ABSTRACT
BACKGROUND: Mediators, genomic and epigenomic characteristics involving in metabolism of arachidonic acid by cyclooxygenase (COX) and lipoxygenase (ALOX) and hepatic activation of clopidogrel have been individually suggested as factors associated with resistance against aspirin and clopidogrel. The present multi-center prospective cohort study evaluated whether the mediators, genomic and epigenomic characteristics participating in arachidonic acid metabolism and clopidogrel activation could be factors that improve the prediction of the aspirin and clopidogrel resistance in addition to cardiovascular risks. METHODS: We enrolled 988 patients with transient ischemic attack and ischemic stroke who were evaluated for a recurrence of ischemic stroke to confirm clinical resistance, and measured aspirin (ARU) and P2Y12 reaction units (PRU) using VerifyNow to assess laboratory resistance 12 weeks after aspirin and clopidogrel administration. We investigated whether mediators, genotypes, and promoter methylation of genes involved in COX and ALOX metabolisms and clopidogrel activation could synergistically improve the prediction of ischemic stroke recurrence and the ARU and PRU levels by integrating to the established cardiovascular risk factors. RESULTS: The logistic model to predict the recurrence used thromboxane A synthase 1 (TXAS1, rs41708) A/A genotype and ALOX12 promoter methylation as independent variables, and, improved sensitivity of recurrence prediction from 3.4% before to 13.8% after adding the mediators, genomic and epigenomic variables to the cardiovascular risks. The linear model we used to predict the ARU level included leukotriene B4, COX2 (rs20417) C/G and thromboxane A2 receptor (rs1131882) A/A genotypes with the addition of COX1 and ALOX15 promoter methylations as variables. The linear PRU prediction model included G/A and prostaglandin I receptor (rs4987262) G/A genotypes, COX2 and TXAS1 promoter methylation, as well as cytochrome P450 2C19*2 (rs4244285) A/A, G/A, and *3 (rs4986893) A/A genotypes as variables. The linear models for predicting ARU (r = 0.291, R2 = 0.033, p < 0.01) and PRU (r = 0.503, R2 = 0.210, p < 0.001) levels had improved prediction performance after adding the genomic and epigenomic variables to the cardiovascular risks. CONCLUSIONS: This study demonstrates that different mediators, genomic and epigenomic characteristics of arachidonic acid metabolism and clopidogrel activation synergistically improved the prediction of the aspirin and clopidogrel resistance together with the cardiovascular risk factors. TRIAL REGISTRATION: URL: https://www. CLINICALTRIALS: gov ; Unique identifier: NCT03823274.
Subject(s)
Aspirin , Clopidogrel , Drug Resistance , Humans , Clopidogrel/therapeutic use , Clopidogrel/pharmacology , Male , Female , Aspirin/therapeutic use , Aspirin/pharmacology , Drug Resistance/genetics , Middle Aged , Aged , Epigenomics , Genomics , Prospective Studies , Platelet Aggregation Inhibitors/therapeutic use , Platelet Aggregation Inhibitors/pharmacology , DNA Methylation/drug effectsABSTRACT
[This corrects the article DOI: 10.2196/50259.].
ABSTRACT
This study proposes a solid-state two-dimensional beam-steering device based on an electro-optical phased array (EOPA) in thin-film lithium niobate (TFLN) and silicon nitride (SiN) hybrid platforms, thereby eliminating the requirement for the direct etching of TFLN. Electro-optic (EO) phase modulator array comprises cascaded multimode interference couplers with an SiN strip-loaded TFLN configuration, which is designed and fabricated via i-line photolithography. Each EO modulator element with an interaction region length of 1.56â cm consumed a minimum power of 3.2 pJ/π under a half-wave voltage of 3.64â V and had an estimated modulation speed of 1.2â GHz. Subsequently, an SiN dispersive antenna with a waveguide grating was tethered to the modulator array to form an EOPA, facilitating the out-of-plane radiation of highly defined near-infrared beams. A prepared EOPA utilized EO phase control and wavelength tuning near 1550â nm to achieve a field-of-view of 22° × 5° in the horizontal and vertical directions. The proposed hybrid integrated platform can potentially facilitate low-power and high-speed beam steering.
