Heterozygous mutations in IDH1 (isocitrate dehydrogenase 1) are found in most grade II and III brain tumors. A slew of mutant IDH1 inhibitors were identified soon after the discovery of IDH1 mutations in brain tumors. But recent reports show that mutant IDH1 inhibitors reverse therapeutic vulnerabilities and activate the oncogenic transcription factor STAT3 in mutant IDH1-expressing cells. Thus, inhibiting mutant IDH1 using mutant IDH1-specific inhibitors can result in drug resistance. Therefore, to block mutant IDH1, it is imperative to identify alternative modes of therapy. In these lines, recent findings show that PROteolysis TArgeting Chimera (PROTAC) molecules can be designed to degrade target proteins in cancer cells. However, it is unknown whether degrading mutant IDH1 leads to STAT3 activation. Therefore, in this study, we asked if degrading mutant IDH1 by employing a PROTAC-based approach leads to STAT3 activation. To answer the question, we adopted the dTAG system, where we fused FKBP12 to mutant IDH1 proteins and used the FKBP12-specific PROTAC, dTAG-13, to degrade mutant IDH1-FKBP12. We assessed STAT3 activation in dTAG-13-treated cells expressing mutant IDH1-FKBP12. We found that fusing FKBP12-HA to mutant IDH1 phenocopies mutant IDH1 with similar expression levels, enzyme activity, and cellular localization. We observed that dTAG-13 degrades mutant IDH1-FKBP12-HA in a dose- and time-responsive manner. Unlike inhibiting, degrading mutant IDH1-FKBP12-HA did not lead to pSTAT3-Y705 activation. We conclude that degrading mutant IDH1 by employing a PROTAC-based approach impairs STAT3 activation. Based on these observations, we suggest that mutant IDH1-specific PROTACs can be developed to degrade mutant IDH1 in gliomas.

Bacterial luciferase (LuxAB) catalyzes the conversion of reduced flavin mononucleotide (FMNH⁻), oxygen, and a long-chain aldehyde to oxidized FMN, the corresponding acid and water with concomitant light emission. This bioluminescence reaction requires the reaction of a flavin reductase such as LuxG (in vivo partner of LuxAB) to supply FMNH⁻ for the LuxAB reaction. LuxAB is a well-known self-sufficient luciferase system because both aldehyde and FMNH⁻ substrates can be produced by the associated enzymes encoded by the genes in the lux operon, allowing the system to be auto-luminous. This makes it useful for in vivo applications. Structural and functional studies have long been performed in efforts to gain a better understanding of the LuxAB reaction. Recently, continued exploration of the LuxAB reaction have elucidated the mechanisms of C4a-hydroperoxyflavin formation and identified key catalytic residues such as His44 that facilitates the generation of flavin intermediates important for light generation. Advancements in protein engineering and synthetic biology have improved the bioluminescence properties of LuxAB. Various applications of LuxAB for bioimaging, bioreporters, biosensing in metabolic engineering and real-time monitoring of aldehyde metabolites in biofuel production pathways have been developed during the last decade. Challenging issues such as achieving red-shifted emissions, optimizing the signal intensity and identifying mechanisms related to the generation of light-emitting species remain to be explored. Nevertheless, LuxAB continues to be a promising tool for diverse biotechnological and biomedical applications.

