Crystal structure of hetero hexameric 11S seed storage protein of hazelnut.

Edible plant seeds provide a relatively inexpensive source of protein and make up a large part of nutrients for humans. Plant seeds accumulate storage proteins during seed development. Seed storage proteins act as a reserve of nutrition for seed germination and seedling growth. However, seed storage proteins may be allergenic, and the prevalence of food allergy has increased rapidly in recent years. The 11S globulins account for a significant number of known major food allergens. They are of interest to the public and the agricultural industry because of food safety concerns and the need for crop enhancement. We sought to determine the crystal structure of Cor a 9, the 11 S storage protein of hazelnut and a food allergen. The structure was refined to 1.92 Å, and the R and Rfree for the refined structure are 17.6% and 22.5%, respectively. The structure of Cor a 9 showed a hetero hexamer of an 11S seed storage protein for the first time. The hexamer was two trimers associated back-to-back. Two long alpha helixes at the C-terminal end of the acidic domain of one of the Cor a 9 isoforms lay at the trimer-trimer interface's groove. These data provided much-needed information about the allergenicity of the 11S seed proteins. The information may also facilitate a better understanding of the folding and transportation of 11S seed storage proteins.

Drug Repurposing of ACT001 to Discover Novel Promising Sulfide Prodrugs with Improved Safety and Potent Activity for Neutrophil-Mediated Antifungal Immunotherapy.

Neutrophil-mediated immunotherapy is a promising strategy for treating Candida albicans infection due to its potential in dealing with drug-resistant events. Our previous study found that ACT001 exhibited good antifungal immunotherapeutic activity by inhibiting PD-L1 expression in neutrophils, but its strong cytotoxicity and high BBB permeability hindered its antifungal application. To address these deficiencies, a series of novel sulfide derivatives were designed and synthesized based on a slow-release prodrug strategy. Among these derivatives, compound 16 exhibited stronger inhibition of PD-L1 expression, less cytotoxicity to neutrophils, and lower BBB permeability than ACT001. Compound 16 also significantly enhanced neutrophil-mediated antifungal immunity in C. albicans infected mice, with acceptable pharmacokinetic properties and good oral safety. Moreover, pharmacological mechanism studies demonstrated that ACT001 and compound 16 reduced PD-L1 expression in neutrophils by directly targeting STAT3. Briefly, this study provided a novel prototype compound 16 which exhibited great potential in neutrophil-mediated antifungal immunotherapy.

Pharmacological inhibition of Kir4.1 evokes rapid-onset antidepressant responses.

Major depressive disorder, a prevalent and severe psychiatric condition, necessitates development of new and fast-acting antidepressants. Genetic suppression of astrocytic inwardly rectifying potassium channel 4.1 (Kir4.1) in the lateral habenula ameliorates depression-like phenotypes in mice. However, Kir4.1 remains an elusive drug target for depression. Here, we discovered a series of Kir4.1 inhibitors through high-throughput screening. Lys05, the most potent one thus far, effectively suppressed native Kir4.1 channels while displaying high selectivity against established targets for rapid-onset antidepressants. Cryogenic-electron microscopy structures combined with electrophysiological characterizations revealed Lys05 directly binds in the central cavity of Kir4.1. Notably, a single dose of Lys05 reversed the Kir4.1-driven depression-like phenotype and exerted rapid-onset (as early as 1 hour) antidepressant actions in multiple canonical depression rodent models with efficacy comparable to that of (S)-ketamine. Overall, we provided a proof of concept that Kir4.1 is a promising target for rapid-onset antidepressant effects.

Trilogy of drug repurposing for developing cancer and chemotherapy-induced heart failure co-therapy agent.

