Approximately one-third of people with diabetes have diabetic kidney disease (DKD), and DKD remains a leading cause of kidney failure in the United States. DKD is characterized by injury to the glomerular filtration barrier and renal tubulointerstitium, arising from complex and convergent metabolic, hemodynamic, and inflammatory insults. Tyrosine phosphorylation, a reversible post-translational modification, is a central regulatory mechanism in these pathways and is dynamically controlled by protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs). Aberrant tyrosine phosphorylation is a hallmark of DKD and implicates PTPs as key modulators of podocyte, endothelial, tubular, and immune cell dysfunction. Over 100 genes code for PTPs in humans, with 38 ‘classical’ PTPs that specifically target tyrosine motifs, including receptor-type and non-receptor-type subtypes. Several classical PTPs are implicated in DKD pathogenesis, while others are increasingly associated with signaling cascades that drive renal metabolism, inflammation, fibrosis, and cell death. There is growing interest in PTPs as their contributions to the disease emerge, though the role(s) of some remain uncharacterized in DKD. Moreover, recent breakthroughs in pharmaceuticals targeting PTPs for metabolic diseases and cancer have ignited interest in repurposing or tailoring PTP‑directed therapeutics for DKD. This review summarizes current knowledge and recent advances of classical PTPs in DKD, including their roles in insulin signaling, cellular architecture, and glomerular and tubular cell injury. It further discusses existing DKD treatments and highlights emerging strategies to pharmacologically modulate PTP activity, such as active‑site inhibitors, allosteric agents, and antibody‑targeting approaches. By integrating mechanistic insights with drug discovery efforts, this review outlines how targeting specific PTPs could complement current standard‑of‑care therapies and open new avenues for precision treatment of DKD.
Personalized neoantigen cancer vaccine is a promising strategy for precision immunotherapy by targeting patient-specific and mutation-derived tumor antigens. Early clinical studies have demonstrated the feasibility, safety, and immunogenicity of these vaccines across multiple solid tumors, with encouraging outcomes particularly when combined with immune checkpoint blockade. However, broader clinical translation remains limited by sequential bottlenecks across the vaccine development pipeline, including false-positive neoantigen selection, imperfect modeling of antigen processing and HLA presentation, limited prediction of T-cell receptor recognition, and challenges in formulation, delivery, and manufacturing. Artificial intelligence and advanced computational workflows are increasingly integrated into this pipeline to improve candidate prioritization and support more reproducible decision-making. In this review, we summarize clinical progress and key translational barriers in personalized neoantigen vaccination, and discuss how AI-enabled approaches may contribute across four major stages: multi-omics integration for neoantigen discovery, processing-aware HLA presentation prediction, structure-aware and TCR-informed immunogenicity modeling, and data-driven formulation optimization, particularly for lipid nanoparticle-based delivery systems. These approaches are able to help narrow biological and chemical search spaces, improve prioritization, and provide mechanistic insights into antigen presentation and immune recognition rather than replacing experimental validation. This articlefurther addresses future implementation challenges, including dataset diversity, model interpretability, prospective benchmarking, manufacturing traceability, and evolving regulatory frameworks for individualized mRNA cancer immunotherapies. Integrating computational innovation with rigorous immunological validation, scalable manufacturing, and regulatory oversight will be essential for advancing personalized neoantigen vaccines toward broader clinical implementation.
Cardiac fibrosis is a pathological remodeling process that contributes to the development and progression of heart failure. Although glucose-dependent insulinotropic polypeptide (GIP) may exert antifibrotic effects, how GIP receptor activation regulates cardiac fibrogenesis and modulates cardiac function in heart failure remains unclear. This study investigated whether GIP receptor agonism suppresses cardiac fibroblast activity and improves heart failure and explored the underlying mechanisms by using cellular and animal models. Human cardiac fibroblasts were treated with [D-Ala2]GIP (DA-GIP; 10, 100, or 300 nM) for 24 h or left untreated (control). Fibroblast migration, collagen production, and intracellular signaling were examined using wound healing, immunoblotting, enzyme-linked immunosorbent, and fluorometric assays. Cardiac structure, function, and fibrosis were assessed through echocardiography and Masson’s trichrome staining in rats with isoproterenol-induced heart failure with and without DA-GIP (24 nM/kg, twice daily for 2 weeks) administration. Compared with control cells, DA-GIP (300 nM)-treated cardiac fibroblasts exhibited a significantly lower migratory activity and reduced expression levels of pro-collagen IA1, pro-collagen III, and transforming growth factor-β1 proteins. Additionally, DA-GIP increased nitric oxide (NO) production and promoted endothelial NO synthase (eNOS) and protein kinase B (Akt) activation in cardiac fibroblasts. Notably, Akt inhibition blocked DA-GIP-induced eNOS activation, and treatment with Nω-nitro-L-arginine methyl ester (a NO synthase inhibitor, 100 μM) attenuated the antifibrotic effect of DA-GIP. In heart failure rats, DA-GIP reduced myocardial fibrosis, chamber dilatation, and systolic dysfunction. DA-GIP suppresses cardiac fibroblast activity through Akt-dependent eNOS activation and subsequent NO production, thereby improving cardiac remodeling and function in experimental heart failure.