ABSTRACT
BACKGROUND: Limited awareness, social stigma, and access to mental health professionals hinder early detection and intervention of internet gaming disorder (IGD), which has emerged as a significant concern among young individuals. Prevalence estimates vary between 0.7% and 15.6%, and its recognition in the International Classification of Diseases, 11th Revision and Diagnostic and Statistical Manual of Mental Disorders, 5th Edition underscores its impact on academic functioning, social isolation, and mental health challenges. OBJECTIVE: This study aimed to uncover digital phenotypes for the early detection of IGD among adolescents in learning settings. By leveraging sensor data collected from student tablets, the overarching objective is to incorporate these digital indicators into daily school activities to establish these markers as a mental health screening tool, facilitating the early identification and intervention for IGD cases. METHODS: A total of 168 voluntary participants were engaged, consisting of 85 students with IGD and 83 students without IGD. There were 53% (89/168) female and 47% (79/168) male individuals, all within the age range of 13-14 years. The individual students learned their Korean literature and mathematics lessons on their personal tablets, with sensor data being automatically collected. Multiple regression with bootstrapping and multivariate ANOVA were used, prioritizing interpretability over predictability, for cross-validation purposes. RESULTS: A negative correlation between IGD Scale (IGDS) scores and learning outcomes emerged (r166=-0.15; P=.047), suggesting that higher IGDS scores were associated with lower learning outcomes. Multiple regression identified 5 key indicators linked to IGD, explaining 23% of the IGDS score variance: stroke acceleration (ß=.33; P<.001), time interval between keys (ß=-0.26; P=.01), word spacing (ß=-0.25; P<.001), deletion (ß=-0.24; P<.001), and horizontal length of strokes (ß=0.21; P=.02). Multivariate ANOVA cross-validated these findings, revealing significant differences in digital phenotypes between potential IGD and non-IGD groups. The average effect size, measured by Cohen d, across the indicators was 0.40, indicating a moderate effect. Notable distinctions included faster stroke acceleration (Cohen d=0.68; P=<.001), reduced word spacing (Cohen d=.57; P=<.001), decreased deletion behavior (Cohen d=0.33; P=.04), and longer horizontal strokes (Cohen d=0.34; P=.03) in students with potential IGD compared to their counterparts without IGD. CONCLUSIONS: The aggregated findings show a negative correlation between IGD and learning performance, highlighting the effectiveness of digital markers in detecting IGD. This underscores the importance of digital phenotyping in advancing mental health care within educational settings. As schools adopt a 1-device-per-student framework, digital phenotyping emerges as a promising early detection method for IGD. This shift could transform clinical approaches from reactive to proactive measures.
Subject(s)
Early Diagnosis , Internet Addiction Disorder , Students , Adolescent , Female , Humans , Male , Internet Addiction Disorder/epidemiology , Internet Addiction Disorder/diagnosis , Phenotype , Republic of Korea/epidemiology , Students/psychologyABSTRACT
BACKGROUND: In patients with non-small cell lung cancer (NSCLC), interstitial lung abnormalities (ILA) have been linked to mortality and can be identified on computed tomography (CT) scans. In the present study we aimed to evaluate the predictive value of automatically quantified ILA based on the Fleischner Society definition in patients with stage I NSCLC. METHODS: We retrospectively reviewed 948 patients with pathological stage I NSCLC who underwent pulmonary resection between April 2009 and October 2022. A commercially available deep learning-based automated quantification program for ILA was used to evaluate the preoperative CT data. The Fleischner Society definition, quantitative results, and interdisciplinary discussion led to the division of patients into normal and ILA groups. The sum of the fibrotic and nonfibrotic ILA components constituted the total ILA component and more than 5%. RESULTS: Of the 948 patients with stage I NSCLC, 99 (10.4%) patients had ILA. Shorter overall survival and recurrence-free survival was associated with the presence of ILA. After controlling for confounding variables, the presence of ILA remained significant for increased risk of death (hazard ratio [HR] = 3.09; 95% confidence interval [CI]: 1.91-5.00; p < 0.001) and the presence of ILA remained significant for increased recurrence (HR = 1.96; 95% CI: 1.16-3.30; p = 0.012). CONCLUSIONS: The automated CT quantification of ILA, based on the Fleischner Society definition, was significantly linked to poorer survival and recurrence in patients with stage I NSCLC.
