some oncogenic clients of Hsp70, representing a new class of antitumor candidates. Hsp70-Bim PPI exhibited cancer-dependent role as a potential anti-cancer target.

A "Three-in-One" Multifunctional Probe for Bcl-2/Mcl-1 Profiling and Visualizing in Situ.

Bcl-2 and Mcl-1, the two arms of the anti-apoptotic Bcl-2 family proteins, have been identified as key regulators of apoptosis and effective therapeutic targets of cancer. However, no small-molecular probe is capable of profiling and visualizing both Bcl-2 and Mcl-1 simultaneously in situ. Herein, we report a multifunctional molecular probe (BnN3 -OPD-Alk) by a "three-in-one" molecular designing strategy, which integrated the Bcl-2/Mcl-1 binding ligand, fluorescent reporter group and photoreactive group azido into the same scaffold. BnN3 -OPD-Alk exhibited sub-micromolar affinities to Bcl-2/Mcl-1 and bright green self-fluorescence. It was then successfully applied for Bcl-2/Mcl-1 labeling, capturing, enriching, and bioimaging both in vitro and in cells. This strategy could facilitate the precise early diagnosis and effective therapy of dual Bcl-2/Mcl-1-related diseases.

Targeting the Allosteric Pathway That Interconnects the Core-Functional Scaffold and the Distal Phosphorylation Sites for Specific Dephosphorylation of Bcl-2.

Protein phosphorylation is the most significant post-translational modification for regulating cellular activities, but site-specific modulation of phosphorylation is still challenging. Using three-dimensional NMR spectra, molecular dynamics simulations, and alanine mutations, we identified that the interaction network between pT69/pS70 and R106/R109 residues prevents the phosphorylation sites from exposure to phosphatase and subsequent dephosphorylation. A Bcl-2-dephosphorylation probe, S1-6e, was designed by installing a carboxylic acid group to a Bcl-2 inhibitor. The carboxyl group competitively disrupts the interaction network between R106/R109 and pT69/pS70 and subsequently facilitates Bcl-2 dephosphorylation in living cells. As a result, S1-6e manifests a more effective apoptosis induction in pBcl-2-dependent cancer cells than other inhibitors exhibiting a similar binding affinity for Bcl-2. We believe that targeting the allosteric pathways interconnecting the core-functional domain and the phosphorylation site can be a general strategy for a rational design of site-specific dephosphorylating probes, since the allosteric pathway has been discovered in a variety of proteins.

The chaperone Hsp70 is a BH3 receptor activated by the pro-apoptotic Bim to stabilize anti-apoptotic clients.

The chaperone heat shock protein 70 (Hsp70) is crucial for avoiding protein misfolding under stress, but is also up-regulated in many kinds of cancers, where its ability to buffer cellular stress prevents apoptosis. Previous research has suggested Hsp70 interacts with pro-apoptotic Bcl-2 family proteins, including Bim and Bax. However, a definitive demonstration of this interaction awaits, and insights into the structural basis and molecular mechanism remain unclear. Earlier studies have identified a Bcl-2 homology 3 (BH3) domain present in Bcl-2 family members that engages receptors to stimulate apoptosis. We now show that Hsp70 physically interacts with pro-apoptotic multidomain and BH3-only proteins via a BH3 domain, thereby serving as a novel BH3 receptor, using in vitro fluorescent polarization (FP), isothermal titration calorimetry (ITC), and cell-based co-immunoprecipitation (co-IP) experiments, 1H-15N-transverse relaxation optimized spectroscopy (TROSY-HSQC), trypsin proteolysis, ATPase activity, and denatured rhodanese aggregation measurements further demonstrated that BimBH3 binds to a novel allosteric site in the nucleotide-binding domain (NBD) of Hsp70, by which Bim acts as a positive co-chaperone to promote the ATPase activity and chaperone functions. A dual role of Hsp70's anti-apoptotic function was revealed that when it keeps Bim in check to inhibit apoptosis, it simultaneously stabilizes oncogenic clients including AKT and Raf-1 with the aid of Bim. Two faces of Bim in cell fate regulation were revealed that in opposite to its well-established pro-apoptotic activator role, Bim could help the folding of oncogenic proteins.