Abstract Background Lupus nephritis (LN) is a severe manifestation of systemic lupus erythematosus (SLE) associated with significant morbidity. Although reduced production of reactive oxygen species (ROS) by neutrophils correlates with severe SLE, the specific mechanisms linking ROS deficiency to heightened renal inflammation remain unknown. We aimed to elucidate the role of NOX2-derived ROS in LN pathogenesis and identify potential therapeutic targets. Methods We conducted an in vivo study using a pristane-induced lupus model in Ncf1 −/− (NOX2-deficient) mice and wild-type controls. We assessed LN severity and characterized renal immune infiltration using flow cytometry and single-cell RNA sequencing (scRNA-seq). We performed transcriptomic analysis to evaluate the function of NOX2-deficient neutrophils. Finally, we tested the therapeutic efficacy of an IL-1 receptor antagonist in ameliorating disease severity in the NOX2-deficient mice. Results NOX2 deficiency exacerbated LN severity compared to wild-type controls, demonstrated by increased serum anti-dsDNA antibody titers and worsened LN scores. Through scRNA-seq, we identified a distinct, activated neutrophil subset in Ncf1 −/− mice featuring a robust interferon signature and high Nlrp3 expression. Transcriptomic analysis confirmed the upregulation of core NLRP3 pathway within these cells. Crucially, treating NOX2-deficient mice with an IL-1 receptor antagonist reduced LN activity scores. Conclusion Our results showed that NOX2 deficiency was associated with the expansion of a highly inflammatory renal neutrophil subset that may contribute to aggravated renal inflammation. These findings suggest that NOX2 functions as a negative regulator of the NLRP3 inflammasome and IL-1β blockade represents a promising precision therapeutic strategy for patients with LN who exhibit impaired ROS production.
Abstract Background SARS-CoV-2 remains a global health threat because ongoing viral evolution and immune evasion reduce the effectiveness of existing therapies. SNS812 is an inhaled small interfering RNA targeting a highly conserved region of the viral RNA-dependent RNA polymerase gene, representing a strategy that may enable broader antiviral activity against emerging variants. Methods In this phase 2, double-blind, randomised, placebo-controlled trial, adults with mild-to-moderate COVID-19 within 3 days of symptom onset were randomly assigned (1:1:1) to receive placebo or inhaled SNS812 (100 mg or 200 mg) once daily for 7 days (ClinicalTrials.gov identifier: NCT05941793). Safety was the primary endpoint. Secondary endpoints included the time to sustained alleviation (TTSA) and resolution (TTSR) for prespecified composite target symptoms and individual symptoms. Virological outcomes were exploratory. Results A total of 135 participants were enrolled, with more than 90% infected with immune-evasive SARS-CoV-2 variants. No treatment-related adverse events or serious adverse events were reported. In exploratory analyses, the 200 mg SNS812 group showed a shorter median time to SARS-CoV-2 antigen negativity (2.9 vs 3.6 days; p = 0.007) and a faster viral load reduction rate (− 0.755 vs − 0.652; p = 0.040), demonstrating dose-dependent virological effects. In the modified intention-to-treat population, exploratory symptom analyses showed shorter TTSA and TTSR for prespecified target symptoms with SNS812 200 mg compared with placebo (median TTSR 6.1 vs 8.1 days; adjusted hazard ratio 2.07, 95% CI 1.30–3.29; median TTSA 3.6 vs 6.5 days; adjusted hazard ratio 1.81, 95% CI 1.14–2.88). Conclusion Inhaled SNS812 was safe and well tolerated and showed dose-dependent antiviral activity, with exploratory signals of symptomatic improvement in adults with mild-to-moderate COVID-19 infected with immune-evasive variants. These findings support further evaluation in larger, adequately powered trials, including older and higher-risk populations.
