Comprehensive pan-cancer analysis of KLRB1-CLEC2D pair and identification of small molecule inhibitors to disrupt their interaction.

The interplay between immune checkpoints KLRB1 and CLEC2D is crucial for tumor progression and immune evasion, yet the interaction dynamics are not fully understood. This study aims to elucidate the interaction across various cancers and identify small molecule inhibitors that can disrupt it. We perform a comprehensive pan-cancer analysis of the KLRB1-CLEC2D pair, including mRNA expression patterns, pathological stages, survival outcomes, and single-cell omics, immune infiltration, copy number variations, and DNA methylation profiles. Our findings reveal a consistently higher CLEC2D/KLRB1 ratio in most cancer types compared to normal tissues, and this ratio also increased with advancing pathological stages. Lower KLRB1 expression correlated with higher mortality in most cancers, opposite to CLEC2D. Expression variations were attributed to differential lymphocyte infiltration, CNV, and DNA methylation. Structure-based virtual screening analysis identified compounds including forsythiaside A and RGD peptides as effective inhibitors of the KLRB1-CLEC2D interaction, validated through microscale thermophoresis. This research advances understanding of the KLRB1-CLEC2D interaction within the tumor microenvironment and introduces novel therapeutic strategies to modulate this interaction.

Chirality-driven strong thioredoxin reductase inhibition.

Overexpression of thioredoxin reductase (TXNRD) plays crucial role in tumorigenesis. Therefore, designing TXNRD inhibitors is a promising strategy for targeted anticancer drug development. However, poor selectivity has always been a challenge, resulting in unavoidable toxicity in clinic. Herein we demonstrate a strategy to develop highly selective chiral metal complexes-based TXNRD inhibitors. By manipulating the conformation of two distinct weakly interacting groups, we optimize the compatibility between the drug and the electrophilic group within the active site of TXNRD to enhance their non-covalent interaction, thus effectively avoids the poor selectivity deriving from covalent drug interaction, on the basis of ensuring the strong inhibition. Detailed experimental and computational results demonstrate that the chiral isomeric drugs bind to the active site of TXNRD, and the interaction strength is well modulated by chirality. Especially, the meso-configuration, in which the two large sterically hindered active groups are positioned on opposite sides of the drug, exhibits the highest number of non-covalent interactions and most effective inhibition on TXNRD. Taken together, this work not only provides a novel approach for developing highly selective proteinase inhibitors, but also sheds light on possible underlying mechanisms for future application.

A Validation of the Equivalence of the Cell-Based Potency Assay Method with a Mouse LD50 Bioassay for the Potency Testing of OnabotulinumtoxinA.

(1) Background: At present, the only potency assay approved in China for the in-country testing of botulinum toxin type A for injection products is the mouse bioassay (MBA). The Chinese market for neurotoxin products is rapidly expanding, but MBAs are subject to high variability due to individual variations in mice, as well as variations in injection sites, in addition to the limited number of batches tested for one MBA. Compared with the mLD50 method, the cell-based potency assay (CBPA) developed for the potency testing of onabotulinumtoxinA (BOTOX) by AbbVie not only does not use any experimental animals but also allows for significant time and cost savings. Due to the significant benefits conferred by the replacement of the mLD50 assay with CBPA in China, the CBPA method has been transferred, validated, and cross-validated to demonstrate the equivalence of the two potency methods. (2) Methods: The differentiated SiMa cells were treated with both BOTOX samples and the reference standard, and the cleaved SNAP25197 in the cell lysates was quantified using Chemi-ECL ELISA. A 4-PL model was used for the data fit and sample relative potency calculation. The method accuracy, linearity, repeatability, and intermediate precision were determined within the range of 50% to 200% of the labeled claim. A statistical equivalence of the two potency methods (CBPA and mLD50) was initially demonstrated by comparing the AbbVie CBPA data with NIFDC mLD50 data on a total of 167 commercial BOTOX lots (85 50U lots and 82 100U lots). In addition, six lots of onabotulinumtoxinA (three 50U and three 100U) were re-tested as cross-validation by these two methods for equivalence. (3) Results: The overall assay's accuracy and intermediate precision were determined as 104% and 9.2%, and the slope, R-square, and Y-intercept for linearity were determined as 1.071, 0.998, and 0.036, respectively. The repeatability was determined as 6.9%. The range with the acceptable criteria of accuracy, linearity, and precision was demonstrated as 50% to 200% of the labeled claim. The 95% equivalence statistic test using margins [80%, 125%] indicates that CBPA and mLD50 methods are equivalent for both BOTOX strengths (i.e., 50U and 100U). The relative potency data from cross-validation were within the range of ≥80% to ≤120%. (4) Conclusions: The CBPA meets all acceptance criteria and is equivalent to mLD50. The replacement of mLD50 with CBPA is well justified in terms of ensuring safety and efficacy, as well as for animal benefits.

The complete chloroplast genome of the medicinally important plant Plumbago zeylanica L. (plumbaginaceae) and phylogenetic analysis.

