Search Results (203)

Over the past few decades, Fc fragment-conjugated proteins, such as Protein A, have been extensively utilized across a range of applications, including antibody purification, site-specific immobilization of antibodies, and the development of biosensing platforms. In this study, building upon our group prior research, we designed and screened an affinity DNA functional ligand (A-DNAFL) and experimentally validated its binding affinity (K_D = 6.59 × 10⁻⁸) toward mouse IgG antibodies, whose binding performance was comparable to that of protein A. Systematic evaluations were performed to assess the binding efficiency under varying pH levels and ionic strength conditions. Optimal antibody immobilization was achieved in PBST-B buffer under physiological pH 7.2–7.4 and containing approximately 154 mM Na⁺ and 4 mM K⁺. Two competitive binding assays confirmed that the A-DNAFL binds to the Fc fragment of murine IgG antibody. Furthermore, molecular docking simulations were employed to investigate the interaction mode, revealing key residues involved in binding as well as the contributions of hydrogen bonding and hydrophobic interactions to complex stabilization. Leveraging these insights, A-DNAFL was utilized as a tool for oriented immobilization of antibodies on the sensing interface, enabling the construction of an immunosensor for ricin detection. Following optimization of immobilization parameters, the biosensor exhibited a detection limit of 30.5 ng/mL with the linear regression equation is lg(Response) = 0.329 lg(C_ricin) − 2.027 (N = 9, R = 0.938, p < 0.001)—representing a 64-fold improvement compared to conventional protein A-based methods. The system demonstrated robust resistance to nonspecific interference. Sensing interface reusability was also evaluated, showing only 8.55% signal reduction after two regeneration cycles, indicating that glycine effectively elutes bound antibodies while preserving sensor activity. In summary, the A-DNAFL presented in this study represents a novel antibody-directed immobilization material that serves as a promising alternative to protein A. It offers several advantages, including high modifiability, low production cost, and a relatively small molecular weight. These features collectively contribute to its broad application potential in biosensing, antibody purification, and other areas of life science research. Full article

Cellular senescence, a state of stable cell cycle arrest accompanied by a complex senescence-associated secretory phenotype (SASP), is a fundamental biological process implicated as a key driver of lung aging and lung age-related diseases (LARDs). This review provides a comprehensive overview of the rapidly evolving field of senotyping based on cellular heterogeneity in lung development and aging in health and disease. It also delves into the molecular mechanisms driving senescence and SASP production, highlighting pathways such as p53/p21, p16^INK4a/RB, mTOR, and p38 MAPK as therapeutic targets. The involvement of various novel SASP proteins, such as GDP15, cytokines/chemokines, growth factors, and DNA damage response proteins. We further highlight the effectiveness of senotherapeutics in mitigating the detrimental effects of senescent cell (SnC) accumulation within the lungs. It also outlines two main therapeutic approaches: senolytics, which selectively trigger apoptosis in SnCs, and senomorphics (also known as senostatics), which mitigate the detrimental effects of the SASP without necessarily removing the senescent cells. Various classes of senolytic and senomorphic drugs are currently in clinical trials including natural products (e.g., quercetin, fisetin, resveratrol) and repurposed drugs (e.g., dasatinib, navitoclax, metformin, rapamycin) that has demonstrated therapeutic promise in improving tissue function, alleviating LARDs, and extending health span. We discuss the future of these strategies in lung research and further elaborate upon the usability of novel approaches including HSP90 inhibitors, senolytic CAR-T cells, Antibody drug conjugate and galactose-modified prodrugs in influencing the field of personalized medicine in future. Overall, this comprehensive review highlights the progress made so far and the challenges faced in the field of cellular senescence including SnC heterogeneity, states of senescence, senotyping, immunosenescence, drug delivery, target specificity, long-term safety, and the need for robust cell-based biomarkers. Future perspectives, such as advanced delivery systems, and combination therapies, are considered critical for translating the potential of senotherapeutics into effective clinical applications for age-related pulmonary diseases/conditions. Full article