Design and preparation of quaternized pectin-Montmorillonite hybrid film for sustained drug release.

Montmorillonite (MMT) presents nonocclusive lamellar structure which restricts the potential use for sustained drug release. To solve the limitation, the quaternized pectin (QP) was synthesized and firstly introduced to form QP-MMT hybrid film containing 5-FU. The Fourier transform infrared spectroscopy (FT-IR) and X-Ray diffraction (XRD) were employed to determine the variation of the functional group and crystallinity between pectin and QP. The resultant composite film was characterized by FT-IR, XRD and Field Emission Scanning Electron Microscope. The results of the characterization indicated that intercalation reaction happened in the blending process. The optimum film showed high value of drug encapsulation efficiency (36.50%) and loading efficiency (80.30%). The in vitro drug release studies revealed that the MMT significantly improved the sustained-release performance in all simulated mediums. The cumulative release rate of sample QP10-MMT0.1 was all around 20% in the first half-hour in all simulated mediums and sustained increased for more than 8 h. The cytotoxicity assay was performed to prove the great biocompatibility of QP-MMT hybrid film. The present study introduced a facile route to prepare the composite film which presented sustained drug release performance.

Structure-based design, synthesis, and evaluation of Bcl-2/Mcl-1 dual inhibitors.

Based on our previously reported Bcl-2/Mcl-1 dual inhibitor 4-thiomorpholinyl-2-cyano-3-amidinophenalenone (A1) that simultaneously occupies the p2 and p4 hydrophobic pockets of Bcl-2 and Mcl-1, we optimized molecules with different bond angles of the groups extending to the p4 pocket and bulky hydrophobic groups to explore p2. Research on structure-activity relationship resulted in a new derivative B4 that is capable of occupying both the p2 and p4 more deeply and completely than A1, with Ki values determined by fluorescence polarization assay (FPAs) improving to 0.31 µM for Bcl-2 and 0.16 µM for Mcl-1. Furthermore, B4 exhibited selective lethality on cancer cells over normal cells. It showed stronger apoptosis induction than (-)-gossypol on a Bcl-2/Mcl-1-dependent cancer cell line and killed an Mcl-1-dependent cell line which is resistant to ABT-199 treatment.

Deconstruction of oriented crystalline cellulose by novel levulinic acid based deep eutectic solvents pretreatment for improved enzymatic accessibility.

Discovering green solvents and their inner mechanisms for efficient deconstruction of lignocellulosic biomass recalcitrance are receiving growing interests. In this work, eco-friendly levulinic acid (LA) based deep eutectic solvents (DES) were proposed for pretreatment on moso bamboo by combining acetamide (Am), betaine (Ba) and choline chloride (ChCl) as hydrogen bonding acceptors. LA/ChCl pretreated materials showed optimal enzymatic accessibility with the highest glucose yield (79.07%) because of its higher lignin removal, morphological disruption and decreased crystallinity. Moreover, the microvoids (averagely 30 nm) and cracks were observed for cellulose microfibrils in anisotropic directions, which resulted in shorter microfibrils and crystallites facilitating the enzymatic hydrolysis. The studies on recyclability revealed that LA/Ba DES had better recycling performance due to its maintaining capability of lignin extraction. Series of supramolecular changes on oriented crystalline cellulose were determined in this work by novel LA based DES, which may provide new alternatives for biomass pretreatments.

High Purity and Low Molecular Weight Lignin Nano-Particles Extracted from Acid-Assisted MIBK Pretreatment.