Chemotherapy-induced complications, particularly lethal cardiovascular diseases, pose significant challenges for cancer survivors. The intertwined adverse effects, brought by cancer and its complication, further complicate anticancer therapy and lead to diminished clinical outcomes. Simple supplementation of cardioprotective agents falls short in addressing these challenges. Developing bi-functional co-therapy agents provided another potential solution to consolidate the chemotherapy and reduce cardiac events simultaneously. Drug repurposing was naturally endowed with co-therapeutic potential of two indications, implying a unique chance in the development of bi-functional agents. Herein, we further proposed a novel "trilogy of drug repurposing" strategy that comprises function-based, target-focused, and scaffold-driven repurposing approaches, aiming to systematically elucidate the advantages of repurposed drugs in rationally developing bi-functional agent. Through function-based repurposing, a cardioprotective agent, carvedilol (CAR), was identified as a potential neddylation inhibitor to suppress lung cancer growth. Employing target-focused SAR studies and scaffold-driven drug design, we synthesized 44 CAR derivatives to achieve a balance between anticancer and cardioprotection. Remarkably, optimal derivative 43 displayed promising bi-functional effects, especially in various self-established heart failure mice models with and without tumor-bearing. Collectively, the present study validated the practicability of the "trilogy of drug repurposing" strategy in the development of bi-functional co-therapy agents.

Protective factors for children with autism spectrum disorder during COVID-19-related strict lockdowns: a Shanghai autism early developmental cohort study.

BACKGROUND: COVID-19 lockdowns increased the risk of mental health problems, especially for children with autism spectrum disorder (ASD). However, despite its importance, little is known about the protective factors for ASD children during the lockdowns. METHODS: Based on the Shanghai Autism Early Developmental Cohort, 188 ASD children with two visits before and after the strict Omicron lockdown were included; 85 children were lockdown-free, while 52 and 51 children were under the longer and the shorter durations of strict lockdown, respectively. We tested the association of the lockdown group with the clinical improvement and also the modulation effects of parent/family-related factors on this association by linear regression/mixed-effect models. Within the social brain structures, we examined the voxel-wise interaction between the grey matter volume and the identified modulation effects. RESULTS: Compared with the lockdown-free group, the ASD children experienced the longer duration of strict lockdown had less clinical improvement (ß = 0.49, 95% confidence interval (CI) [0.19-0.79], p = 0.001) and this difference was greatest for social cognition (2.62 [0.94-4.30], p = 0.002). We found that this association was modulated by parental agreeableness in a protective way (-0.11 [-0.17 to -0.05], p = 0.002). This protective effect was enhanced in the ASD children with larger grey matter volumes in the brain's mentalizing network, including the temporal pole, the medial superior frontal gyrus, and the superior temporal gyrus. CONCLUSIONS: This longitudinal neuroimaging cohort study identified that the parental agreeableness interacting with the ASD children's social brain development reduced the negative impact on clinical symptoms during the strict lockdown.

pLM4Alg: Protein Language Model-Based Predictors for Allergenic Proteins and Peptides.

The rising prevalence of allergy demands efficient and accurate bioinformatic tools to expedite allergen identification and risk assessment while also reducing wet experiment expenses and time. Recently, pretrained protein language models (pLMs) have successfully predicted protein structure and function. However, to our best knowledge, they have not been used for predicting allergenic proteins/peptides. Therefore, this study aims to develop robust models for allergenic protein/peptide prediction using five pLMs of varying sizes and systematically assess their performance through fine-tuning with a convolutional neural network. The developed pLM4Alg models have achieved state-of-the-art performance with accuracy, Matthews correlation coefficient, and area under the curve scoring 93.4-95.1%, 0.869-0.902, and 0.981-0.990, respectively. Moreover, pLM4Alg is the first model capable of handling prediction tasks involving residue-missed sequences and sequences containing nonstandard amino acid residues. To facilitate easy access, a user-friendly web server (https://f6wxpfd3sh.us-east-1.awsapprunner.com) has been established. pLM4Alg is expected to become the leading machine learning-based prediction model for allergenic peptides and proteins. Its collaboration with other predictors holds great promise for accelerating allergy research.

Water-soluble fraction of pea protein isolate is critical for the functionality of protein-glucose conjugates obtained via wet-heating Maillard reaction.