Plumbago zeylanica L. 1753 is a medicinally-important herb in family Plumbaginaceae. In this study, we assembled and reported the complete chloroplast genome of P. zeylanica. The plastome of P. zeylanica was 169,178 bp, including a large single-copy region of 92,135 bp, a small single-copy region (SSC) of 13,455 bp and a pair of inverted repeat regions (IRs) of 31,794 bp. It contained 124 genes, including 79 protein-coding genes, 37 tRNA genes and eight rRNA genes. Phylogenetic analysis showed that P. zeylanica formed a close relationship with P. auriculata in Plumbago. The first complete chloroplast genome report of P. zeylanica providing an opportunity to explore the genetic diversity, and would be also helpful in the species identification and conservation.

MEF-UNet: An end-to-end ultrasound image segmentation algorithm based on multi-scale feature extraction and fusion.

Ultrasound image segmentation is a challenging task due to the complexity of lesion types, fuzzy boundaries, and low-contrast images along with the presence of noises and artifacts. To address these issues, we propose an end-to-end multi-scale feature extraction and fusion network (MEF-UNet) for the automatic segmentation of ultrasound images. Specifically, we first design a selective feature extraction encoder, including detail extraction stage and structure extraction stage, to precisely capture the edge details and overall shape features of the lesions. In order to enhance the representation capacity of contextual information, we develop a context information storage module in the skip-connection section, responsible for integrating information from adjacent two-layer feature maps. In addition, we design a multi-scale feature fusion module in the decoder section to merge feature maps with different scales. Experimental results indicate that our MEF-UNet can significantly improve the segmentation results in both quantitative analysis and visual effects.

Discovery of Pyroptosis-inducing Drugs and Antineoplastic Activity Based on the ROS/ER Stress/Pyroptosis Axis.

BACKGROUND: Pyroptosis, a cell death process triggered by chemotherapy drugs, has emerged as a highly promising mechanism for combating tumors in recent years. As the lead of new drugs, natural products play an important role in the discovery of anticancer drugs. Compared to other natural products, the medicine food homologous natural products (MFHNP) exhibit a superior safety profile. Among a series of MFHNP molecular skeletons, this study found that only benzylideneacetophenone (1) could induce cancer cell pyroptosis. However, the anti-cancer activity of 1 remains to be improved. AIMS: This study aimed to find a pyroptosis inducer with highly effective antitumor activity by modifying the chalcone structure. METHODS: To examine the effect of the Michael receptor in compound 1 on the induction of pyroptosis, several analogs were synthesized by modifying the Michael acceptor. Subsequently, the anticancer activity was tested by MTT assay, and morphological indications of pyroptosis were observed in human lung carcinoma NCI-H460 and human ovarian cancer CP-70 cell lines. Furthermore, to improve the activity of the chalcone skeleton, the anticancer group 3,4,5- trimethoxyphenyl was incorporated into the phenyl ring. Subsequently, compounds 2-22 were designed, synthesized, and screened in human lung cancer cells (NCI-H460, H1975, and A549). Additionally, a quantitative structure-activity relationship (QSAR) model was established using the eXtreme Gradient Boosting (XGBoost) machine learning library to identify the pharmacophore. Furthermore, both in vitro and in vivo experiments were conducted to investigate the molecular mechanisms of pyroptosis induced by the active compound. RESULTS: α, ß-unsaturated ketone was the functional group of the chalcone skeleton and played a pivotal role in inducing cancer cell pyroptosis. QSAR models showed that the regression coefficients (R2) were 0.992 (A549 cells), 0.990 (NCI-H460 cells), and 0.998 (H1975 cells). Among these compounds, compound 7 was selected to be the active compound. Moreover, compound 7 was found to induce pyroptosis in lung cancer cells by upregulating the expression of CHOP by increasing the ROS level. Furthermore, it effectively suppressed the growth of lung cancer xenograft tumors. CONCLUSION: Compound 7 exhibits antineoplastic activity by regulating the ROS/ER stress/pyroptosis axis and is a kind of promising pyroptosis inducer.

Comparative proteomic analysis of cerebral cortex revealed neuroprotective mechanism of esculentoside A on Alzheimer's disease.

Esculentoside A (EsA), isolated from phytolacca esculenta, is a saponin showing neuroprotective effect in the mouse models of Alzheimer's disease (AD). To investigate its action target and underlying mechanism, this study used the proteomics technique of isobaric tags for relative and absolute quantification (iTRAQ) to analyze the differentially expressed proteins (DEPs) in the cerebral cortex of EsA-treated and untreated triple-transgenic 3 × Tg-AD model mice. Proteomic comparison revealed 250, 436, and 903 DEPs in three group pairs, i.e. AD/Wild-type (WT), AD+5 mg/kg EsA/AD, AD+10 mg/kg EsA/AD, respectively. Among them 28 DEPs were commonly shared by three group pairs, and 25 of them showed reversed expression levels in the diseased group under the treatment of both doses of EsA. Bioinformatics analysis revealed that these DEPs were mainly linked to metabolism, synapses, apoptosis, learning and memory. EsA treatment restored the expression of these proteins, including amyloid precursor protein (APP), cathepsin B (Cstb), 4-aminobutyrate aminotransferase (Abat), 3-phosphoinositide-dependent protein kinase-1 (PDK1), carnitine palmitoyltransferase1 (Cpt1) and synaptotagmin 17 (Syt17), thereby ameliorated the spatial learning and memory of AD mice. Collectively, this study reveals for the first time the profound effect of EsA on the cerebral cortex of AD mice, which might be a potential therapeutic agent for the treatment of AD.