Tuberculosis (TB), primarily caused by Mycobacterium tuberculosis (Mtb), remains a leading cause of infectious disease mortality worldwide. Global TB control efforts face several hurdles, including the lack of a broadly effective vaccine, limited sensitivity of current diagnostics, particularly for paucibacillary and extrapulmonary TB, and significant adverse effects associated with prolonged small-molecule drug regimens. The growing prevalence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains further underscores the urgent need for innovative therapeutic strategies. We outline characteristics of next-generation TB therapeutics. We show that antibody (Ab)-drug conjugates (ADCs) satisfy many of those desirable characteristics. Since a major hurdle to this approach lies in Mtb-specific Abs, we highlight an open-access resource comprising a broad panel of Mtb-specific mouse monoclonal antibodies targeting key factors involved in Mtb survival, immune evasion, and pathogenesis. These critical Mtb virulence factors include heat shock proteins (GroES, DnaK, and HspX), surface-associated or secreted proteins (LAM, Ag85, HBHA, Mpt64/CFP-21, and PhoS1/PstS1), cell wall/envelope-associated proteins (LprG/p27), and detoxifying enzymes (KatG and SodA). The resource provides full-length sequences of the immunoglobulin variable regions, enabling antibody engineering and facilitating translational TB research across vaccine design, diagnostic development, and immunotherapeutic applications, in addition to ADCs. This ADC targeted delivery strategy holds promise for overcoming TB heterogeneity and eliminating both active and dormant Mtb populations within a single therapeutic formulation and offers a novel avenue for precision TB treatment. Full article

Fatty acid binding proteins FABP3 and FABP4 act as intracellular lipid chaperones that influence fatty acid transport and metabolism in mammary tissue, and genetic variation in these genes may affect milk composition. We examined the associations between FABP3 and FABP4 polymorphisms and milk composition and fatty acid profiles in 200 lactating Native Southern Yellow (NSY) cows. DNA from each cow was PCR-amplified and Sanger-sequenced for FABP3 and FABP4; genotypes were tested for their association with milk fatty acid concentrations and standard composition traits using linear models adjusted for relevant covariates. We detected a missense variant in FABP3 (c.3656G > A; p.Val45Met) and an intronic SNP in FABP4 (g.3509T > C). The FABP3 p.Val45Met AA genotype was associated with higher concentrations of butyric, palmitic, oleic, and α-linolenic acids. Cows with the FABP4 TC genotype exhibited elevated levels of myristoleic, γ-linolenic, conjugated linoleic, and arachidic acids, along with increased fat-free dry matter, protein, and lactose. In silico analyses provided mixed evidence for the structural effects of p.Val45Met, molecular docking suggested altered ligand affinity for several fatty acids, and splice site prediction implicated g.3509T > C in possible transcript processing changes. These variants constitute candidate markers for milk fatty acid composition in NSY cattle; replication in independent cohorts and functional validation are recommended to confirm their utility for milk quality improvement. Full article

Effective treatment of ovarian cancer is limited by late-stage detection and chemotherapy resistance. There is a clinical need for the discovery of novel molecular targets to enable the development of innovative theranostic approaches. We investigated the coagulation factor II receptor/protease-activated receptor 1 (F2R/PAR1) as a potential diagnostic/prognostic biomarker and therapeutic target for ovarian cancer treatment. Public RNA sequence and DNA microarray data were used to analyze F2R gene expression in ovarian cancers, with protein expression confirmed in tumor samples by flow cytometry, immunofluorescence, and immunohistochemistry (IHC). Functional assays were conducted to study effects of F2R suppression on tumor progression. Our analysis confirmed elevated F2R mRNA and protein expression in ovarian cancers, notably in patients with metastatic and chemotherapy-resistant disease. Kaplan–Meier survival analysis demonstrated an association between high F2R protein detection and reduced progression-free survival. F2R suppression in ovarian cancer cell lines reduced tumor cell motility, invasion, spheroid formation, and metabolism and enhanced carboplatin sensitivity. F2R is a compelling diagnostic/prognostic and therapeutic target that could be used to treat chemotherapy-resistant and metastatic disease. The evaluation of novel F2R targeting strategies, using antibody-conjugated drugs or F2R ligand-decorated drug carriers, could lead to the development of effective therapeutics for patients with ovarian cancer. Full article