Subject(s)
Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , Neoplasm Staging , Tomography, X-Ray Computed , Humans , Carcinoma, Non-Small-Cell Lung/pathology , Carcinoma, Non-Small-Cell Lung/surgery , Carcinoma, Non-Small-Cell Lung/diagnostic imaging , Male , Female , Lung Neoplasms/pathology , Lung Neoplasms/surgery , Lung Neoplasms/diagnostic imaging , Lung Neoplasms/mortality , Prognosis , Tomography, X-Ray Computed/methods , Retrospective Studies , Aged , Middle Aged , Lung Diseases, Interstitial/pathology , Lung Diseases, Interstitial/diagnostic imagingABSTRACT
Macroautophagy/autophagy is a complex degradation process with a dual role in cell death that is influenced by the cell types that are involved and the stressors they are exposed to. Ferroptosis is an iron-dependent oxidative form of cell death characterized by unrestricted lipid peroxidation in the context of heterogeneous and plastic mechanisms. Recent studies have shed light on the involvement of specific types of autophagy (e.g. ferritinophagy, lipophagy, and clockophagy) in initiating or executing ferroptotic cell death through the selective degradation of anti-injury proteins or organelles. Conversely, other forms of selective autophagy (e.g. reticulophagy and lysophagy) enhance the cellular defense against ferroptotic damage. Dysregulated autophagy-dependent ferroptosis has implications for a diverse range of pathological conditions. This review aims to present an updated definition of autophagy-dependent ferroptosis, discuss influential substrates and receptors, outline experimental methods, and propose guidelines for interpreting the results.Abbreviation: 3-MA:3-methyladenine; 4HNE: 4-hydroxynonenal; ACD: accidentalcell death; ADF: autophagy-dependentferroptosis; ARE: antioxidant response element; BH2:dihydrobiopterin; BH4: tetrahydrobiopterin; BMDMs: bonemarrow-derived macrophages; CMA: chaperone-mediated autophagy; CQ:chloroquine; DAMPs: danger/damage-associated molecular patterns; EMT,epithelial-mesenchymal transition; EPR: electronparamagnetic resonance; ER, endoplasmic reticulum; FRET: Försterresonance energy transfer; GFP: green fluorescent protein;GSH: glutathione;IF: immunofluorescence; IHC: immunohistochemistry; IOP, intraocularpressure; IRI: ischemia-reperfusion injury; LAA: linoleamide alkyne;MDA: malondialdehyde; PGSK: Phen Green™ SK;RCD: regulatedcell death; PUFAs: polyunsaturated fatty acids; RFP: red fluorescentprotein;ROS: reactive oxygen species; TBA: thiobarbituricacid; TBARS: thiobarbituric acid reactive substances; TEM:transmission electron microscopy.
Subject(s)
Autophagy , Ferroptosis , Ferroptosis/physiology , Humans , Autophagy/physiology , Animals , ConsensusABSTRACT
The emergence of antibiotic-resistant bacteria (ARBs) and genes (ARGs) in aquaculture underscores the urgent need for alternative veterinary strategies to combat antimicrobial resistance (AMR). These measures are vital to reduce the likelihood of entering a post-antibiotic era. Identifying environmentally friendly biotechnological solutions to prevent and treat bacterial diseases is crucial for the sustainability of aquaculture and for minimizing the use of antimicrobials, especially antibiotics. The development of probiotics with quorum-quenching (QQ) capabilities presents a promising non-antibiotic strategy for sustainable aquaculture. Recent research has demonstrated the effectiveness of QQ probiotics (QQPs) against a range of significant fish pathogens in aquaculture. QQ disrupts microbial communication (quorum sensing, QS) by inhibiting the production, replication, and detection of signalling molecules, thereby reducing bacterial virulence factors. With their targeted anti-virulence approach, QQPs have substantial promise as a potential alternative to antibiotics. The application of QQPs in aquaculture, however, is still in its early stages and requires additional research. Key challenges include determining the optimal dosage and treatment regimens, understanding the long-term effects, and integrating QQPs with other disease control methods in diverse aquaculture systems. This review scrutinizes the current literature on antibiotic usage, AMR prevalence in aquaculture, QQ mechanisms and the application of QQPs as a sustainable alternative to antibiotics.