Wet-heating Maillard reaction (MR) has been applied to improve the function of proteins by conjugating with soluble carbohydrates. However, the impact of soluble solutes particularly in plant protein on the degree of MR and the properties of the corresponding conjugates has yet to be discussed. In this study, high-intensity ultrasound (HIUS) was utilized to pretreat commercial pea protein isolate in order to improve its solubility. Two different fractions including soluble fraction (SUPPI) and whole solution (UPPI) of HIUS treated PPI were conjugated with glucose (G) to prepare SUPPI-G and UPPI-G, respectively, over a course of 24 h wet-heating at 80 °C. Conjugation was confirmed by the degree of glycation, SDS-PAGE, FTIR, and intrinsic fluorescence analysis. Color change and glucose content analysis showed that the degree of MR was greater when using SUPPI rather than UPPI. The solubility of SUPPI-G was further improved by 24 h of MR while it remained unchanged for UPPI-G. The emulsifying activity index and foaming capability of SUPPI-G were similar to those of UPPI-G. Interfacial properties determined by dynamic adsorption and dilatational rheology at both oil-water and air-water interface suggested that insoluble fraction of UPPI is essential to make stable emulsions and foams. In conclusion, the proportion of soluble protein in PPI is critical to its wet-heating MR based conjugation with glucose and the solubility of the conjugates.

New failure mechanism for evaluating ultimate inclined load adjacent to slope.

A new failure mechanism is proposed for calculating the ultimate inclined load adjacent to the slope, i.e., the slope is in the limit state when the critical slope contour and the slope surface are at the critical position where two intersections will occur. The conventional view is that the critical slope contour calculated by the method of characteristics has only a concave shape. This study found that the critical slope contour changes from concave to convex when the inclined load imposed on the slope top surface increases. The feasibility of the proposed method is verified by the finite element limit analysis (FELA) and the definition of the ultimate load. The parametric analysis showed that the current method of characteristics (CMOC) overestimated the ultimate inclined load and gave an incorrect conclusion since it assumed larger failure models at a low strength ratio or large friction angle. The proposed method does not require assumption or search of the failure models, and it can solve the shortcomings of CMOC.

Structural repurposing of SGLT2 inhibitor empagliflozin for strengthening anti-heart failure activity with lower glycosuria.

Sodium-glucose cotransporter 2 (SGLT2) inhibitors have been reapproved for heart failure (HF) therapy in patients with and without diabetes. However, the initial glucose-lowering indication of SGLT2i has impeded their uses in cardiovascular clinical practice. A challenge of SGLT2i then becomes how to separate their anti-HF activity from glucose-lowering side-effect. To address this issue, we conducted structural repurposing of EMPA, a representative SGLT2 inhibitor, to strengthen anti-HF activity and reduce the SGLT2-inhibitory activity according to structural basis of inhibition of SGLT2. Compared to EMPA, the optimal derivative JX01, which was produced by methylation of C2-OH of the glucose ring, exhibited weaker SGLT2-inhibitory activity (IC50 > 100 nmol/L), and lower glycosuria and glucose-lowering side-effect, better NHE1-inhibitory activity and cardioprotective effect in HF mice. Furthermore, JX01 showed good safety profiles in respect of single-dose/repeat-dose toxicity and hERG activity, and good pharmacokinetic properties in both mouse and rat species. Collectively, the present study provided a paradigm of drug repurposing to discover novel anti-HF drugs, and indirectly demonstrated that SGLT2-independent molecular mechanisms play an important role in cardioprotective effects of SGLT2 inhibitors.

UniDL4BioPep: a universal deep learning architecture for binary classification in peptide bioactivity.

Identification of potent peptides through model prediction can reduce benchwork in wet experiments. However, the conventional process of model buildings can be complex and time consuming due to challenges such as peptide representation, feature selection, model selection and hyperparameter tuning. Recently, advanced pretrained deep learning-based language models (LMs) have been released for protein sequence embedding and applied to structure and function prediction. Based on these developments, we have developed UniDL4BioPep, a universal deep-learning model architecture for transfer learning in bioactive peptide binary classification modeling. It can directly assist users in training a high-performance deep-learning model with a fixed architecture and achieve cutting-edge performance to meet the demands in efficiently novel bioactive peptide discovery. To the best of our best knowledge, this is the first time that a pretrained biological language model is utilized for peptide embeddings and successfully predicts peptide bioactivities through large-scale evaluations of those peptide embeddings. The model was also validated through uniform manifold approximation and projection analysis. By combining the LM with a convolutional neural network, UniDL4BioPep achieved greater performances than the respective state-of-the-art models for 15 out of 20 different bioactivity dataset prediction tasks. The accuracy, Mathews correlation coefficient and area under the curve were 0.7-7, 1.23-26.7 and 0.3-25.6% higher, respectively. A user-friendly web server of UniDL4BioPep for the tested bioactivities is established and freely accessible at https://nepc2pvmzy.us-east-1.awsapprunner.com. The source codes, datasets and templates of UniDL4BioPep for other bioactivity fitting and prediction tasks are available at https://github.com/dzjxzyd/UniDL4BioPep.

