Molecular phylogeny reveals Varroa mites are not a separate family but a subfamily of Laelapidae.

Varroa mites, notorious for parasitizing honeybees, are generally classified as Varroidae. Their extremely modified morphologies and behaviors have led to debates regarding their phylogenetic position and classification as an independent family. In this study, two different datasets were employed to reconstruct the phylogenies of Varroa mites and related Laelapidae species: (1) 9257 bp from the whole 13 mitochondrial protein-coding genes of 24 taxa, (2) 3158 bp from 113 taxa using Sanger sequencing of four nuclear loci. Both mitochondrial and nuclear analyses consistently place Varroa mites within the Laelapidae. Here we propose to place Varroa mites in the subfamily Varroinae stat. nov., which represents a highly morphologically adapted group within the Laelapidae. Ancestral state reconstructions reveal that bee-associated lifestyles evolved independently at least three times within Laelapidae, with most phoretic traits originating from free-living ancestors. Our revised classification and evolutionary analyses will provide new insight into understanding the Varroa mites.

Clinical, cognitive, and functional characteristics of recent-onset psychosis with autistic features: A 2-year longitudinal study.

Though categorized as separate illnesses, schizophrenia and autism are known to exhibit shared characteristics. This study explored the distinctions in clinical, cognitive, and functional characteristics among individuals with recent-onset psychosis, considering the severity of their autistic symptoms, involving longitudinal examinations. We analyzed 671 patients with recent-onset psychosis from Korean Early Psychosis Cohort Study (KEPS), and used the PANSS Autism Severity Score (PAUSS) to categorize patient into 'autistic', 'moderate', and 'non-autistic' groups. The autistic group had the highest rate of schizophrenia diagnosis, and the lowest incidence of comorbid psychiatric disorders. Schizophrenia diagnosis predicted membership of the autistic group. More severe autistic symptoms correlated with worse overall symptoms and functional outcomes, which significantly predicted membership of the autistic group. Cognitive impairments and emotional recognition difficulties increased with the severity of autistic symptoms. 2-year longitudinal assessments demonstrated that group differences in autistic features and overall symptoms, and functional outcomes remained consistent, and membership of the autistic group significantly predicted symptomatic remission and functional recovery. In conclusion, the presence of autistic symptoms has a significant impact on the overall symptomatology and functional capabilities. They are enduring attributes rather than temporary state variables, and serve as a significant predictor for both symptomatic and functional recovery.

Ultrastable 3D Heterogeneous Integration via N-Heterocyclic Carbene Self-Assembled Nanolayers.

The commercialization of 3D heterogeneous integration through hybrid bonding has accelerated, and accordingly, Cu-polymer bonding has gained significant attention as a means of overcoming the limitations of conventional Cu-SiO2 hybrid bonding, offering high compatibility with other fabrication processes. Polymers offer robust bonding strength and a low dielectric constant, enabling high-speed signal transmission with high reliability, but suffer from low thermomechanical stability. Thermomechanical stability of polymers was not achieved previously because of thermal degradation and unstable anchoring. To overcome these limitations, wafer-scale Cu-polymer bonding via N-heterocyclic carbene (NHC) nanolayers was presented for 3D heterogeneous integration, affording ultrastable packing density, crystallinity, and thermal properties. NHC nanolayers were deposited on copper electrodes via electrochemical deposition, and wafer-scale 3D heterogeneous integration was achieved by adhesive bonding at 170 °C for 1 min. Ultrastable conductivity and thermomechanical properties were observed by the spatial mapping of conductivity, work function, and force-distance curves. With regard to the characterization of NHC nanolayers, low-temperature bonding, robust corrosion inhibition, enhanced electrical conductivity, back-end-of-line process compatibility, and fabrication process reduction, NHC Cu/polymer bonding provides versatile advances in 3D heterogeneous integration, indicating that NHC Cu/polymer bonding can be utilized as a platform for future 3D vertical chip architectures.

Diabetic Ketoacidosis as an Effect of Sodium-Glucose Cotransporter 2 Inhibitor: Real World Insights.