Subject(s)
Aquaculture , Fish Diseases , Probiotics , Quorum Sensing , Quorum Sensing/drug effects , Aquaculture/methods , Probiotics/pharmacology , Animals , Fish Diseases/prevention & control , Fish Diseases/microbiology , Fishes , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/administration & dosage , Drug Resistance, BacterialABSTRACT
The implications of administration of mRNA vaccines to individuals with chronic inflammatory diseases, including myocarditis, rheumatoid arthritis (RA), and inflammatory bowel disease (IBD), are unclear. We investigated mRNA vaccine effects in a chronic inflammation mouse model implanted with an LPS pump, focusing on toxicity and immunogenicity. Under chronic inflammation, mRNA vaccines exacerbated cardiac damage and myocarditis, inducing mild heart inflammation with heightened pro-inflammatory cytokine production and inflammatory cell infiltration in the heart. Concurrently, significant muscle damage occurred, with disturbances in mitochondrial fusion and fission factors signaling impaired muscle repair. However, chronic inflammation did not adversely affect muscles at the vaccination site or humoral immune responses; nevertheless, it partially reduced the cell-mediated immune response, particularly T-cell activation. These findings underscore the importance of addressing mRNA vaccine toxicity and immunogenicity in the context of chronic inflammation, ensuring their safe and effective utilization, particularly among vulnerable populations with immune-mediated inflammatory diseases.
ABSTRACT
Epizootics of Koi herpesvirus (KHV) cause mass mortality in koi carp (Cyprinus rubrofuscus) and common carp (Cyprinus carpio) worldwide. Rapid and accurate virus detection technology is crucial for preventing pathogen spread and minimizing damage. Although several diagnostic assays have been developed for KHV, the analytical and diagnostic performance of the detection methods has not been evaluated. In this study, we developed and validated the diagnostic performance of two molecular diagnostic assays, cross-priming amplification-based lateral flow assay (CPA-LFA) and TaqMan probe-based real-time polymerase chain reaction (PCR). To detect KHV, primers and probe were designed based on the thymidine kinase (TK) genes. The detection limits of developed CPA-LFA and real-time PCR assays were determined to be 675.69 copies/µL and 8.384 copies/µL, respectively. The diagnostic sensitivity and specificity of the developed assay were determined using fish samples (n = 179). CPA-LFA was found to be 93.67% and 100%, respectively, and real-time PCR was found to be 100% and 100%, respectively. Therefore, the newly developed CPA-LFA and real-time PCR assays accurately and rapidly detect KHV. CPA-LFA is particularly suitable for point-of-care diagnosis because of its simple diagnostic process, and real-time PCR analysis is most suitable for precise diagnosis because it can detect low viral loads.
Subject(s)
Carps , Fish Diseases , Herpesviridae Infections , Herpesviridae , Animals , Herpesviridae Infections/diagnosis , Herpesviridae Infections/veterinary , Real-Time Polymerase Chain Reaction , Cross-Priming , Fish Diseases/diagnosis , Herpesviridae/geneticsABSTRACT
Objectives: This study aims to evaluate the practicality of handheld digital X-ray in determining the position of the electrode array following Cochlear implantation (CI). Methods: A retrospective study was conducted involving 11 patients (12 ears) who underwent intraoperative imaging via handheld X-ray (MINE ALNU®, OTOM, Gwangju, South Korea) post-CI between December 2021 and January 2023. Immediate confirmation of the correct electrode array placement in the cochlea was achieved, with subsequent comparisons made to C-arm image and postoperative transorbital view X-ray. Results: Rapid intraoperative imaging was achieved in all instances. The electrode types used included 9 Nucleus slim modiolar electrodes, 1 Nucleus contour electrode, and 2 Medel flex26 electrodes. A malpositioned electrode array was detected in one patient. The handheld digital X-ray also adeptly visualized the electrodes implanted in pediatric patients. Conclusions: The use of intraoperative handheld digital X-ray using MINE ALNU® proves to be a safe, efficient, straightforward, and reliable method for immediate identification of an inserted electrode array. It has potential to replace the traditional C-arm X-ray for verifying electrode positioning in the operating room.
ABSTRACT
BACKGROUND: Acral lentiginous melanoma (ALM), a cutaneous melanoma subtype, exhibits a poorer prognosis than nonacral cutaneous melanoma (NACM). The neutrophil-to-lymphocyte ratio (NLR) is emerging as a prognostic indicator across diverse cancers. OBJECTIVE: We explored the baseline NLR disparities between ALM and NACM, and the NLR's prognostic significance in patients with ALM. METHODS: We reviewed records of patients with ALM and NACM diagnosed between 1997 and 2022, analyzing medical data. RESULTS: Among 327 and 159 patients with ALM and NACM, respectively, baseline NLR varied based on distinct clinicopathologic factors between ALM and NACM. In stage 3 to 4 melanomas, the median NLR for ALM (2.18; IQR, 1.70-3.08) significantly surpassed NACM (1.74; IQR, 1.33-2.53) (P = .029). In patients with ALM, high NLR (hazard ratio, 1.64; 95% CI, 1.02-2.66; P = .043) was independently correlated with poor progression-free survival when adjusting for ulceration, Breslow thickness of ≥2 mm, and nodal invasion. LIMITATIONS: Single-center, retrospective design. CONCLUSION: Advanced-stage ALM exhibited a significantly higher baseline NLR compared with that of NACM. Evaluating baseline NLR could provide valuable prognostic insights for patients with ALM.