Abstract Background Pathogenic sequence alterations in the SLC26A4 gene, which encodes the solute carrier SLC26A4/pendrin, lead to Pendred syndrome and non-syndromic autosomal recessive deafness type B4 (DFNB4), two of the most common forms of hearing loss worldwide. Many pathogenic SLC26A4 protein variants exhibit reduced cellular levels due to ubiquitin-proteasome system (UPS)-mediated degradation, and UPS inhibition rescues their plasma membrane expression and ion transport function. However, the underlying molecular mechanisms remain unclear and may involve interactions with novel molecular partners. Methods A candidate SLC26A4 protein partner was found by a yeast two-hybrid screening. The biological significance of this interaction has been studied by immunohistochemistry and co-localization in the mouse inner ear and kidney, co-immunoprecipitation of endogenous and recombinant proteins, Liquid Chromatography-Tandem Mass Spectrometry, and Fluorescence Resonance Energy Transfer. Results We identified the zinc finger and BTB domain-containing protein ZBTB16 as a novel SLC26A4-interacting partner. ZBTB16 co-localized with SLC26A4 in the outer sulcus and spiral prominence epithelial cells of the mouse cochlea and in the apical membrane of non-alpha non-beta intercalated cells of the distal nephron. ZBTB16 was found to be part of a ubiquitin-ligase complex comprising the scaffold protein Cullin 3 and the ubiquitin ligase RocI, and to bind with its C-terminal zinc finger region a unique amino acid sequence within the C-terminal Sulfate Transporter and Anti-Sigma factor Antagonist (STAS) domain of SLC26A4. This direct molecular interaction leads to increased site- and variant-specific ubiquitination and accelerated degradation of SLC26A4. Finally, using AI-based structure prediction, we provide an atomistic model of the complete SLC26A4/ZBTB16/Cullin 3/RocI complex in agreement with our experimental results. Conclusion These findings describe a primary mechanism of SLC26A4 regulation in the inner ear and kidney and of SLC26A4 loss of function in Pendred syndrome and deafness DFNB4, and identify potential novel molecular targets for therapeutic intervention.
Abstract Background To enhance the efficiency of identifying rare variants within the Taiwanese population and to support genome-wide association studies (GWAS) and imputation studies for genetic risk prediction in the Han population, we have developed the National Health Research Institutes (NHRI) reference panel (NHRI-RP-1). Methods NHRI-RP-1 is based on 2,561 whole genome sequences taken from the in-house NHRI datasets. Our objective was to optimize conditions of sample sizes (0.5K, 1K, 1.5K, 2K, 2.5K), minor allele frequency (MAF) thresholds (MAF ≥ 0.05, 0.01, 0.001, 2 × 10 –4 ), and imputation quality (r 2 ≥ 0, 0.3, 0.5) to build an aggregated genome reference panel for genetic medicine by comparing with worldwide references. Clinical applications and GWAS were then evaluated to demonstrate the capability of the reference panel. Results Among different combinations of relevant parameters, the NHRI-RP-1 (with a 2,500-sample size, MAF ≥ 2 × 10 –4 , r 2 ≥ 0) demonstrated a superior F1 score on local match, genotype concordance and r-squared, as compared to those using worldwide reference genomes, particularly for rare MAFs. Furthermore, NHRI-RP-1 achieved over 95% accuracy for nine pathogenic variants present in the Taiwan Biobank and 93.49% and 92.9% accuracy for imputing two DRD1 variants. Conclusions The use of different reference panels can influence the outcomes of GWAS. Our case studies demonstrate the utility of the NHRI-RP-1 for genetic medicine. Incorporating a population-specific panel such as NHRI-RP-1 can facilitate the development of prediction models using polygenic risk scores for diseases that are common in the Han population.
Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer with limited treatment options. Although PARP inhibitor (PARPi) offers great promise in treating TNBC with deficiency in homologous recombination (HR), most TNBC patients are HR-proficient. Furthermore, acquired resistance to PARPi remains as a challenge. Thus, there is an unmet need to identify new therapeutic target for developing advanced TNBC treatment strategy. I-SceI reporter assay and alkaline comet assay were used to analyze the role of Smyca in HR repair. Ingenuity pathway analysis was used to identify upstream regulators of Smyca-regulated transcriptome. RNA immunoprecipitation and RNA pull down were used to examine Smyca-FOXM1 interaction. Chromatin immunoprecipitation followed by sequencing was performed to identify FOXM1 target genes that are regulated by Smyca. Chromatin isolation by RNA purification was used to determine Smyca loading onto the promoters of FOXM1 target genes. Patient-derived organoid and xenograft mouse models were performed to evaluate the effect of Smyca on chemoresistance. Nanoparticle-assisted gapmer antisense oligonucleotides delivery was used to target Smyca in vivo. Co-culture of CD3 + T cells with TNBC cells and syngeneic mouse model were used to examine the effect of Smyca-FOXM1 targeting on anti-tumor immunity. The long non-coding RNA Smyca is highly expressed in TNBC. We show that Smyca is induced by genotoxic agents to enhance HR repair. Mechanistically, Smyca binds FOXM1 and promotes the recruitment of FOXM1 to the promoters of a set of HR and nucleotide metabolism genes, thereby promoting their expression. Smyca ablation induces BRCAness in HR-proficient TNBC, thereby sensitizing these tumors to platinum or PARPi. Furthermore, targeting Smyca-FOXM1 complex in combination with platinum or PARPi activates cGAS/STING pathway and tumor immunogenicity to enhance anti-tumor immune surveillance. Clinically, Smyca expression in breast cancer patients correlates positively with therapy resistance and negatively with HR deficiency, interferon signature, and infiltration of anti-tumor immune cells. Our study identifies an unprecedented role of Smyca in HR repair to promote TNBC survival and immune evasion in response to therapy and suggests Smyca as a potential target for sensitizing TNBC to chemotherapy, PARPi, or immunotherapy.
Individuals with asymptomatic SARS-CoV-2 infection can unknowingly transmit the virus, yet identifying such subclinical infections in post-vaccinated populations remains challenging. We conducted a longitudinal study of 129 infection-naïve vaccine recipients immunized with various combinations of SARS-CoV-2 spike (S) protein vaccine platforms. Sera were collected before the first dose (v1), at 2 weeks (v7) and 6 months (v8) after the third dose. Taiwan’s first major COVID-19 outbreak occurred between v7 and v8. We measured anti-nucleocapsid (anti-N) and anti-S IgG antibody titers by ELISA and assessed virus-neutralizing activity using live virus and pseudovirus assays. By developing an iterative serial screening method, we identified asymptomatic breakthrough (post-vaccination) infections among unconfirmed cases. Our v7-v8 paired cohort resolved into three distinct groups: confirmed cases (21
Chikungunya virus (CHIKV), a mosquito-borne alphavirus, causes frequent global epidemics of chikungunya fever, characterized by severe joint pain and debilitating arthritis. With no specific antiviral therapies available, these outbreaks pose a major public health challenge, particularly in tropical regions. There is thus urgent need to develop novel antivirals. Drug repurposing is an attractive strategy to identify potential antivirals targeting CHIKV replication. The Spectrum collection of approved drugs was screened using high-content cell imaging to identify potential CHIKV replication inhibitors. Efficacy of these CHIKV inhibitors was evaluated in a C57BL/6 mouse model of chikungunya disease, monitoring viremia and clinical symptoms like joint swelling. The CHIKV inhibitor's effect on virus uptake, replication, RNA synthesis, and protein production was studied in ERMS cells. In silico molecular docking was used to study the inhibitor’s binding to the CHIKV proteins. Binding was validated in vitro using microscale thermophoresis and isothermal titration calorimetry. CHIKV helicase, protease, and ATPase activities were measured in the presence of emetine dihydrochloride (ED) to determine its mechanism of action. Out of the four CHIKV inhibitors identified by high content screening, ED potently inhibited CHIKV replication in the mouse model, yielding significantly lower viremia levels. Notably, CHIKV-infected mice treated with ED showed no clinical symptoms of joint swelling. In ERMS cells, ED blocked CHIKV uptake and early replication by suppressing viral RNA synthesis, which in turn prevented viral protein production. Computational modelling predicted ED’s binding around the RNA-binding site of the CHIKV nsP2 helicase domain. In vitro validation confirmed dose-dependent ED binding with CHIKV nsP2. ED specifically inhibited helicase unwinding in a concentration-dependent manner without affecting ATPase or protease activity. This study demonstrates that ED inhibits CHIKV replication in cultured cells and a mouse model of infection through binding to CHIKV nsP2 and inhibition of its helicase activity. While this is a promising virus-targeted mechanism of ED’s antiviral action, additional host-directed mechanisms warrant further investigation.