One of the notable adverse effects of sodium-glucose cotransporter 2 (SGLT2) inhibitor is diabetic ketoacidosis (DKA) often characterized by euglycemia. In this retrospective review of patients with DKA from 2015 to 2023, 21 cases of SGLT2 inhibitorassociated DKA were identified. Twelve (57.1%) exhibited euglycemic DKA (euDKA) while nine (42.9%) had hyperglycemic DKA (hyDKA). More than 90% of these cases were patients with type 2 diabetes mellitus. Despite similar age, sex, body mass index, and diabetes duration, individuals with hyDKA showed poorer glycemic control and lower C-peptide levels compared with euDKA. Renal impairment and acidosis were worse in the hyDKA group, requiring hemodialysis in two patients. Approximately one-half of hyDKA patients had concurrent hyperosmolar hyperglycemic state. Common symptoms included nausea, vomiting, general weakness, and dyspnea. Seizure was the initial manifestation of DKA in two cases. Infection and volume depletion were major contributors, while carbohydrate restriction and inadequate insulin treatment also contributed to SGLT2 inhibitor-associated DKA. Despite their beneficial effects, clinicians should be vigilant for SGLT2 inhibitor risk associated with DKA.

Real-time analysis and prediction method of ion concentration using the effect of O-H stretching bands in aqueous solutions based on ATR-FTIR spectroscopy.

Analyzing the concentration of ions in aqueous solutions in real-time plays an important role in the fields of chemistry and biology. Traditional methods for measuring ion concentrations, such as concentration analysis by measuring electrical conductivity, inductively coupled plasma mass spectrometry, and ion chromatography, have been used in many research fields. However, these methods are limited in determining ion concentrations instantaneously. Fourier-transform infrared-attenuated total reflectance (ATR-FTIR) spectroscopy provides a new approach for determining ion concentrations in aqueous solutions. This allows for fast analysis without pretreatment and is scalable for real-time measurements. In this study, we present a method for measuring ion concentrations by examining ion-water interactions in the O-H stretching band of aqueous solutions using ATR-FTIR spectroscopy. Five aqueous solutions, namely LiCl + HCl, LiOH + HCl, LiOH, Li3PO4, and NaCl were used in the experiments and prepared at concentrations between 0.5-2 M. The ion concentrations in the prepared aqueous solutions were measured using ATR-FTIR spectroscopy. We observed that the difference in absorbance increased and decreased linearly with changes in concentration. The concentration of ions in the aqueous solution could be measured by validating the designed linear regression analysis function model. In this study, we proposed five linear regression analysis function models, all of which showed high coefficients of determination above 0.9, with the highest coefficient of determination reaching 0.9969. These results show that ATR-FTIR spectroscopy has the potential to be applied as a rapid and simple concentration analysis system.

Effects of statin use on serum creatinine phosphokinase levels in normal thyroid function.

BACKGROUND/AIMS: Statins are common lipid-lowering agents used in dyslipidemia. However, they increase serum creatinine phosphokinase (CPK) levels. Currently, there are no studies on the effect of thyroid-stimulating hormone (TSH) levels on CPK levels after statin administration. Therefore, this study aimed to investigate CPK level alterations after statin administration according to TSH quartiles in participants with euthyroidism. METHODS: This retrospective analysis included 25,047 patients with euthyroidism. CPK levels were measured before and 6 months after statin administration. Normal TSH levels were divided into four quartiles, and the CPK levels and proportions of patients with normal CPK levels after statin administration for each TSH quartile were evaluated. RESULTS: The baseline CPK level was significantly higher in the lowest TSH quartile (Q1) compared to the other quartiles but decreased after statin administration. Thus, the difference between the CPK levels and the other quartile groups was not significant. The proportion of patients with normal CPK levels was also significantly lowest in Q1 before statin administration; however, no significant difference was noted in the ratio among each group after statin administration. These findings were consistent with the findings of the analysis according to statin intensity. CONCLUSION: In patients in the lowest TSH quartile of the normal TSH range, the CPK level decreased, and the proportion of normal CPK levels increased significantly after statin administration. However, similar changes were not observed in other TSH quartiles. Therefore, further studies are required to mechanistically confirm these conclusions.

Quantitative Comparison of the Hydration Capacity of Surface-Bound Dextran and Polyethylene Glycol.