Acer tegmentosum Maxim (AT) has a variety of pharmacological activities, however, the effects of AT on liver injury and gut microbiota in alcoholic liver disease (ALD) mice is still unclear. This study aimed to evaluate the preventive effect of AT extract on acute alcoholic liver disease. Six-week-old male C57BL/6J mice were randomly divided into 6 groups. Each group was intragastrically treated saline or different concentration of AT extract solution for 5 weeks continuously. After the last gavage, except for the NC group, all the other groups were gavaged twice with 56% alcohol to establish the acute ALD model and biochemical indexes, histopathological, and gut microbiota were analyzed. Established an acute ALD mouse model and detected serum, liver oxidation levels, and alcohol metabolism-related gene expressions. Through 16S rRNA sequencing, analyzed gut microbiota, explored the relationship between gut microbiota and liver indicators. AT extract significantly decreased lipid levels, promoted ADH, ALDH, and increased the antioxidant activities. Meanwhile, AT extract significantly downregulated the expression of lipid oxidation and inflammatory factors, upregulated alcohol metabolism genes. In addition, 16S rRNA sequencing and analysis showed that AT extract effectively regulated the gut microbiota diversity of ALD mice, significantly improved the structural disturbance of intestinal microflora. AT extract regulated gut microbiota and had a strong correlation with serum, liver-related indexes, and gene expression levels. All these results showed that AT can alleviate alcohol induced liver injury by regulating oxidative stress, inflammatory response, alcohol metabolism, and gut microbiota disorder.

Non-alcoholic steatohepatitis (NASH) is the progressive form of non-alcoholic fatty liver disease (NAFLD) which is the most common chronic liver disease worldwide. Hypoxia-inducible factor-1α (HIF1α) inhibitor is emerging as a promising therapeutic strategy for diseases. However, the role of HIF1α inhibitor in NASH is still unclear. A choline-deficient, l-amino acid-defined, high-fat diet (CDAHFD) -induced NASH mouse model was established to identify the impacts of HIF1α inhibitor KC7F2 on the development of NASH. We found that KC7F2 treatment substantially aggravated lipid accumulation, inflammation, and fibrosis in the liver of NASH mice presumably via increasing Tsukushi (TSKU) expression in the liver. Mechanistically, KC7F2 up-regulated expression of TSKU in hepatocyte in vitro, which led to increased hepatocellular lipid accumulation and was reversed when TSKU was knockdown in hepatocyte. Our findings indicated that HIF1α inhibitor promotes the development of NASH presumably via increasing TSKU expression in the liver, suggesting that HIF1α attenuates NASH, and that we should assess the potential liver toxicity when use HIF1α inhibitor or medicines that can decrease the expression of HIF1α to therapy other diseases.

Polarization of microglia following spinal cord injury (SCI) is a pivotal pathological process of secondary injury. Although differentiation antagonistic nonprotein coding RNA (DANCR) has been implicated in immune and inflammatory responses across various diseases, its role in SCI still unclear. This research aimed to clarify the underlying mechanisms of DANCR in SCI recovery by investigating its expression pattern in microglia. Our findings indicate that the DANCR level in microglia is increased after SCI and that its knockdown can promote microglial M2-type polarization; suppress inflammatory cytokines, oxidative stress, and neuronal apoptosis; and facilitate nerve regeneration as well as spinal cord functional recovery. Further investigations suggest that DANCR's effects are mediated through the ACTN4/STAT3 axis. These results provide potential targets for enhancing functional recovery following SCI.

Human ventriculum myosin (βmys) powers contraction sometimes in complex with myosin binding protein C (MYBPC3). The latter regulates βmys activity and impacts cardiac function. Single residue variants (SRVs) change protein sequence in βmys or MYBPC3 causing inheritable heart diseases by affecting the βmys/MYBPC3 complex. Muscle genetics encode instructions for contraction informing native protein construction, functional integration, and inheritable disease impairment. A digital model decodes these instructions and evolves by processing new information content from diverse data modalities using a human partner-driven virtuous cycle optimization.

Ischemia-reperfusion injury (IRI) often results in renal impairment. While the presence of neutrophil extracellular traps (NETs) is consistently observed, their specific impact on IRI is not yet defined. Sivelestat sodium, an inhibitor of neutrophil elastase which is crucial for NET formation, may offer a therapeutic approach to renal IRI, warranting further research.