Abstract Background Persistent injury and impaired regeneration of the alveolar epithelium are key contributors to the pathogenesis of pulmonary fibrosis. In idiopathic pulmonary fibrosis (IPF), type 2 alveolar epithelial (AT2) cells fail to fully differentiate into type 1 alveolar epithelial (AT1) cells, remaining instead in a transitional state. Histone deacetylase (HDAC) inhibitors are promising therapeutic agents for pulmonary fibrosis. Therefore, this study investigated whether MPT0E028, a pan-HDAC inhibitor, ameliorated bleomycin (BLM)-induced pulmonary fibrosis in a therapeutic model of mice by promoting AT2-to-AT1 cell differentiation. Methods The effects of MPT0E028 on pulmonary fibrosis were assessed by evaluating the expression of fibrogenic proteins and cell markers of AT1 (T1α and aquaporin 5 [AQP5]), AT2 (surfactant protein C [SPC]) and alveolar epithelial transitional cells (Keratin 8 [KRT8]) in a therapeutic model of BLM-induced pulmonary fibrosis in mice. The role of the ataxia-telangiectasia mutated (ATM)/AMP-activated protein kinase (AMPK)/forkhead box O1 (FoxO1) signaling pathway in MPT0E028-induced T1α expression was examined in murine AT2 cells (MLE-12 cells). Results Administration of MPT0E028 significantly reduced fibrosis scores; suppressed the expression of connective tissue growth factor, collagen I, fibronectin, and α-smooth muscle actin; and improved lung function in the therapeutic model of BLM-induced pulmonary fibrosis in mice. MPT0E028 enhanced the expression of T1α and AQP5 but reduced the expression of SPC and KRT8 in lung tissues from BLM-treated mice. In MLE-12 cells and primary human AT2 cells, MPT0E028 upregulated T1α and AQP5 expression in a time-dependent manner, with this accompanied by a decrease in SPC expression. AS1842856, an FoxO1 inhibitor, and FoxO1 siRNA transfection inhibited MPT0E028-stimulated T1α expression, whereas transfection with FoxO3 siRNA had no effect. FoxO1 siRNA transfection also inhibited MPT0E028-stimulated T1α-luciferase activity. MPT0E028 induced FoxO1 serine phosphorylation, increased FoxO1 recruitment to the T1α promoter, and enhanced FoxO1-luciferase activity. Compound C, an AMPK inhibitor, and AMPK siRNA transfection suppressed MPT0E028-stimulated T1α expression, and compound C also inhibited MPT0E028-promoted FoxO1 recruitment to the T1α promoter. MPT0E028 induced AMPK phosphorylation in a time-dependent manner and increased ATM acetylation and phosphorylation in MLE-12 cells. ATM siRNA transfection suppressed MPT0E028-induced T1α expression, AMPK and FoxO1 serine phosphorylation, FoxO1 recruitment to the T1α promoter, and FoxO1-luciferase activity. MPT0E028 induced FoxO1 phosphorylation in AT2 cells in the therapeutic model of BLM-induced pulmonary fibrosis in mice. Conclusions MPT0E028 is the first pan-HDAC inhibitor shown to activate ATM acetylation-mediated AMPK/FoxO1 signaling to induce AT2-to-AT1 differentiation in a therapeutic model of BLM-induced pulmonary fibrosis in mice. Administration of MPT0E028 after BLM challenge effectively ameliorated pulmonary fibrosis by suppressing fibrogenic protein expression and promoting AT2-to-AT1 cell differentiation. These results suggest that MPT0E028 holds potential as a therapeutic agent for IPF treatment.
Photodynamic therapy (PDT) demonstrates remarkable versatility by activating diverse non-apoptotic cell death pathways, effectively circumventing apoptosis resistance and enhancing tumor eradication. Key mechanisms include autophagic cell death, regulated necrosis-driven immune activation, ferroptosis-mediated oxygen replenishment, pyroptosis-induced immunogenic cell death (ICD), and paraptosis. These pathways collectively highlight PDT’s capacity to disrupt tumor survival mechanisms and stimulate systemic anti-tumor immunity. However, several challenges remain, including precise spatiotemporal control of ROS, hypoxia mitigation, and selective photosensitizer delivery. Advances in nanotechnology, hypoxia-responsive agents, and combination therapies (e.g., immune checkpoint inhibitors or chemotherapy) hold promise for overcoming these limitations. In this review, we discuss multiple types of non-apoptotic cell death pathways activated by PDT and the underlying mechanisms of each distinct cell death process. We also review the clinical applications and current challenges of leveraging these pathways to enhance tumor treatment in PDT. Future research should prioritize the development of subcellular-targeted photosensitizers, deeper light-penetration technologies, and biomarker-guided personalized regimens for more precise and effective PDT.