Background/Objectives: The conventional treatment of glioblastoma (GBM) with alkylating agents is not curative. The protein O6-methylguanine DNA methyltransferase (MGMT) is a significant limitation, being able to repair drug-induced DNA damage. Thus, exploring non-alkylating agents already approved by the FDA is imperative. The antidepressant fluoxetine (FL) has been explored due to its anti-cancer properties. However, its first-pass effect and its non-targeted distribution to brain tissue are major limitations of FL’s administration, which is conventionally orally administered. Thus, the primary objective of this work was the development of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) tailored with folic acid (FA) for FL delivery to GBM cells. Methods: A Central Composite Design (CCD) was applied to optimize the NPs. Results: The developed FA-functionalized PLGA NPs exhibited physicochemical properties suitable for brain-targeted delivery. The final formulation presented an average diameter of 167 ± 8 nm, a polydispersity index (PdI) of 0.23 ± 0.07, and a zeta potential of −22.2 ± 0.3 mV. The encapsulation efficiency (EE) and loading capacity (LC) values were 44.4 ± 3.8% and 3.1 ± 0.3%, respectively. In vitro studies demonstrated that the NPs are stable in storage and simulated physiological conditions and can maintain a controlled and slow-release profile of FL for 17 days. In vitro cell uptake experiments demonstrated that conjugation with FA enhances the NPs’ internalization in GBM cells overexpressing folate receptors through endocytosis mediated by this receptor. Furthermore, in vitro cytotoxicity experiments demonstrated that the FL encapsulation in the developed NPs maintains drug efficacy, as well as it was able to increase cell sensitivity to treatment with an alkylating agent. Conclusions: These results suggest that the developed NPs are effective nanocarriers, either as a standalone therapy or as a chemosensitizer in combination with the standard GBM treatment. Full article

Antioxidative neuroprotection is effective at preventing ischemic stroke (IS). Ferulic acid (FA) offers benefits in the treatment of many diseases, mostly due to its antioxidant activities. In this study, a glycosylated ferulic acid conjugate (FA-Glu), with 1,2,3-triazole as a linker and bioisostere between glucose at the C6 position and FA at the C4 position, was designed and synthesized. The hydrophilicity and chemical stability of FA-Glu were tested. FA-Glu’s protection against DNA oxidative cleavage was tested using pBR322 plasmid DNA under the Fenton reaction. The cytotoxicity of FA-Glu was examined via the PC12 cell and bEnd.3 cell tests. Antioxidative neuroprotection was evaluated, in vitro, via a H₂O₂-induced PC12 cell test, measuring cell viability and ROS levels. Antioxidative alleviation of cerebral ischemia–reperfusion injury (CIRI), in vivo, was evaluated using a rat middle cerebral artery occlusion (MCAO) model. The results indicated that FA-Glu was water-soluble (LogP −1.16 ± 0.01) and chemically stable. FA-Glu prevented pBR322 plasmid DNA cleavage induced via •OH radicals (SC% 88.00%). It was a non-toxic agent based on PC12 cell and bEnd.3 cell tests results. FA-Glu significantly protected against H₂O₂-induced oxidative damage in the PC12 cell (cell viability 88.12%, 100 μM) and inhibited excessive cell ROS generation (45.67% at 100 μM). FA-Glu significantly reduced the infarcted brain areas measured using TTC stain observation, quantification (FA-Glu 21.79%, FA 28.49%, I/R model 43.42%), and H&E stain histological observation. It sharply reduced the MDA level (3.26 nmol/mg protein) and significantly increased the GSH level (139.6 nmol/mg protein) and SOD level (265.19 U/mg protein). With superior performance to FA, FA-Glu is a safe agent with effective antioxidative DNA and neuronal protective actions and an ability to alleviate CIRI, which should help in the prevention of IS. Full article