Acyl thiourea scaffolds are frequently employed in drug development to discern unique and essential therapies for the eradication of the most challenging diseases. Hence, we developed a library of novel cyclopropyl incorporating acyl thiourea derivatives (4a-j) and evaluated their antimicrobial, α-amylase, and proteinase K inhibition potential. Compound (4h) (4-methoxy) demonstrated the strongest α-amylase inhibition (IC = 1.572 ± 0.017 μM), while compound (4j) (3,4,5-trimethoxy) exhibited potent proteinase K inhibition (IC = 1.718 ± 0.061 μM), comparable to the standard acarbose (IC = 1.063 ± 0.013 μM) and phenyl methyl sulfonyl fluoride (IC = 0.119 ± 0.014 μM). The unsubstituted compound (4a) emerged as the most potent antifungal agent (17 mm zone of inhibition), outperforming the positive control Terbinafine (zone of inhibition 16 mm). These compounds (4a-j) also displayed moderate antibacterial activity. SAR analysis revealed the influences of various substitutions on the acyl thiourea scaffold. Computational studies, including DFT, molecular docking, and ADMET predictions, supported the biological findings and identified these compounds as promising inhibitors of α-amylase, proteinase K, and microbial pathogens.

Diffuse large B-cell lymphoma (DLBCL) is a prevalent and aggressive form of non-Hodgkin's lymphoma with a complex etiology. NOP2/Sun domain 2 (NSUN2) is an RNA methyltransferase that has been linked to the regulation of gene expression in various cancers. However, the function of NSUN2 in DLBCL, specifically its contribution to exosome-driven tumor progression, remains to be thoroughly elucidated.

The aim of the current study was to investigate the potential therapeutic effect of kaurenoic acid (KA) against Monosodium Urate Crystals (MSU)-induced acute gout by downregulation of NF-κB signaling pathway, mitigating inflammation and oxidative stress. KA potentially targeted NF-κB pathway activation and provided comprehensive insights through multiple approaches. This was accomplished by advanced analytical techniques. This methodology highlighted the efficacy of KA in acute gout attacks offering new approach for gout management.

Cancer is among the leading causes of death worldwide. The effectiveness of conventional chemotherapy has some drawbacks, therefore, there is an urgency to develop novel strategies to fight this disease. Ornithine decarboxylase (ODC) is the most finely tuned enzyme of the polyamine (PA) biosynthesis pathway as it is regulated at different levels: transcriptional, translational, post-translational, and by feedback inhibition. In cancer, this enzyme is overexpressed due to its regulation by the protooncogene c-Myc, thus it has been proposed as a drug target against this disease. This review describes information regarding the biochemistry and regulation of ODC at different levels and its role in cancer. Moreover, we discuss the molecules aiming on the inhibition of the ODC activity that have been tested as therapeutic options. ODC remains as a therapeutic opportunity that needs to be more explored.

The mitochondrial flavoenzymes proline dehydrogenase (PRODH) and hydroxyproline dehydrogenase (PRODH2) catalyze the first steps of proline and hydroxyproline catabolism, respectively. The enzymes are targets for chemical probe development because of their roles in cancer cell metabolism (PRODH) and primary hyperoxaluria (PRODH2). Mechanism-based inactivators of PRODH target the FAD by covalently modifying the N5 atom, with N-propargylglycine (NPPG) being the current best-in-class of this type of probe. Here we investigated a close analog of NPPG, but-3-yn-2-ylglycine (B32G), distinguished by having a methyl group adjacent to the ethynyl group of the propargyl warhead. UV-visible spectroscopy shows that a bacterial PRODH catalyzes the oxidation of the S-enantiomer of B32G, a necessary first step in mechanism-based inactivation. In contrast, the enzyme does not react with the R-enantiomer. Enzyme activity assays show that S-B32G inhibits bacterial PRODH in a time-dependent manner consistent with covalent inactivation; however, the inactivation efficiency is ∼600-times lower than NPPG. We generated the crystal structure of PRODH inactivated by S-B32G at 1.68 Å resolution and found that inactivation induces a covalent link between the FAD N5 and the ε-nitrogen of an active site lysine, confirming that S-B32G follows the same mechanism as NPPG. Despite its lower inactivation efficiency at the purified bacterial enzyme, S-B32G exhibited comparable activity to NPPG against PRODH and PRODH2 in human cells and mouse livers. Molecular modeling is used to rationalize the stereospecificity of B32G.