Production of Fish Analogues from Plant Proteins: Potential Strategies, Challenges, and Outlook.

Fish products are consumed by human beings as a high-quality protein source. However, overfishing, and pollution puts out an urgent call to seek a new strategy to substitute fish protein for secure eco-sustainability. Plant-based fish analogs, which mimic the structure, texture, and flavor of fish meat products, are a rapid-growing segment of the food products. The purpose of this review is to discuss the feasibility and potential strategies for developing plant-based fish analog. The nutritional properties, especially the protein quality of plant-based fish analogs, were discussed. Furthermore, a thorough comparison was made between fish and terrestrial animal muscle structures, including both macroscopical and microscopical structures. Potential processing technologies for producing plant-based fish analogs from plant proteins and approaches for the characterization of the fish analog structures were elaborated. Comparing all the current processing techniques, extrusion is the predominately used technique in the current industry. At the same time, 3D-printing and electrospinning have shown the prominent potential of mimicking fish muscle structure as bottom-up approaches. Finally, key challenges and future research were discussed for the potential commercialization of plant-based fish analogues. The primary focus of this review covers the innovative works that were indexed in the Web of Science Core Collection in the past five years.

New failure mechanism for evaluating seismic and static undrained bearing capacity adjacent to 2D slope.

The seismic or static undrained slip line field theory and Cauchy, Riemann, mixed boundary value problems for undrained soil slopes are derived. A new failure mechanism is proposed to determine the undrained bearing capacity adjacent to slopes. The effects of geometric or strength parameters and seismic forces on static and seismic undrained bearing capacity are investigated. The convergence of the proposed method is proved. The static and seismic undrained bearing capacities predicted by the proposed method are close to those of the currently existing methods. The proposed method does not need to assume or search the failure modes, and a new limit state evaluation index is given.

A Heterogeneity-Aware Replacement Policy for the Partitioned Cache on Asymmetric Multi-Core Architectures.

In an asymmetric multi-core architecture, multiple heterogeneous cores share the last-level cache (LLC). Due to the different memory access requirements among heterogeneous cores, the LLC competition is more intense. In the current work, we propose a heterogeneity-aware replacement policy for the partitioned cache (HAPC), which reduces the mutual interference between cores through cache partitioning, and tracks the shared reuse state of each cache block within the partition at runtime to guide the replacement policy to keep cache blocks shared by multiple cores in multithreaded programs. In the process of updating the reuse state, considering the difference of memory accesses to LLC by heterogeneous cores, the cache replacement policy tends to keep cache blocks required by big cores, to better improve the LLC access efficiency of big cores. Compared with LRU and the SRCP, which are the state-of-the-art cache replacement algorithms, the performance of big cores can be significantly improved by HAPC when running multithreaded programs, while the impact on little cores is almost negligible, thus improving the overall performance of the system.

N-Acylamino Saccharin as an Emerging Cysteine-Directed Covalent Warhead and Its Application in the Identification of Novel FBPase Inhibitors toward Glucose Reduction.

With a resurgence of covalent drugs, there is an urgent need for the identification of new moieties capable of cysteine bond formation. Herein, we report on the N-acylamino saccharin moieties capable of novel covalent reactions with cysteine. Their utility as alternative electrophilic warheads was demonstrated through the covalent modification of fructose-1,6-bisphosphatase (FBPase), a promising target associated with cancer and type 2 diabetes. The cocrystal structure of title compound W8 bound with FBPase unexpectedly revealed that the N-acylamino saccharin moiety worked as an electrophile warhead that covalently modified the noncatalytic C128 site in FBPase while releasing saccharin, suggesting a previously undiscovered covalent reaction mechanism of saccharin derivatives with cysteine. Treatment of title compound W8 displayed potent inhibition of glucose production in vitro and in vivo. This newly discovered reactive warhead supplements the current repertoire of cysteine covalent modifiers while avoiding some of the limitations generally associated with established moieties.

Discovery of novel dual RAGE/SERT inhibitors for the potential treatment of the comorbidity of Alzheimer's disease and depression.