We have quantified and compared the hydration capacity (i.e., capability to incorporate water molecules) of the two surface-bound hydrophilic polymer chains, dextran (dex) and poly(ethylene glycol) (PEG), in the form of poly(l-lysine)-graft-dextran (PLL-g-dex) and poly(l-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG), respectively. The copolymers were attached to a negatively charged silica-titania surface through the electrostatic interaction between the PLL backbone and the surface in neutral aqueous media. While the molecular weights of PLL and PEG were fixed, that of dex and the grafting density of PEG or dex on the PLL were varied. The hydration capacity of the polymer chains was quantified through the combined experimental approach of optical waveguide lightmode spectroscopy (OWLS) and quartz crystal microbalance with dissipation monitoring (QCM-D) to yield a value for areal solvation (Ψ), i.e., mass of associated solvent molecules within the polymer chains per unit substrate area. For the two series of copolymers with comparable stretched chain lengths of hydrophilic polymers, namely, PLL(20)-g-PEG(5) and PLL(20)-g-dex(10), the Ψ values gradually increased as the initial grafting density on the PLL backbone increased or as g decreased. However, the rate of increase in Ψ was higher for PEG than dextran chains, which was attributed to higher stiffness of the dextran chains. More importantly, the number of water molecules per hydrophilic group was clearly higher for PEG chains. Given that the -CH2CH2O- units that make up the PEG chains form a cage-like structure with 2-3 water molecules, these "strongly bound" water molecules can account for the slightly more favorable behavior of PEG compared to dextran in both aqueous lubrication and antifouling behavior of the copolymers.

Comparative analysis of resting-state EEG-based multiscale entropy between schizophrenia and bipolar disorder.

BACKGROUND: Studies that use nonlinear methods to identify abnormal brain dynamics in patients with psychiatric disorders are limited. This study investigated brain dynamics based on EEG using multiscale entropy (MSE) analysis in patients with schizophrenia (SZ) and bipolar disorder (BD). METHODS: The eyes-closed resting-state EEG data were collected from 51 patients with SZ, 51 patients with BD, and 51 healthy controls (HCs). Patients with BD were further categorized into type I (n = 23) and type II (n = 16), and then compared with patients with SZ. A sample entropy-based MSE was evaluated from the bilateral frontal, central, and parieto-occipital regions using 30-s artifact-free EEG data for each individual. Correlation analyses of MSE values and psychiatric symptoms were performed. RESULTS: For patients with SZ, higher MSE values were observed at higher-scale factors (i.e., 41-70) across all regions compared with both HCs and patients with BD. Furthermore, there were positive correlations between the MSE values in the left frontal and parieto-occipital regions and PANSS scores. For patients with BD, higher MSE values were observed at middle-scale factors (i.e., 13-40) in the bilateral frontal and central regions compared with HCs. Patients with BD type I exhibited higher MSE values at higher-scale factors across all regions compared with those with BD type II. In BD type I, positive correlations were found between MSE values in all left regions and YMRS scores. CONCLUSIONS: Patients with psychiatric disorders exhibited group-dependent MSE characteristics. These results suggest that MSE features may be useful biomarkers that reflect pathophysiological characteristics.

Pharmacokinetic interactions between fexuprazan, a potassium-competitive acid blocker, and nonsteroidal anti-inflammatory drugs in healthy males.

Fexuprazan, a novel potassium-competitive acid blocker, is expected to be used for the prevention of nonsteroidal anti-inflammatory drugs (NSAIDs) induced ulcer. This study aimed to evaluate pharmacokinetic (PK) interactions between fexuprazan and NSAIDs in healthy subjects. A randomized, open-label, multicenter, six-sequence, one-way crossover study was conducted in healthy male subjects. Subjects randomly received one of the study drugs (fexuprazan 40 mg BID, celecoxib 200 mg BID, naproxen 500 mg BID, or meloxicam 15 mg QD) for 5 or 7 days in the first period followed by the combination of fexuprazan and one of NSAIDs for the same days and the perpetrator additionally administered for 1-2 days in the second period. Serial blood samples for PK analysis were collected until 48- or 72-h post-dose at steady state. PK parameters including maximum plasma concentration at steady state (Cmax,ss) and area under plasma concentration-time curve over dosing interval at steady state (AUCτ,ss) were compared between monotherapy and combination therapy. The PKs of NSAIDs were not significantly altered by fexuprazan. For fexuprazan, differences in PK parameters (22% in Cmax, 19% in AUCτ,ss) were observed when co-administered with naproxen, but not clinically significant. The geometric mean ratio (90% confidence interval) of combination therapy to monotherapy for Cmax,ss and AUCτ,ss was 1.22 (1.02-1.46) and 1.19 (1.00-1.43), respectively. There were no significant changes in the systemic exposure of fexuprazan by celecoxib and meloxicam. Fexuprazan and NSAIDs did not show clinically meaningful PK interactions.