Subject(s)
Melanoma , Skin Neoplasms , Humans , Melanoma/pathology , Skin Neoplasms/pathology , Prognosis , Retrospective Studies , Neutrophils/pathology , Lymphocytes/pathologyABSTRACT
Vibrio harveyi strain 22FBVib0145 was isolated from a diseased olive flounder farmed in Jeju, Korea. Here, we report the draft genome sequence of this strain. It is 6,238,277 bp in length with a G + C content of 44.8%.
ABSTRACT
Efficient management of low energy states is vital for cells to maintain basic functions and metabolism and avoid cell death. While autophagy has long been considered a critical mechanism for ensuring survival during energy depletion, recent research has presented conflicting evidence, challenging the long-standing concept. This recent development suggests that cells prioritize preserving essential cellular components while restraining autophagy induction when cellular energy is limited. This essay explores the conceptual discourse on autophagy regulation during energy stress, navigating through the studies that established the current paradigm and the recent research that has challenged its validity while proposing an alternative model. This exploration highlights the far-reaching implications of the alternative model, which represents a conceptual departure from the established paradigm, offering new perspectives on how cells respond to energy stress.
Subject(s)
AMP-Activated Protein Kinases , Autophagy , AMP-Activated Protein Kinases/metabolism , Autophagy/physiologyABSTRACT
Macroautophagy/autophagy research often involves overexpressing proteins to investigate their localization, function and activity. However, this approach can disturb the inherent balance of cellular components, potentially affecting the integrity of the autophagy process. With the advent of genome-editing techniques like CRISPR-Cas9, it is now possible to tag endogenous proteins with fluorescent markers, enabling the study of their behaviors under more physiologically relevant conditions. Nevertheless, conventional microscopy methods have limitations in characterizing the behaviors of proteins expressed at endogenous levels. This challenge can be overcome by single-molecule localization microscopy (SMLM) methods, which provide single-molecule sensitivity and super-resolution imaging capabilities. In our recent study, we used SMLM in combination with genome editing to explore the behavior of endogenous ULK1 during autophagy initiation, yielding unprecedented insights into the autophagy initiation process.Abbreviation: ATG13: autophagy related 13; ATG14: autophagy related 14; ATG16L1: autophagy related 16 like 1; BECN1: beclin 1; ER: endoplasmic reticulum; GABARAPL1: GABA type A receptor associated protein like 1; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MTORC1: mechanistic target of rapamycin kinase complex 1; PALM: photo-activated localization microscopy; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PIK3R4/VPS15: phosphoinositide-3-kinase regulatory subunit 4; PtdIns3P: phosphatidylinositol-3-phosphate; SMLM: single-molecule localization microscopy; ULK1: unc-51 like autophagy activating kinase 1; WIPI2: WD repeat domain, phosphoinositide interacting 2.
Subject(s)
Autophagy , Class III Phosphatidylinositol 3-Kinases , Autophagy/physiology , Autophagy-Related Proteins/metabolism , Beclin-1/metabolism , Autophagy-Related Protein-1 Homolog/metabolism , Class III Phosphatidylinositol 3-Kinases/metabolism , PhosphatidylinositolsABSTRACT
The perovskite solar cell (PSC), which has achieved efficiencies of more than 26%, is expected to be a promising technology that can alternate silicon-based solar cells. However, the performance of PSCs is still limited due to defects and ion migration that occur at the large number of grain boundaries present in perovskite thin films. In this study, the mixed ammonium ligands passivation strategy (MAPS) is demonstrated, which combines n-octylammonium iodide (OAI) and 1,3-diaminopropane (DAP) can effectively suppress the grain boundary defects and ion migration through grain boundaries by the synergistic effect of OAI and DAP, resulting in improved efficiency and stability of PSCs. It has also been revealed that MAPS not only enhances crystallinity and reduces grain boundaries but also improves charge transport while suppressing charge recombination. The MAPS-based opaque PSC shows the best power conversion efficiency (PCE) of 21.29% with improved open-circuit voltage (VOC ) and fill factor (FF), and retained 84% of its initial PCE after 1900 h at 65 °C in N2 atmosphere. Amazingly, the MAPS-based semi-transparent PSC (STP-PSC) retained 94% of their maximum power (21.00% at around 10% AVT) after 1000 h under 1 sun illumination and MAPS-based perovskite submodule (PSM) achieved a PCE of 19.59%, which is among the highest values reported recently.