Depression is identified as one of the most common psychiatric symptoms in Alzheimer's disease (AD). The comorbidity of AD and depression increases the burden of clinical treatment and care in elderly patients. In order to find new treatment options, we first proposed the dual RAGE/SERT inhibitors by fusing the key pharmacophore of vilazodone and azeliragon for the potential treatment of AD with comorbid depression. After a series of structural modifications, 34 dual-target directed ligands were designed and synthesized, and their RAGE and SERT inhibitory activities were systematically evaluated. Among them, compound 12 showed good dual-target bioactivities against RAGE (IC50 = 8.26 ± 1.12 µM) and SERT (IC50 = 31.09 ± 5.15 nM) in vitro, better safety profile than azeliragon, good liver microsomal stability, weak CYP inhibition, and acceptable pharmacokinetic properties. Moreover, 12 ameliorated Aß25-35-induced neurotoxicity in SH-SY5Y cells and alleviated the depressive symptom in tail suspension test. In brief, these results indicated that 12 is a prospective prototype for the potential treatment of AD with comorbid depression.

Identification of Guaifenesin-Andrographolide as a Novel Combinatorial Drug Therapy for Epilepsy Using Network Virtual Screening and Experimental Validation.

Combinatorial drug therapy has attracted substantial attention as an emerging strategy for the treatment of diseases with complex pathological mechanisms. We previously developed a potentially universal computational screening approach for combination drugs and used this approach to successfully identify some beneficial combinations for the treatment of heart failure. Herein, this screening approach was used to identify novel combination drugs for the treatment of epilepsy in an approved drug library. The combination of guaifenesin-andrographolide was first discovered as a promising therapy with synergistic anticonvulsant activities in maximal electroshock (MES)- and subcutaneous pentylenetetrazol (sc-PTZ)-induced epilepsy models in vivo. The studies of network analysis, fluorescence imaging, and N-methyl-d-aspartate (NMDA)-induced cytotoxicity further revealed that guaifenesin-andrographolide might synergistically affect NMDA receptors and then alleviate the pathogenesis of epilepsy. Therefore, we report that the combination of guaifenesin-andrographolide exerts effects against epilepsy through a novel synergistic mechanism and is thus a potential treatment for epilepsy, providing a promising mechanism for the design of novel combinatorial drug treatments against epilepsy.

Development of a TCR-like antibody and chimeric antigen receptor against NY-ESO-1/HLA-A2 for cancer immunotherapy.

BACKGROUND: The current therapeutic antibodies and chimeric antigen receptor (CAR) T cells are capable of recognizing surface antigens, but not of intracellular proteins, thus limiting the target coverage for drug development. To mimic the feature of T-cell receptor (TCR) that recognizes the complex of major histocompatibility class I and peptide on the cell surface derived from the processed intracellular antigen, we used NY-ESO-1, a cancer-testis antigen, to develop a TCR-like fully human IgG1 antibody and its derivative, CAR-T cells, for cancer immunotherapy. METHODS: Human single-chain variable antibody fragment (scFv) phage library (~10∧11) was screened against HLA-A2/NY-ESO-1 (peptide 157-165) complex to obtain target-specific antibodies. The specificity and affinity of those antibodies were characterized by flow cytometry, ELISA, biolayer interferometry, and confocal imaging. The biological functions of CAR-T cells were evaluated against target tumor cells in vitro. In vivo antitumor activity was investigated in a triple-negative breast cancer (TNBC) model and primary melanoma tumor model in immunocompromised mice. RESULTS: Monoclonal antibody 2D2 identified from phage-displayed library specifically bound to NY-ESO-1157-165 in the context of human leukocyte antigen HLA-A*02:01 but not to non-A2 or NY-ESO-1 negative cells. The second-generation CAR-T cells engineered from 2D2 specifically recognized and eliminated A2+/NY-ESO-1+tumor cells in vitro, inhibited tumor growth, and prolonged the overall survival of mice in TNBC and primary melanoma tumor model in vivo. CONCLUSIONS: This study showed the specificity of the antibody identified from human scFv phage library and demonstrated the potential antitumor activity by TCR-like CAR-T cells both in vitro and in vivo, warranting further preclinical and clinical evaluation of the TCR-like antibody in patients. The generation of TCR-like antibody and its CAR-T cells provides the state-of-the-art platform and proof-of-concept validation to broaden the scope of target antigen recognition and sheds light on the development of novel therapeutics for cancer immunotherapy.