Lipid Variability Induces Endothelial Dysfunction by Increasing Inflammation and Oxidative Stress.

BACKGRUOUND: This study investigates the impact of fluctuating lipid levels on endothelial dysfunction. METHODS: Human aortic and umbilical vein endothelial cells were cultured under varying palmitic acid (PA) concentrations: 0, 50, and 100 µM, and in a variability group alternating between 0 and 100 µM PA every 8 hours for 48 hours. In the lipid variability group, cells were exposed to 100 µM PA during the final 8 hours before analysis. We assessed inflammation using real-time polymerase chain reaction, Western blot, and cytokine enzyme-linked immunosorbent assay (ELISA); reactive oxygen species (ROS) levels with dichlorofluorescin diacetate assay; mitochondrial function through oxygen consumption rates via XF24 flux analyzer; and endothelial cell functionality via wound healing and cell adhesion assays. Cell viability was evaluated using the MTT assay. RESULTS: Variable PA levels significantly upregulated inflammatory genes and adhesion molecules (Il6, Mcp1, Icam, Vcam, E-selectin, iNos) at both transcriptomic and protein levels in human endothelial cells. Oscillating lipid levels reduced basal respiration, adenosine triphosphate synthesis, and maximal respiration, indicating mitochondrial dysfunction. This lipid variability also elevated ROS levels, contributing to a chronic inflammatory state. Functionally, these changes impaired cell migration and increased monocyte adhesion, and induced endothelial apoptosis, evidenced by reduced cell viability, increased BAX, and decreased BCL2 expression. CONCLUSION: Lipid variability induce endothelial dysfunction by elevating inflammation and oxidative stress, providing mechanistic insights into how lipid variability increases cardiovascular risk.

Evaporation-Driven Energy Generation Using an Electrospun Polyacrylonitrile Nanofiber Mat with Different Support Substrates.

Water evaporation-driven energy harvesting is an emerging mechanism for contributing to green energy production with low cost. Herein, we developed polyacrylonitrile (PAN) nanofiber-based evaporation-driven electricity generators (PEEGs) to confirm the feasibility of utilizing electrospun PAN nanofiber mats in an evaporation-driven energy harvesting system. However, PAN nanofiber mats require a support substrate to enhance its durability and stability when it is applied to an evaporation-driven energy generator, which could have additional effects on generation performance. Accordingly, various support substrates, including fiberglass, copper, stainless mesh, and fabric screen, were applied to PEEGs and examined to understand their potential impacts on electrical generation outputs. As a result, the PAN nanofiber mats were successfully converted to a hydrophilic material for an evaporation-driven generator by dip-coating them in nanocarbon black (NCB) solution. Furthermore, specific electrokinetic performance trends were investigated and the peak electricity outputs of Voc were recorded to be 150.8, 6.5, 2.4, and 215.9 mV, and Isc outputs were recorded to be 143.8, 60.5, 103.8, and 121.4 µA, from PEEGs with fiberglass, copper, stainless mesh, and fabric screen substrates, respectively. Therefore, the implications of this study would provide further perspectives on the developing evaporation-induced electricity devices based on nanofiber materials.

Real-world efficacy and safety of capecitabine with oxaliplatin in patients with advanced adenocarcinoma of the ampulla of Vater.