Despite the development of a wide plethora of different anticancer agents, most of them are not used for patient treatment due to adverse effects caused by untargeted cytotoxicity. To prevent this unwanted toxicity, it is necessary to develop therapies discriminating between healthy and cancerous cells. One possible method is to target proteins overexpressed in cancer but not in normal cells. CD320 is a receptor responsible for the uptake of the transcobalamin-bound fraction of vitamin B12 (cobalamin), which is necessary for DNA synthesis, and thus, cell proliferation. CD320 was shown to be overexpressed in many cancers and its potential role as an early cancer biomarker was confirmed in several studies. Consequently, CD320 may represent a promising anti-cancer therapy target. This review summarizes the current advances and perspectives of anti-cancer CD320 targeting therapy, including therapeutic conjugates of vitamin B12, CD320-specific antibodies and nanobodies, nanoparticles loaded with cytotoxic drugs, porphyrin, and the potential of targeted CD320 therapy in attenuation of tumor tissues. Given the growing interest in CD320 as a novel target for anti-cancer therapy, further in vivo studies are required for the investigation of CD320 targeting effects on systemic cytotoxicity. Full article

Background/Objectives: Ovarian cancer is the most lethal gynecologic malignancy due to its late diagnosis, aggressive disease course, and high likelihood of recurrence. In the last few years, with the advent of high-throughput genomic methodologies, our understanding of ovarian cancer genetics and biology has grown. In this review, we discuss current monitoring techniques, as well as biomarker-directed therapies, recently developed for ovarian cancer treatment. Methods: The primary literature and review articles were obtained through PUBMED searches of “ovarian cancer”, “biomarkers”, “CA125”, “circulating tumor DNA”, “BRCA”, “HER2”, “TROP2”, and “FOLR1.” Results and Conclusions: The detection and quantification of CA125, a protein biomarker, remains the primary test used in the clinic for ovarian cancer diagnosis and monitoring. However, liquid biopsy techniques involving circulating tumor DNA, used alone or in combination with CA125, are increasingly used to enhance diagnostic accuracy and provide a more comprehensive picture of tumor genomic changes, including single-nucleotide variants, copy number variations, and epigenetic alterations. In the last few years, the use of BRCA, HER2, TROP2, and FOLR1 as biomarkers for targeted treatment has demonstrated promising results, both preclinically and clinically. The detection of BRCA1/2 mutations is routinely used as a strong predictor of response to PARP inhibitors, while HER2, TROP2, and FOLR1 expressions have emerged as primary targets for the treatment of recurrent ovarian cancer patients using novel antibody–drug conjugates (ADCs). Full article

Background/Objectives: Gene therapy has emerged as a promising strategy for treating a wide range of diseases. However, a major challenge remains in developing efficient and safe delivery systems for genetic material. Nanoparticles, particularly chitosan nanoparticles (CNPs), have gained significant attention as a potential solution. This study focuses on designing a SARS-CoV-2 plasmid DNA (pDNA) conjugated with CNPs and evaluating its in vitro delivery efficiency. Methods: The Omicron Spike DNA sequence was inserted into the pIRES2-eGFP expression vector, and CNPs were synthesized with optimized physicochemical properties to enhance stability, cellular uptake, and transfection efficiency. The conjugate was characterized using UV-Vis, FT-IR, DLS, and TEM techniques. Transfection efficiency was assessed and compared to the commercially available TurboFect reagent as a control. Results: CNPs-pDNA polyplexes with an average size of 159.0 ± 33.1 nm (TEM), a zeta potential of +19.7 ± 0.3 mV, and 100% ± 0.0 encapsulation efficiency were developed as a non-viral delivery system. CNPs efficiently serve as a delivery vehicle for the constructed pDNA without altering cell morphology, achieving transfection efficiencies of 62–74%, compared to 55–70% for TurboFect. Furthermore, RT-qPCR confirmed the expression of Spike mRNA, and Western blot assays validated the expression of Spike protein. Notably, Spike protein expression from CNPs was found to be two-fold higher than the control at 96 h post-transfection. Conclusions: These findings suggest that CNPs are a promising and versatile platform for delivering genetic material. Importantly, this study highlights the intrinsic properties of chitosan, without the use of additional ligands, as a key factor in achieving efficient gene delivery. Full article