4-Phenol oxidases are proposed to be involved in the utilization of lignin-derived aromatic compounds. While enzymes with selectivity towards 4-hydroxyphenyl and guaiacyl motifs are well described, we identified the first syringyl-specific oxidase from Streptomyces cavernae (Sc4ASO) only very recently. Here, in-depth studies were conducted to unravel the molecular origins of the outstanding selectivity of Sc4ASO. Kinetic experiments revealed high activities on dimethoxylated substrates (up to 2.9 ± 0.1 s), but also strong cooperativity between both protein subunits, as well as substrate inhibition in dependency of ortho methoxylation and chain length of the para substituent. Rapid mixing kinetics in combination with the determination of the crystal structure in complex with three substrates allowed to connect the kinetic behavior with never-observed positioning of the conserved residue Y471. Ultimately, the catalytic potential of Sc4ASO was investigated in a 100 mL scale cascade reaction to produce the natural product syringaresinol.

Hepatocellular carcinoma (HCC) is one of the most lethal malignancies worldwide and the most common form of liver cancer. Despite global efforts toward early diagnosis and effective treatments, HCC is often diagnosed at advanced stages, where conventional therapies frequently lead to resistance and/or high recurrence rates. Therefore, novel biomarkers and promising medications are urgently required. Epi-drugs, or epigenetic-based medicines, have recently emerged as a promising therapeutic modality. Since the epigenome of the cancer cells is always dysregulated and this is followed by apoptosis-resistance, reprogramming the epigenome of cancer cells by epi-drugs (such as HDAC inhibitors (HDACis), and DNMT inhibitors (DNMTis)) could be an alternative approach to use in concert with established treatment protocols. C-FLIP, an anti-apoptotic protein, and Ku70, a member of the DNA repair system, bind together and make a cytoplasmic complex in certain cancers and induce resistance to apoptosis. Many epi-drugs, such as HDACis, can dissociate this complex through Ku70 acetylation and activate cellular apoptosis. The novel compounds for dissociating this complex could provide an innovative insight into molecular targeted HCC treatments. In this review, we address the innovative therapeutic potential of targeting c-FLIP/Ku70 complex by epi-drugs, particularly HDACis, to overcome apoptosis resistance of HCC cells. This review will cover the mechanisms by which the c-FLIP/Ku70 complex facilitates cancer cell survival, the impact of epigenetic alterations on the complex dissociation, and highlight HDACis potential in combination therapies, biomarker developments and mechanistic overviews. This review highlights c-FLIP ubiquitination and Ku70 acetylation levels as diagnostic and prognostic tools in HCC management.

Astaxanthin (ASX), a fat-soluble carotenoid mainly sourced from Haematococcus pluvialis, shows promise for clinical applications in chronic inflammatory diseases. This study investigates whether ASX can mitigate atherosclerosis (AS) by modulating macrophage ferroptosis and provides astaxanthin-loaded polylactic acid-glycolic acid nanoparticles (ASX-PLGA NPs) as comparison.

Yes-associated protein (YAP), a focal point of current biological research, is involved in regulating various life processes. In this report, live-cell fluorescence resonance energy transfer (FRET) imaging was employed to unravel the YAP complexes in MCF-7 cells. Fluorescence imaging of living cells co-expressing CFP (cyan fluorescent protein)-YAP and YFP (yellow fluorescent protein)-LATS1 (large tumor suppressor 1) plasmids revealed that YAP promoted LATS1 oligomerization around mitochondria. Moreover, FRET two-hybrid assay showed that YAP directly interacted with LATS1 to form dimer. Similarly, we found that YAP directly interacted with large tumor suppressor 2 (LATS2) to form a heterotrimer with 1:2 in cytoplasm and around mitochondria. In addition, YAP directly interacted with angiomotin (AMOT) to form a heterodimer in cytoplasm. However, YAP did not interact with O-linked N-acetylglucosamine transferase (OGT). Furthermore, FRET assay also indicated that YAP exhibited a higher affinity with AMOT, followed by LATS1, and least with LATS2. In summary, YAP directly interacts with LATS1 and AMOT to form a heterodimer, with LATS2 to form a heterotrimer with 1:2, and shows a preference for binding to AMOT, followed by LATS1, and lastly LATS2, providing new insights into the Hippo-YAP signaling pathway.