BACKGROUND: Adenocarcinoma of the ampulla of Vater (AoV) is one of the rare periampullary cancers, and due to its anatomical location, it is categorized into various histologic subtypes. Its rarity and diversity pose challenges in treatment decision-making for patients with advanced AoV carcinoma. This study investigated the efficacy and safety of the combined regimen of capecitabine and oxaliplatin (CAPOX) in a real-world clinical setting. METHODS: This investigation encompassed patients with advanced AoV carcinoma who underwent CAPOX treatment. Histologic phenotypes were identified through a combination of histopathological analysis and protein expression markers, including MUC1, CDX2, CK20, and MUC2. The correlation between histopathological determinants and survival outcomes was explored, in addition to an evaluation of the safety profile of CAPOX therapy. RESULTS: From January 2010 to June 2023, 42 patients received CAPOX. Of these, 14 patients (33.3%) had not received any prior palliative chemotherapy, while 28 patients (66.7%) had undergone one prior line of chemotherapy. At a median follow up of 9.0 months, the median progression-free survival (PFS) was 4.38 months (95% CI, 2.78-5.69) and the median overall survival (OS) was 9.57 months (95% CI 7.56-11.6). The objective response and disease control rates were 38.1% and 61.9%, respectively. Patients who received CAPOX as a second-line treatment had poorer PFS (HR = 2.62; 95% CI, 1.49-4.90, p = 0.003) and OS (HR = 2.82, 95% CI, 1.47-5.38, p = 0.001) compared to those who received CAPOX as a first-line chemotherapy. There were no statistically significant differences in PFS (p = 0.185) and OS (p = 0.097) between groups based on histologic subtypes. Neutropenia (14.3%) emerged as the predominant grade 3-4 toxicity. Notably, treatment cessation occurred in select instances owing to grade 3 fatigue (9.5%) and peripheral neuropathy (9.5%). CONCLUSIONS: This study confirmed the therapeutic efficacy and safety of CAPOX in a real-world setting, consistent with prior phase II trial results. While CAPOX proved feasible for advanced AoV carcinoma regardless of histologic subtype, its reduced effectiveness in second-line settings necessitates further research to determine its optimal palliative use.

Clinical Implication of Maumgyeol Basic Biotypes-Electroencephalography- and Photoplethysmogram-Based Bwave State Inventory.

OBJECTIVE: The development of individual subtypes based on biomarkers offers a cost-effective and timely avenue to comprehending individual differences pertaining to mental health, independent from individuals' subjective insights. Incorporating 2-channel electroencephalography (EEG) and photoplethysmogram (PPG), we sought to establish a subtype classification system with clinical relevance. METHODS: One hundred healthy participants and 99 patients with psychiatric disorders were recruited. Classification thresholds were determined using the EEG and PPG data from 2,278 individuals without mental disorders, serving to classify subtypes in our sample of 199 participants. Multivariate analysis of variance was applied to examine psychological distinctions among these subtypes. K-means clustering was employed to verify the classification system. RESULTS: The distribution of subtypes differed between healthy participants and those with psychiatric disorders. Cognitive abilities were contingent upon brain subtypes, while mind subtypes exhibited significant differences in symptom severity, overall health, and cognitive stress. K-means clustering revealed that the results of our theory-based classification and data-driven classification are comparable. The synergistic assessment of both brain and mind subtypes was also explored. CONCLUSION: Our subtype classification system offers a concise means to access individuals' mental health. The utilization of EEG and PPG signals for subtype classification offers potential for the future of digital mental healthcare.

Efficacy and Safety of Metformin and Atorvastatin Combination Therapy vs. Monotherapy with Either Drug in Type 2 Diabetes Mellitus and Dyslipidemia Patients (ATOMIC): Double-Blinded Randomized Controlled Trial.