Abstract Background Circulating monocyte trans-endothelial migration is critical for vascular diseases. The monocyte surface proteins, i.e., integrins and membrane receptors are central in this process. Previous studies demonstrate that ablation of the mitochondrial arginase-2 ( ARG2 ) reduces monocyte/macrophage infiltration in cardiovascular disease. We further investigate whether ARG2 regulates integrin and surface receptor levels in monocytes and facilitates trans-endothelial migration, contributing to atherogenesis. Methods For this purpose, human THP1 cells deficient in ARG2 gene (THP1 ARG2–/– ) were generated by CRISPR-U ™ -mediated genome engineering. Atherosclerotic Apoe –/– Arg2 + / + mice and Apoe –/– Arg2 –/– mouse models are used. Proteomic profiling, cellular/molecular biology methods, and confocal microscopy were utilized for gene or protein expression analysis. Results As compared to the control THP1 WT cells, the THP1 ARG2–/– cells reveal decreased adhesion and trans-endothelial migratory activities towards chemoattractants in a transwell co-culture system, accompanied by lower levels of αL and α4 integrins, CD99 and PECAM1 (CD31) (surface molecules for trans-endothelial activity) and CCR2 (the chemoattractant receptor). The monocyte adhesion to endothelial cells is reduced by blocking LFA-1 (αLβ2) or VLA-4 (α4β1) integrin. Pro-inflammatory polarization of the THP1 cells with LPS does not affect the integrin and surface receptor levels, however, enhances release of several pro-inflammatory cytokines, which is reduced in THP1 ARG2–/– cells due to reduced TLR4–ERK–NF-κB signaling. Conditioned medium from the LPS-primed THP1 WT reveals higher capacity to enhance endothelial VCAM-1 and ICAM-1 levels than the THP1 ARG2–/– cells, that is partly mediated by IL-1β. Moreover, ARG2-dependent TGF-β signaling was found to selectively regulate αL expression in monocytes. Finally, as compared to Apoe –/– Arg2 + / + mice, the ApoE –/– Arg2 –/– mice show significantly decreased macrophage integrin, chemoattractant receptor levels, and atherosclerosis. Conclusions Our study identifies ARG2 as a key regulator of monocyte-mediated vascular inflammation and atherogenesis by controlling integrin, chemoattractant receptor expression, and pro-inflammatory cytokine release through TLR4-ERK-NFκB signaling, highlighting ARG2 as a potential therapeutic target for vascular and chronic inflammatory diseases.
Antifungal resistance represents a growing global health challenge, driven by limited therapeutic options, inadequate use of both clinical and environmental drugs, and the remarkable adaptive capacity of fungal pathogens. While resistance has classically been attributed to genetic alterations affecting drug targets and efflux systems, accumulating evidence indicates that regulatory mechanisms play a critical role in shaping antifungal susceptibility. This review highlights that acetyltransferases constitute important and active components of antifungal resistance, operating through multiple integrated mechanisms. Notably, recent studies have extended this framework by identifying the direct enzymatic inactivation of antifungal agents by acetyltransferases, which emerges as an additional resistance mechanism and establishes a functional parallel with antibiotic acetylation in bacteria. These findings highlight acetyltransferases as clinically relevant drivers of antifungal resistance and suggest that targeting these enzymes may enhance the efficacy of existing antifungal therapies and reduce treatment failure.
Background Calcium (Ca2+) signalling and phosphorylation are independent mechanisms that regulate diverse biological processes. It is, however, not appreciated that a normal function of phospho-mimic amino acids (aspartate/glutamate) is to interact with Ca2+ at the atomic level. Methods In this work, we have utilised a combination of molecular virology, phosphorylation proteomics mass spectrometry, surface plasmon resonance, nuclear magnetic resonance, AlphaFold structural prediction, and electron microscopy imaging to systematically evaluated the biology of calcium - phosphorylation interplay. Results Here, we leveraged HIV-Ca2+ biology to enable polarised targeting in primary cells to describe an unknown layer of regulatory processes via Ca2+-phosphate (PO43-) bridge to support protein complex formation. We identified novel HIV phosphorylation sites proximal to Endosomal Sorting Complex Required for Transport (ESCRT) binding motifs and potential Ca2+ binding sites through phospho-proteomics. Integrating primary cells, molecular virology, structural biology, biophysical and ultrastructural analyses, we presented multiple examples of coordination between calcium and phosphorylated amino acids to support HIV assembly and function. Our data suggest Ca2+-PO43- could form a bridge between intrinsically disordered protein regions of HIV to: (i) stabilise Pr55(Gag)-Pr160(GagPol) complex for virus function; (ii) mediate p6(Pol) dimerization to support virion maturation; and (iii) modulate viral complex formation to package both viral enzymatic- and cellular- proteins. Conclusions Our work has identified the coordination between calcium and phosphorylated amino acids (presumably via the formation of a salt-bridge) is an unknown layer of regulation that support protein complex formation. In the context of HIV, these Ca2+-PO43- cooperation via the intrinsically disordered protein regions regulates the assembly of HIV complex and modulates its interactions with ESCRT cellular proteins. Importantly, the convergent enrichment of calcium-phosphorylation domains across a wide range of viral and cellular intrinsically disordered protein regions implies Ca2+-PO43- cooperation to be a general regulatory principle in biology.