Diabetes Mellitus (DM), one of the oldest known metabolic disorders, dates back to 3000 BC and continues to have a profound impact on health and the economy. Nutrition plays a critical role in managing diabetes and enhancing overall quality of life. It is also vital for immune system function, as well as in the prevention and treatment of aging-related diseases. A key factor contributing to the global rise in obesity is the excessive consumption of fructose/glucose (corn) syrup, which leads to various metabolic complications. Uncontrolled intake of carbohydrates, particularly sugars like fructose, triggers the Maillard Reaction, a chemical process that occurs between sugars and proteins, resulting in advanced glycation end-products (AGEs). This process is accelerated in diabetic patients due to hyperglycemia, leading to increased glycation of plasma proteins such as immunoglobulins, which play an essential role in the immune system. Studies show that individuals with Diabetes Mellitus experience a higher susceptibility to infections due to increased viral entry, impaired immune responses, reduced viral clearance, and dysregulated inflammatory cytokine production. In this study, human IgG proteins were glycated in vitro using fructose, simulating the damaging effects seen in diabetic conditions. A mixture containing antioxidants like glutathione, oleuropein, and selenium was prepared and incubated with the glycated IgG to assess its protective properties. Lymphocyte cells from healthy volunteers were also treated with fructose and subjected to similar experiments. Results demonstrated that fructose significantly compromises immune function by damaging key proteins, but the antioxidant mixture effectively mitigates this damage, offering a protective mechanism against glycation in the immune system.

Our previous study revealed that lipid flip-flop inducing phytochemicals from Gymnema sylvestre increase membrane permeability of antimicrobials in S. aureus. However, their lipid flipping and membrane permeabilizing effect on methicillin resistant S. aureus (MRSA) membrane that has intrinsically higher aminoacylated lipid content compared to methicillin sensitive S. aureus (MSSA) is poorly characterized. Gymnema saponins, gymnemic acid I and IV significantly increased the antibiotic susceptibility in both MSSA and MRSA. MRSA exhibited a rigid membrane with lipid diffusion coefficient 0.0002 μm/s compared to the MSSA membrane lipids with diffusion coefficient 1.48 μm/s. Further, unlike MSSA, MRSA cells inhibited fusion of fluid liposomes with their plasma membrane. In vitro assay on reconstituted membrane vesicles revealed that Gymnema saponins induced 60 % lipid flipping in MSSA membrane compared to only 20 % lipid flipping in MRSA, indicating significantly lower Gymnema saponin-induced trans-bilayer lipid mobility in MRSA. Gymnema saponins induced significantly lower crystal violet uptake, release of cellular protein, cell shrinkage and lysis in MRSA compared to MSSA. Gymnema saponins led to dose-dependent inhibition of lipid-aminoacylation in both MSSA and MRSA making their membranes more negative compared to untreated control cells. In silico analysis reveals binding of both gymnemic acid I and IV to multiple peptide resistance factor (binding energy ∼ 7.5 kCal), the protein responsible for lipid aminoacylation in S. aureus. For the first time, our study reveals that MRSA membrane with higher aminoacyl-PG compared to MSSA shows significantly lower rate of diffusion and trans-bilayer flip-flop of lipids. Further, gymnemic acids are useful probes for identification, characterization and drug sensitization of rigid membrane MRSA phenotypes.