Background: It is well known that a large number of patients with diabetes also have dyslipidemia, which significantly increases the risk of cardiovascular disease (CVD). This study aimed to evaluate the efficacy and safety of combination drugs consisting of metformin and atorvastatin, widely used as therapeutic agents for diabetes and dyslipidemia. Methods: This randomized, double-blind, placebo-controlled, parallel-group and phase III multicenter study included adults with glycosylated hemoglobin (HbA1c) levels >7.0% and <10.0%, low-density lipoprotein cholesterol (LDL-C) >100 and <250 mg/dL. One hundred eighty-five eligible subjects were randomized to the combination group (metformin+atorvastatin), metformin group (metformin+atorvastatin placebo), and atorvastatin group (atorvastatin+metformin placebo). The primary efficacy endpoints were the percent changes in HbA1c and LDL-C levels from baseline at the end of the treatment. Results: After 16 weeks of treatment compared to baseline, HbA1c showed a significant difference of 0.94% compared to the atorvastatin group in the combination group (0.35% vs. -0.58%, respectively; P<0.0001), whereas the proportion of patients with increased HbA1c was also 62% and 15%, respectively, showing a significant difference (P<0.001). The combination group also showed a significant decrease in LDL-C levels compared to the metformin group (-55.20% vs. -7.69%, P<0.001) without previously unknown adverse drug events. Conclusion: The addition of atorvastatin to metformin improved HbA1c and LDL-C levels to a significant extent compared to metformin or atorvastatin alone in diabetes and dyslipidemia patients. This study also suggested metformin's preventive effect on the glucose-elevating potential of atorvastatin in patients with type 2 diabetes mellitus and dyslipidemia, insufficiently controlled with exercise and diet. Metformin and atorvastatin combination might be an effective treatment in reducing the CVD risk in patients with both diabetes and dyslipidemia because of its lowering effect on LDL-C and glucose.

Pioglitazone-induced alterations of purine metabolism in healthy male subjects.

Pioglitazone is class of thiazolidinediones that activates peroxisome proliferator-activated receptors (PPARs) in adipocytes to improve glucose metabolism and insulin sensitivity and has been used as a treatment for type 2 diabetes. However, the underlying mechanisms of associated pioglitazone-induced effects remain unclear. Our study aimed to investigate endogenous metabolite alterations associated with pioglitazone administration in healthy male subjects using an untargeted metabolomics approach. All subjects received 30 mg of pioglitazone once daily in the assigned sequence and period. Urine samples were collected before pioglitazone administration and for 24 h after 7 days of administration. A total of 1465 compounds were detected and filtered using a coefficient of variance below 30% and 108 metabolites were significantly altered upon pioglitazone administration via multivariate statistical analysis. Fourteen significant metabolites were identified using authentic standards and public libraries. Additionally, pathway analysis revealed that metabolites from purine and beta-alanine metabolisms were significantly altered after pioglitazone administration. Further analysis of quantification of metabolites from purine metabolism, revealed that the xanthine/hypoxanthine and uric acid/xanthine ratios were significantly decreased at post-dose. Pioglitazone-dependent endogenous metabolites and metabolic ratio indicated the potential effect of pioglitazone on the activation of PPAR and fatty acid synthesis. Additional studies involving patients are required to validate these findings.

Clinical and Genetic Characteristics Associated With Survival Outcome in Late-Onset Huntington's Disease in South Korea.

BACKGROUND AND PURPOSE: The onset of Huntington's disease (HD) usually occurs before the age of 50 years, and the median survival time from onset is 15 years. We investigated survival in patients with late-onset HD (LoHD) (age at onset ≥60 years) and the associations of the number of mutant CAG repeats and age at onset (AAO) with survival in patients with HD. METHODS: Patients with genetically confirmed HD at six referral centers in South Korea between 2000 and 2020 were analyzed retrospectively. Baseline demographic, clinical, and genetic characteristics and the survival status as at December 2020 were collected. RESULTS: Eighty-seven patients were included, comprising 26 with LoHD (AAO=68.77±5.91 years, mean±standard deviation; 40.54±1.53 mutant CAG repeats) and 61 with common-onset HD (CoHD) (AAO=44.12±8.61 years, 44.72±4.27 mutant CAG repeats). The ages at death were 77.78±7.46 and 53.72±10.86 years in patients with LoHD and CoHD, respectively (p<0.001). The estimated survival time was 15.21±2.49 years for all HD patients, and 10.74±1.95 and 16.15±2.82 years in patients with LoHD and CoHD, respectively. More mutant CAG repeats and higher AAO were associated with shorter survival (hazard ratio [HR]=1.05, 95% confidence interval [CI]=1.01-1.09, p=0.019; and HR=1.17, 95% CI=1.03-1.31, p=0.013; respectively) for all HD patients. The LoHD group showed no significant factors associated with survival after disease onset, whereas the number of mutant CAG repeats had a significant effect (HR=1.12, 95% CI=1.01-1.23, p=0.034) in the CoHD group. CONCLUSIONS: Survival after disease onset was shorter in patients with LoHD than in those with CoHD. More mutant CAG repeats and higher AAO were associated with shorter survival in patients with HD.