Abstract Background Dengue virus (DENV) infection drives pathological inflammation through coordinated activation of pattern recognition receptors (PRRs) in myeloid cells, yet no approved immunomodulatory therapy exists to interrupt this process. CLEC5A and Toll-like receptor 2 (TLR2) have each been implicated in DENV-induced cytokine production, but whether their co-engagement represents a convergent and therapeutically targetable axis remains unclear. Methods We define the cytokine landscape of human macrophages infected with all four DENV serotypes and reverse genetics DENV2 (rgDV2) strains carrying NS1 mutations derived from the severe 2015 Taiwan outbreak. We further demonstrate the potential therapeutic effect of bispecific antibodies simultaneously targeting CLEC5A and TLR2—engineered in both IgG1 and IgG4 formats. Results Human macrophages infected with all four DENV serotypes, as well as reverse genetics DENV2 (rgDV2), exhibit a consistently dominant pro-inflammatory program driven by CLEC5A-TLR2 co-signaling. Treatment with bispecific antibodies, in both IgG1 and IgG4 formats, potently suppress the production of TNF-α, IL-6, IL-8, and MCP-1 production across all viral strains tested. Notably, the IgG4 format preserves robust inhibitory efficacy while minimizing Fc receptor engagement, thereby offering a rational strategy to mitigate the risk of antibody-dependent enhancement (ADE). This dual-targeting approach also remains effective against NS1 mutant strains linked to enhanced virulence and cytokine storm. Conclusion Together, these findings identify CLEC5A-TLR2 co-signaling as a convergent inflammatory axis in dengue pathogenesis and establish bispecific dual-receptor blockade as a mechanistically grounded, ADE-aware immunomodulatory strategy to mitigate cytokine-driven pathology.
Itaconate has garnered significant attention in recent years due to its immunomodulatory and antimicrobial functions. During inflammation and pathogenic infections, itaconate is formed through the decarboxylation of cis-aconitate in the mitochondrial tricarboxylic acid cycle and accumulates in large quantities to counteract excessive inflammation and pathogenic infections. However, many pathogenic bacteria have also evolved pathways to directly degrade itaconate or indirectly resist its stimulatory effects. We first review the research history and metabolic pathways of itaconate. Then we focus on exploring its direct mechanism of inhibiting pathogenic bacteria growth and reproduction by post-translational modification of metabolic enzymes such as isocitrate lyase, aldolase, and IMP dehydrogenase. Additionally, we examine its indirect mechanism of coordinating immune cell functions to eliminate pathogenic bacteria. Pathogenic bacteria counteract this by directly degrading itaconate through the IcT-IcH-CcL cascade reaction or by adapting through metabolic reprogramming to enable chronic infection. Subsequently, we discuss the spatiotemporal specificity of itaconate during the early and late stages of pathogenic bacteria infection, highlighting its role in regulating immune defense strategies at different phases. Finally, we discuss the potential and limitations of itaconate-related interventions as adjunctive strategies for bacterial disease control, particularly in the context of drug-resistant infections. This review elucidates the mechanism of itaconate in host-microbial crosstalk from the perspective of bidirectional resistance between host and bacteria, emphasizing its crucial role as a metabolic messenger in mediating co-evolutionary, co-developmental, and co-metabolic interactions between the host and bacteria. Current evidence of itaconate-mediated bidirectional interactions may help guide future mechanistic studies and the development of itaconate-related adjunctive strategies for bacterial disease control. Further in vivo, clinical, and field validation is still required before these findings can be translated into therapeutic or agricultural applications.