Performance of alternative drying techniques on hop (Humulus lupulus L.) aroma quality: An HS-SPME-GC-MS-O and chemometrics combined approach.

Economically feasible and effective hop drying strategies are urgently needed to respond to the increasing number of microbrewers in US. In this study, hops were dried by dehydrator-drying (52 °C), oven-drying (52 °C) and freeze-drying (25 °C) until the final moisture content reached 8-10%. Headspace solid-phase microextraction-gas chromatography-mass spectrometry-olfactometry (HS-SPME-GC-MS-O) was employed to analyze the aroma profiles in all dried hops. Methyl octanoate, ß-myrcene, trans-α-bergamotene, linalool and geraniol were perceived as high-intensity aromas in all samples. Generally, dehydrator-dried hops contained the highest contents of aroma compounds among all groups, showing an increase of 5-23% and 6-37% when compared to freeze- and oven-dried hops, respectively. Principal component and hierarchical cluster analyses also revealed aroma content differences from three drying methods. Dehydrator drying at 52 °C was therefore considered as an alternative and promising drying approach for smaller-scale hop processing, which can largely benefit regional producers and local craft breweries.

Common Genomic Aberrations in Mouse and Human Breast Cancers with Concurrent P53 Deficiency and Activated PTEN-PI3K-AKT Pathway.

Simultaneous P53 loss and activation of the PTEN-restricted PI3K-AKT pathway frequently occur in aggressive breast cancers. P53 loss causes genome instability, while PTEN loss and/or activating mutations of PIK3CA and AKT promote cancer cell proliferation that also increases incidences of genomic aberrations. However, the genomic alterations associated with P53 loss and activated PTEN-PI3K-AKT signaling in breast cancer have not been defined. Spatiotemporally controlled breast cancer models with inactivation of both P53 and Pten in adult mice have not been established for studying genomic alterations. Herein, we deleted both floxed Pten and Tp53 genes in the mammary gland epithelial cells in adult mice using a RCAS virus-mediated Cre-expressing system. These mice developed small tumors in 21 weeks, and poorly differentiated larger tumors in 26 weeks. In these tumors, we identified 360 genes mutated by nonsynonymous point mutations and small insertions and deletions (NSPMs/InDels), 435 genes altered by copy number amplifications (CNAs), and 450 genes inactivated by copy number deletions (CNDs). Importantly, 22.2%, 75.9% and 27.3% of these genes were also altered in human breast tumors with P53 and PTEN losses or P53 loss and activated PI3K-AKT signaling by NSPMs/InDels, CNAs and CNDs, respectively. Therefore, inactivation of P53 and Pten in adult mice causes rapid-growing breast tumors, and these tumors recapitulate a significant number of genetic aberrations in human breast tumors with inactivated P53 and activated PTEN-PI3K-AKT signaling. Further characterization of these commonly altered genes in breast cancer should help to identify novel cancer-driving genes and molecular targets for developing therapeutics.

Rapid Repurposing of Novel Combination Drugs for the Treatment of Heart Failure via a Computationally Guided Network Screening Approach.

Combination drugs, characterized by high efficacy and few side effects, have received extensive attention from pharmaceutical companies and researchers for the treatment of complex diseases such as heart failure (HF). Traditional combination drug discovery depends on large-scale high-throughput experimental approaches that are time-consuming and costly. Herein we developed a novel, rapid, and potentially universal computer-guided combination drug-network-screening approach based on a set of databases and web services that are easy for individuals to obtain and operate, and we discovered for the first time that the menthol-allethrin combination screened by this approach exhibited a significant synergistic cardioprotective effect in vitro. Further mechanistic studies indicated that allethrin and menthol could synergistically block calcium channels. Allethrin bound to the central cavity of the voltage-dependent L-type calcium channel subunit alpha-1S (CACNA1S) lead to a conformational change in an allosteric site of CACNA1S, thereby enhancing the binding of menthol to this allosteric site. In summary, we reported a potentially universal computational approach to combination drug screening that has been used to discover a new combination of menthol-allethrin against HF in vitro, providing a new synergistic mechanism and prospective agent for HF treatment.