Targeting CD73 with flavonoids inhibits cancer stem cells and increases lymphocyte infiltration in a triple-negative breast cancer mouse model.

Introduction: Chemotherapy remains the mainstay treatment for triple-negative breast cancer (TNBC) due to the lack of specific targets. Given a modest response of immune checkpoint inhibitors in TNBC patients, improving immunotherapy is an urgent and crucial task in this field. CD73 has emerged as a novel immunotherapeutic target, given its elevated expression on tumor, stromal, and specific immune cells, and its established role in inhibiting anti-cancer immunity. CD73-generated adenosine suppresses immunity by attenuating tumor-infiltrating T- and NK-cell activation, while amplifying regulatory T cell activation. Chemotherapy often leads to increased CD73 expression and activity, further suppressing anti-tumor immunity. While debulking the tumor mass, chemotherapy also enriches heterogenous cancer stem cells (CSC), potentially leading to tumor relapse. Therefore, drugs targeting both CD73, and CSCs hold promise for enhancing chemotherapy efficacy, overcoming treatment resistance, and improving clinical outcomes. However, safe and effective inhibitors of CD73 have not been developed as of now. Methods: We used in silico docking to screen compounds that may be repurposed for inhibiting CD73. The efficacy of these compounds was investigated through flow cytometry, RT-qPCR, CD73 activity, cell viability, tumorsphere formation, and other in vitro functional assays. For assessment of clinical translatability, TNBC patient-derived xenograft organotypic cultures were utilized. We also employed the ovalbumin-expressing AT3 TNBC mouse model to evaluate tumor-specific lymphocyte responses. Results: We identified quercetin and luteolin, currently used as over-the-counter supplements, to have high in silico complementarity with CD73. When quercetin and luteolin were combined with the chemotherapeutic paclitaxel in a triple-drug regimen, we found an effective downregulation in paclitaxel-enhanced CD73 and CSC-promoting pathways YAP and Wnt. We found that CD73 expression was required for the maintenance of CD44highCD24low CSCs, and co-targeting CD73, YAP, and Wnt effectively suppressed the growth of human TNBC cell lines and patient-derived xenograft organotypic cultures. Furthermore, triple-drug combination inhibited paclitaxel-enriched CSCs and simultaneously improved lymphocyte infiltration in syngeneic TNBC mouse tumors. Discussion: Conclusively, our findings elucidate the significance of CSCs in impairing anti-tumor immunity. The high efficacy of our triple-drug regimen in clinically relevant platforms not only underscores the importance for further mechanistic investigations but also paves the way for potential development of new, safe, and cost-effective therapeutic strategies for TNBC.

Structural and tribological studies on the interaction of porcine gastric mucin with non- and cationic-modified ß-lactoglobulins.

ß-lactoglobulin (BLG) is the major whey protein with negative charges at neutral pH in aqueous media. Thus, the interaction with mucins, the major polyanionic component of mucus, is very weak due to the electrostatic repulsion between them. The present study postulates that cationization of BLG molecules may reverse the interaction characteristics between BLG and mucin from repulsive to associative. To this end, cationic-modified BLGs were prepared by grafting positively charged ethylenediamine (EDA) moieties into the negatively charged carboxyl groups on the aspartic and glutamic acid residues and compared with non-modified BLG upon mixing with porcine gastric mucin (PGM). To characterize the structural and conformational features of PGM, non/cationized BLGs, and their mixtures, various spectroscopic approaches, including zeta potential, dynamic light scattering (DLS), and circular dichroism (CD) spectroscopy were employed. Importantly, we have taken surface adsorption with optical waveguide lightmode spectroscopy (OWLS), and tribological properties with pin-on-disk tribometry at the sliding interface as the key approaches to determine the interaction nature between them as mixing PGM with polycations can lead to synergistic lubrication at the nonpolar substrate in neutral aqueous media as a result of an electrostatic association. All the spectroscopic studies and a substantial improvement in lubricity collectively supported a tenacious and associative interaction between PGM and cationized BLGs, but not between PGM and non-modified BLG. This study demonstrates a unique and successful approach to intensify the interaction between BLG and mucins, which is meaningful for a broad range of disciplines, including food science, macromolecular interactions, and biolubrication etc.