A simple and economical biorefinery method, organosolv methyl isobutyl ketone (MIBK) pretreatment assisted by Lewis acid ferric trichloride hydrolysis, was proposed for fractionating the lignin from extractive-free Eucalyptus powder at the nanoscale, accompanied by another product furfural, derived from hemicellulose. Under the conditions (180 °C, 1 h) optimized based on the best yield of furfural, 40.13% of the acid-insoluble lignin (AIL) could be obtained with a high purity of 100%, a low molecular weight of 767 (Mn) and improved thermostability. The extracted lignin was characterized by its chemical structure, thermostability, homogeneity, molecular weight, and morphology as compared with milled wood lignin (MWL). The results showed significant variations in chemical structures of the extracted lignin during the pretreatment. Specifically, the aryl ether linkage and phenylcoumarans were broken severely while the resinols were more resistant. The G-type lignin was more sensitive to degradation than the S-type, and after the pretreatment, H-type lignin was formed, indicating the occurrence of a demethoxylation reaction at high temperature. Moreover, the lignin nano-particles were identified visually by AFM and TEM images. The dynamic light scattering (DLS) showed that the average diameter of the measured samples was 131.8 nm, with the polydispersity index (PDI) of 0.149. The MIBK-lignin nano-particles prepared in our laboratory exhibit high potentials in producing high functional and valuable materials for the application in wide fields.

Using CETSA assay and a mathematical model to reveal dual Bcl-2/Mcl-1 inhibition and on-target mechanism for ABT-199 and S1.

Deepening understanding of how Bcl-2 family proteins protect cancer cells from apoptosis has driven the development of 'BH3 mimetic' drugs that target various anti-apoptotic Bcl-2-like proteins by mimicking their natural inhibitors, the BH3-only proteins. The proof of target engagement and an on-target mechanism validation are critical for evaluating drug development potential. To evaluate target engagement of BH3 mimetics in cells, we measured binding potency of ABT-199, A-1210477 and ABT-737 to Bcl-2 and Mcl-1 proteins by using a dose-response cellular thermal shift assay (CETSA), similar affinity rank-order and selectivity were obtained in comparison with in vitro binding assays. A proof of direct target engagement for S1 and AT-101 was obtained through CETSA assay. By using a previously established mathematical model, we simulated individual death response of various cancer cell lines to ABT-199, S1 or AT-101 in comparison with experimental data. A positive correlation between model predictions and experimental data for ABT-199 and S1 showed that dual Bcl-2 and Mcl-1 target engagement underlies their anticancer efficacy. In contrast, an off-target effect was determined for AT-101.

Heteropoly acids enhanced neutral deep eutectic solvent pretreatment for enzymatic hydrolysis and ethanol fermentation of Miscanthus x giganteus under mild conditions.

To improve the neutral DES (choline chloride/glycerol) pretreatment performance, three environmentally friendly heteropoly acids (phosphotungstic, phosphomolybdic and silicotungstic acids) were used as catalysts. Pretreatment with silicotungstic acid at 120 °C for 3 h resulted in 97.3% of enzymatic digestibility at an enzyme loading of 15FPU/g substrate, which was approximately eight times more than that of raw samples. More importantly, 80% of glucose yield was obtained within 12 h. Simultaneously, 81.8% of ethanol yield was achieved in the SSSF process. The efficient conversion was ascribed to the significant delignification (89.5%), which resulted in the exposure of more accessible specific surface area. This was attributed to that the proton (H+) from heteropoly acids could significantly contribute to the lignin degradation. Intriguingly, trace acetic acid (0.39 g/L) and HMF (0.21-0.95 g/L) in the pretreatment liquor were produced without any significant deleterious effects. These discoveries provide new insights for efficient biomass conversion under mild conditions.

Proteolysis Targeting Chimeras for the Selective Degradation of Mcl-1/Bcl-2 Derived from Nonselective Target Binding Ligands.