Breast cancer is one of the most common cancers found in women worldwide. Besides the availability of clinical drugs, drug resistance and considerable side effects are concerning issues driven the needs for the discovery of novel anticancer agents. Aromatase inhibition is one of the effective strategies for management of hormone-dependent breast cancer. Triazole, coumarin, and isatin are heterocyclic scaffolds holding great attention in the field of drug design. Molecular hybridization is a well-known strategy to achieve new molecules with improved potency and properties. Herein, a set of 27 triazole-based hybrids (i.e., coumarin-triazoles series 5-6 and isatin-triazoles series 7) were synthesized and investigated for their anti-proliferation, apoptosis induction, and aromatase inhibitory potentials. Anti-proliferative study against the hormone-dependent breast cancer (T47D) cell line indicated that coumarin-triazoles 5h (R=NO) and 6i (R=SONH) were the two most potent antiproliferative agents. Particularly, compound 5h showed comparable potency and superior selectivity index than that of the reference drug, doxorubicin. Moreover, the coumarin-triazole 5h induced cellular apoptosis of the estrogen-dependent breast cancer (MCF-7) cells. Additionally, findings from the aromatase inhibitory assay suggested four compounds as potential aromatase inhibitors (i.e., 5i, 6f, 6g and 6i, IC = 1.4-2.4 μM). Two QSAR models with preferable predictive performances were constructed to reveal key properties influencing antiproliferative and aromatase inhibitory effects. Molecular docking was conducted to elucidate the possible binding modalities against the target aromatase enzyme. Key structural features essential for the binding were highlighted. Moreover, the drug-like properties of top-ranking compounds were assessed to ensure their possibilities for successful development.

The possibility of using an oxygen-nitrous oxide mixture for prolonged hypothermic preservation of rat heart for 24 h was investigated. A comparative analysis of restoration of functional activity of hearts in the groups of 24-h preservation at +4 °C with different gases (O, N) and gas mixtures (CO + O, NO + O, N+O, NO + N) was carried out. It was shown that the presence of oxygen in the gas mixture was the key factor for heart preservation. No stable heart preservation was observed in oxygen-free mixtures. At the same time, preservation in pure oxygen showed a significantly lower level of cardiac recovery compared to preservation in gas mixtures O+CO (6.5 atm.) and O+NO (6.5 atm.). LVDP (left ventricular developed pressure) values were 30 ± 19 mmHg and 46 ± 9 mmHg, respectively, with no significant differences found. The decrease in LDVP after 24 h of storage was 26-40 % of the intact control. The results obtained indicate the presence of pronounced synergistic effects of both gases during 24-h heart preservation, which is confirmed by data of marker genes Nfe2l2, Nox1, Prdx1, Hif1a, Nos2, Slc2a4, Ucp-1, Jun, Casp3 expression analysis and myocardial infarction damage level data. The more frequent occurrence of arrhythmias was observed in the oxygen-nitrous oxide group compared with the CO group, and the mechanism of this phenomenon is unclear. Nevertheless, the already medically approved NO + O gas mixture could serve as a balanced choice for future improvements, offering a shorter duration of cardiac preservation compared to the CO + O mixture, while ensuring safety in its use.

In the presented study, we evaluated changes in the molecular structures of lipids and proteins in organs/tissues at the early stage of obesity induced by a high-calorie diet (HCD), using animal models. We examined several different molecular parameters and the organs most affected by obesity. Fourier transform infrared (FTIR) spectroscopy combined with Principal Component Analysis (PCA) and Receiver Operating Characteristic (ROC) analysis were used to evaluate molecular changes in tissues taken from HCD-induced obese Wistar rats and their lean counterparts. We observed that at the early stage of obesity, changes occurred mainly in lipid structures, primarily affecting white epididymal adipose tissue (WAT) and the liver (Lr). No changes in protein molecular structures were observed in any of the examined organs. PCA showed distinctly different organ/tissue compositions, in terms of molecular parameters, for both groups. In turn, ROC analysis indicated that fatty acid chain length (FACL), lipid unsaturation (L_Unsat), and carbonyl/lipid ratio (Carb/L) for WAT, and FACL and lipid/protein ratio (L/P) for Lr, were the molecular parameters, whose levels differentiated the most between both groups. We demonstrated that studies using FTIR spectroscopy combined with advanced data mining methods could deepen the current knowledge about obesity and the biochemical changes occurring in the organs affected by this disease. Thus, they can help in the future with better and faster diagnosis and prevention of obesity and its complications.