The stiffness of the extracellular matrix (ECM) regulates cellular behavior, influencing tumor progression and therapeutic resistance. In cancer, aberrant ECM stiffening can reduce immune cell infiltration and treatment efficacy, which can in turn alter the tumor microenvironment. Here, we review the role of ECM stiffness in cancer biology and its relevance to matrix-targeted therapies and biomaterial design. We discuss three-dimensional (3D) in vitro models that mimic native tissues and the bidirectional interactions between ECM mechanics and therapeutic interventions. A comparative analysis of measurement modalities is presented for characterizing complex 3D environments, including shear-wave-based techniques such as optical and ultrasound elastography and non-shear-wave-based approaches such as atomic force microscopy and rheology. Future directions include developing matrix-modulating therapies, integrating elasticity sensors into microfluidic devices for higher throughputs and physiological relevance, and applying machine learning to interpret heterogeneous mechanical properties. Collectively, these engineering and biological advances highlight ECM stiffness as a tractable target and open translational opportunities for predictive modeling, diagnostic platforms, and matrix-directed therapies to improve cancer treatment.
Abstract Background ADAMTS1, a disintegrin and metalloproteinase with thrombospondin motif 1, plays a role in inflammation, organogenesis, and ovulation. Up or downregulation of ADAMTS1 has been implicated in tissue remodelling leading to cancer. We analysed the expression of ADAMTS1 in different stages/grades of primary and metastatic serous ovarian tumours and ascites-derived tumour cells from patients and assessed the functional role of ADAMTS1 in ovarian cancer (OC) cell lines. Methods The expression and localisation of ADAMTS1 was assessed by immunohistochemistry (IHC) and Opal Multiplex IHC staining of OC tissues. Functional roles of ADAMTS1 in OC cell lines were assessed by using siRNA-mediated knockdown (KD), MTT assay, cell migration by xCELLigence, cell adhesion, ELISA, qRT-PCR, Western blot, immunofluorescence (IF) and activity of Cdc42 GTPase. Results The expression of ADAMTS1 was significantly enhanced in higher stages/grades of ovarian tumours compared to benign tumours. In high-grade tumours, ADAMTS1 was localised more in the nucleus of epithelial cells while localisation in stromal cells was mostly in the cytoplasm. Significantly higher mRNA expression of ADAMTS1 was noted in epithelial compared to mesenchymal ascites-derived tumour cells. The expression of ADAMTS1 was significantly higher in metastatic high-grade tumours compared to primary tumours. KD of ADAMTS1 expression by siRNA in OC cell lines had no effect on cell proliferation but resulted in decreased cell adhesion, increased cell migration accompanied by increased expression of markers (CDH1 and EPCAM) associated with epithelial plasticity. Remodelling of ECM accompanied by increased intra- and extracellular production of VCAN and enhanced Cdc42 GTPase activity was also noted in cell lines with ADAMTS1 KD. Cdc42 GTPase specific inhibitor, ML141, reversed ADAMTS1 KD-mediated enhanced migration. On the other hand, VCAN KD inhibited ADAMTS1 KD-mediated migration and reversed the effect on cell adhesion. Conclusions These results suggest that the expression of ADAMTS1 progressively enriches ovarian tumours and promotes OC progression. Its knock down in in vitro cell culture impacts ECM remodelling through enhanced Cdc42GTPase activity and VCAN production resulting in epithelial cell plasticity, accelerated migration, and reduced cell adhesion.
Abstract Background Ischemic stroke remains a leading cause of death and disability, with limited therapies addressing its pathophysiology. Neuroinflammation, blood–brain barrier disruption, and immune dysregulation critically contribute to both acute neuronal injury and chronic functional decline. Elucidating the endogenous factors that govern these processes and understanding the intricate interplay among them is essential for developing effective therapeutic strategies. Cortistatin, a neuropeptide expressed in the nervous and immune systems, exhibits potent immunomodulatory actions, but its role in neuroinflammatory diseases remains unclear. Methods In this study, we combined human and murine transcriptomic analyses with a preclinical stroke model in cortistatin-deficient mice to investigate the endogenous and therapeutic roles of cortistatin during the acute (48 h) and subacute (7 days) phases of ischemic stroke. Results We demonstrate for the first time that cortistatin deficiency amplified ischemia-induced transcriptional programs, heightening neuroinflammation and glial/neurovascular dysfunction, and worsening neurological outcomes. Interestingly, cortistatin deficiency accelerated aging-associated pathological features, underscoring its essential role in neuroimmune and cerebrovascular homeostasis. Conversely, cortistatin administration improved neuronal survival, mitigated immune dysfunction, and preserved myelin and vascular integrity during both the acute and subacute stages. Conclusions These findings provide the first evidence identifying cortistatin as an endogenous neuroimmune regulator in ischemic stroke and a promising multimodal therapeutic agent for improving acute and long-term outcomes.