Proteolysis targeting chimera (PROTAC) recruits an E3 ligase to a target protein to induce its ubiquitination and subsequent degradation. We reported success in the development of two PROTACs (C3 and C5) that potently and selectively induce the degradation of Mcl-1 and Bcl-2 (DC50 = 0.7 and 3.0 µM), respectively, by introducing the E3 ligase cereblon-binding ligand pomalidomide to Mcl-1/Bcl-2 dual inhibitors S1-6 and Nap-1 with micromolar-range affinity. C3-induced Mcl-1 ubiquitination translated into much more lethality in Mcl-1-dependent H23 cells than the most potent Mcl-1 occupancy-based inhibitor A-1210477 with nanomolar-range affinity. Moreover, structure-activity relationship analysis and molecular dynamic simulations discovered the structural basis for turning nonselective or promiscuous Bcl-2 family ligands into selective PROTACs. C3 and C5 exhibited reversible depletion in living cells, which provides a new potent toolkit for gain-of-function studies to probe the dynamic roles of Bcl-2 and Mcl-1 in apoptosis networks.

Discovery of selective Mcl-1 inhibitors via structure-based design and structure-activity relationship analysis.

Based on Nap-1, a Mcl-1/Bcl-2 dual inhibitor reported by our group, we carried out a structure-guided molecular design and structure-activity relationship (SAR) analysis to study structural features contributing to Mcl-1 binding selectivity and affinity. A series of derivatives of Nap-1 with various pharmacophores were synthesized and among them a dual Mcl-1/Bcl-2 inhibitor A4 with enhanced affinities (IC50 = 0.15 µM for Mcl-1, 0.43 µM for Bcl-2) and a selective Mcl-1 inhibitor B9 with a 20-fold selectivity over Bcl-2 (IC50 = 0.51 µM vs 9.46 µM) were obtained by enzyme linked immunosorbent assay (ELISA). The SAR data and binding modes of A4 and B9 investigated by 2D-NMR derived docking study illustrated that p2 pockets exhibiting different geometry and binding features between Mcl-1 and Bcl-2 contribute to specific binding properties of Mcl-1. In addition, apoptosis-inducing potencies of A4 and B9 were consistent with their binding selectivity determined in vitro.

A novel turn-on fluorescent probe for selective sensing and imaging of glutathione in live cells and organisms.

We synthesized six 1-oxo-1H-phenalene-2,3-dicarbonitrile (OPD)-based probes with various leaving groups using an arylthioether linker and for the first time identified the probe O-NH2 capable of highly selective detection of glutathione over cysteine/homocysteine in vitro and in vivo based on an aromatic nuclear substitution reaction (SNAr) mechanism. The fluorescence of the probe O-NH2 was quenched because of the photoinduced electron transfer (PET) process, but switched on by a glutathione-triggered specific recognition reaction between the probe O-NH2 and glutathione. The recognition mechanism for glutathione was explored and verified by theoretical calculations and ESI-MS analysis. Using O-NH2 as the probe, the GSH fluorescence images were demonstrated in HeLa cells and the intracellular GSH levels in different imatinib-resistant K562 tumor cells were firstly determined. Further, O-NH2 was utilized to detect glutathione in D. magna and zebrafish embryos. The combined results indicate that O-NH2 can be applied as an effective tool for detecting glutathione in biological investigations.

Proteome-Wide Identification of On- and Off-Targets of Bcl-2 Inhibitors in Native Biological Systems by Using Affinity-Based Probes (AfBPs).

Selective inhibition of proteins of the Bcl-2 family by small-molecule inhibitors is a promising new approach in drug discovery. However, information about how these molecules interact with their cellular targets (on- and off-) is highly limited. We have designed and synthesized photoreactive and "clickable" affinity-based probes (AfBPs)-Nap-2 and Nap-5-by introducing photo-crosslinkers onto Nap-1, a fluorescent derivative of small-molecule Bcl-2 inhibitor S1-6. The resulting trifunctional probes Nap-2 and Nap-5 can enrich, visualize, and enable the identification of cellular on- and off-targets of Bcl-2 inhibitors both in vitro and in situ. Tubulin was validated as an off-target of Bcl-2 inhibitors (Nap-1 and S1-6) by large-scale cell-based proteome profiling and pull-down/western blotting (PD/WB) with Nap-2 and Nap-5. It was preliminarily illustrated to be a BH3-containing protein because some well-known Bcl-2 inhibitors can block the labeling of tubulin by Nap-2.