Mesothelioma is characterized by the inactivation of tumor suppressor genes, with frequent mutations in neurofibromin 2 (NF2), BRCA1-associated protein 1 (BAP1), and cyclin-dependent kinase inhibitor 2A (CDKN2A). These mutations lead to disruptions in the Hippo signaling pathway and histone methylation, thereby promoting tumor growth. NF2 mutations result in Merlin deficiency, leading to uncontrolled cell proliferation, whereas BAP1 mutations impair chromatin remodeling and hinder DNA damage repair. Emerging molecular targets in mesothelioma include mesothelin (MSLN), oxytocin receptor (OXTR), protein arginine methyltransferase (PRMT5), and carbohydrate sulfotransferase 4 (CHST4). MSLN-based therapies, such as antibody–drug conjugates and immunotoxins, have shown efficacy in clinical trials. OXTR, upregulated in mesothelioma, is correlated with poor prognosis and represents a novel therapeutic target. PRMT5 inhibition is being explored in tumors with MTAP deletions, commonly co-occurring with CDKN2A loss. CHST4 expression is associated with improved prognosis, potentially influencing tumor immunity. Immune checkpoint inhibitors targeting PD-1/PD-L1 have shown promise in some cases; however, resistance mechanisms remain a challenge. Advances in multi-omics approaches have improved our understanding of mesothelioma pathogenesis. Future research will aim to identify novel therapeutic targets and personalized treatment strategies, particularly in the context of epigenetic therapy and combination immunotherapy. Full article

Exogenous abscisic acid (ABA) and brassinosteroid (BR) play important roles in alleviating cold stress in maize. In this study, two maize inbred lines with differing cold tolerance were treated with exogenous ABA, BR, and their combined solution under cold stress conditions at 10 °C to investigate the effects of these treatments on the physiological characteristics of maize seedlings. The results indicated that cold stress significantly inhibited the growth of maize seedlings. Exogenous hormone treatments enhanced antioxidant enzyme activities and promoted the synthesis of osmolytes, thereby alleviating cold stress; however, the combined treatment (AR) did not significantly improve maize cold tolerance. Transcriptomic analysis revealed that pathways including plant hormone signal transduction, fatty acid elongation, and phenylpropanoid biosynthesis were involved in the interaction between ABA and BR. Weighted gene co-expression network analysis (WGCNA) identified four key candidate genes responsive to exogenous ABA and BR under cold stress, namely Zm00001eb343270, Zm00001eb401890, Zm00001eb206790, and Zm00001eb199820. Based on the gene annotation results, we speculate that ubiquitin-conjugating enzyme E2 O, tubulin–tyrosine ligase-like protein 12, the negative regulator of systemic acquired resistance SNI1, and mRNA stability regulators in response to DNA damage may be involved in regulating maize cold tolerance. These findings provide further evidence for the regulatory mechanisms by which exogenous ABA and BR affect maize cold tolerance and elucidate their interaction under cold stress. Full article

Despite decades of research, cancer continues to be a disease of great concern to millions of people around the world. It has been responsible for a total of 609,820 deaths in the U.S. alone in 2023. Over the years, many drugs have been developed to remove or reduce the disease’s impact, all with varying mechanisms of action and side effects. One class of these drugs is small-molecule mitotic inhibitors. These drugs inhibit cancer cell mitosis or self-replication, impeding cell proliferation and eventually leading to cell death. In this paper, small-molecule mitotic inhibitors are discussed and classified through their discovery, underlying chemistry, and mechanism(s) of action. The binding/inhibition of microtubule-related proteins, DNA damage through the inhibition of Checkpoint Kinase 1 protein, and the inhibition of mitotic kinase proteins are discussed in terms of their anticancer activity to provide an overview of a variety of mitotic inhibitors currently commercially available or under investigation, including those in ongoing clinical trial. Clinical trials for anti-mitotic agents are discussed to track research progress, gauge current understanding, and identify possible future prospects. Additionally, antibody–drug conjugates that use mitotic inhibitors as cytotoxic payloads are discussed as possible ways of administering effective anticancer treatments with minimal toxicity. Full article

The genes that encode the universal stress protein (USP) family domain (pfam00582) aid the survival of bacteria in specific host or habitat-induced stress conditions. Genome sequencing revealed that the genome of Helicobacter pylori, a gastric cancer pathogen, typically contains one USP gene, while related helicobacters have one or two distinct USP genes. However, insights into the functions of Helicobacteraceae (Helicobacter and Wolinella) USP genes are still limited to inferences from large-scale genome sequencing. Thus, we have combined bioinformatics and visual analytics approaches to conduct a more comprehensive data investigation of a set of 1045 universal stress protein sequences encoded in 1014 genomes including 785 Helicobacter pylori genomes. The study generated a representative set of 183 USP sequences consisting of 180 Helicobacter sequences, two Wolinella succinogenes sequences, and a sequence from a related campylobacteria. We used the amino acid residues and positions of the 12 possible functional sites in 1030 sequences to identify 25 functional sites patterns for guiding studies on functional interactions of Helicobacteraceae USPs with ATP and other molecules. Genomic context searches and analysis identified USP genes of gastric and enterohepatic helicobacters that are adjacent or in operons with genes for proteins responsive to DNA-damaging oxidative stress (ATP-dependent proteases: ClpS and ClpA); and DNA uptake proteins (natural competence for transformation proteins: ComB6, ComB7, ComB8, ComB9, ComB10, ComBE, and conjugative transfer signal peptidase TraF). Since transcriptomic evidence indicates that oxidative stress and the presence of virulence-associated genes regulate the transcription of H. pylori USP gene, we recommend further research on Helicobacter USP genes and their neighboring genes in oxidative stress response and virulence of helicobacters. To facilitate the reuse of data and research, we produced interactive analytics resources of a dataset composed of values for variables including phylogeography of H. pylori strains, protein sequence features, and gene neighborhood. Full article

Background. Antibiotic-resistant (AR) bacteria pose an increasing threat to public health, but the dynamics of antibiotic resistance gene (ARG) spread in complex microbial communities are poorly understood. Conjugation is a predominant direct cell-to-cell mechanism for the horizontal gene transfer (HGT) of ARGs. We hypothesized that commensal Escherichia coli donor strains would mediate the conjugative transfer of ARGs to phylogenetically distinct bacteria without antibiotic selection pressure in gastrointestinal tracts of mice carrying a human-derived microbiota with undetectable levels of E. coli. Our objective was to identify a mouse model to study the factors regulating AR transfer by conjugation in the gut. Methods. Two donor E. coli strains were engineered to carry chromosomally encoded red fluorescent protein, and an ARG- and green fluorescent protein (GFP)-encoding broad host range RP4 conjugative plasmid. Mice were orally gavaged with two donor strains (1) E. coli MG1655 or (2) human-derived mouse-adapted E. coli LM715-1 and their colonization assessed by culture over time. Fluorescence-activated cell sorting (FACS) and 16S rDNA sequencing were performed to trace plasmid spread to the microbiota. Results. E. coli LM715-1 colonized mice for ten days, while E. coli MG1655 was not recovered after 72 h. Bacterial cells from fecal samples on days 1 and 3 post inoculation were sorted by FACS. Samples from mice given donor E. coli LM715-1 showed an increase in cells expressing green but not red fluorescence compared to pre-inoculation samples. 16S rRNA gene sequencing analysis of FACS GFP positive cells showed that bacterial families Lachnospiraceae, Clostridiaceae, Pseudomonadaceae, Rhodanobacteraceae, Erysipelotrichaceae, Oscillospiraceae, and Butyricicoccaceae were the primary recipients of the RP4 plasmid. Conclusions. Results show this ARG-bearing conjugative RP4 plasmid spread to diverse human gut bacterial taxa within a live animal where they persisted. These fluorescent marker strategies and human-derived microbiota transplanted mice provided a tractable model for investigating the dynamic spread of ARGs within gut microbiota and could be applied rigorously to varied microbiotas to understand conditions facilitating their spread. Full article