ABSTRACT
The development of anti-solvent free, scalable, and printable perovskite film is crucial to realizing the low-cost roll-to-roll development of perovskite solar cells (PSCs). Herein, large-area perovskite film fabrication is explored using a spray-assisted sequential deposition technique. How propylene carbonate (PC) solvent additive affects the transformation of lead halide (PbI2 ) into perovskite at room temperature is investigated. The result shows that PC-modified perovskite films exhibit a uniform, pinhole-free morphology with oriented grains compared with pristine perovskite films. The PC-modified perovskite film also has a prolonged fluorescence lifetime that indicates lower carrier recombination. The champion PSC devices based on PC-modified perovskite film realize a power conversion efficiency (PCE) of 20.5% and 19.3% at an active area (A) of 0.09 cm2 and 1 cm2 , respectively. The fabricated PSCs are stable and demonstrate ≥85% PCE retention following 60 days of exposure to ambient conditions. Furthermore, perovskite solar modules (A ≈ 13 cm2 ) that yield a PCE of 15.8% are fabricated. These results are among the best reported for the state-of-art spray-coated PSCs. Spray deposition coupled with a PC additive is highly promising for economical and high-output preparation of PSCs.
ABSTRACT
BACKGROUND: As the nasal mucosa is the initial site of infection for COVID-19, intranasal vaccines are more favorable than conventional vaccines. In recent clinical studies, intranasal immunization has been shown to generate higher neutralizing antibodies; however, there is a lack of evidence on sterilizing immunity in the upper airway. Previously, we developed a recombinant measles virus encoding the spike protein of SARS-CoV-2 (rMeV-S), eliciting humoral and cellular immune responses against SARS-CoV-2. OBJECTIVES: In this study, we aim to provide an experiment on nasal vaccines focusing on a measles virus platform as well as injection routes. STUDY DESIGN: Recombinant measles viruses expressing rMeV-S were prepared, and 5 × 105 PFUs of rMeV-S were administered to Syrian golden hamsters via intramuscular or intranasal injection. Subsequently, the hamsters were challenged with inoculations of 1 × 105 PFUs of SARS-CoV-2 and euthanized 4 days post-infection. Neutralizing antibodies and RBD-specific IgG in the serum and RBD-specific IgA in the bronchoalveolar lavage fluid (BALF) were measured, and SARS-CoV-2 clearance capacity was determined via quantitative reverse-transcription PCR (qRT-PCR) analysis and viral titer measurement in the upper respiratory tract and lungs. Immunohistochemistry and histopathological examinations of lung samples from experimental hamsters were conducted. RESULTS: The intranasal immunization of rMeV-S elicits protective immune responses and alleviates virus-induced pathophysiology, such as body weight reduction and lung weight increase in hamsters. Furthermore, lung immunohistochemistry demonstrated that intranasal rMeV-S immunization induces effective SARS-CoV-2 clearance that correlates with viral RNA content, as determined by qRT-PCR, in the lung and nasal wash samples, SARS-CoV-2 viral titers in lung, nasal wash, BALF samples, serum RBD-specific IgG concentration, and RBD-specific IgA concentration in the BALF. CONCLUSION: An intranasal vaccine based on the measles virus platform is a promising strategy owing to the typical route of infection of the virus, the ease of administration of the vaccine, and the strong immune response it elicits.
Subject(s)
COVID-19 , Measles , Orthopoxvirus , Vaccines , Animals , Cricetinae , SARS-CoV-2 , Measles virus/genetics , COVID-19/prevention & control , Spike Glycoprotein, Coronavirus , Immunization , Nasal Mucosa , Antibodies, Neutralizing , Immunoglobulin A , Immunoglobulin G , Antibodies, Viral , Administration, IntranasalABSTRACT
Recombinant immunotoxins (RITs) are fusion proteins consisting of a targeting domain linked to a toxin, offering a highly specific therapeutic strategy for cancer treatment. In this study, we engineered and characterized RITs aimed at mesothelin, a cell surface glycoprotein overexpressed in various malignancies. Through an extensive screening of a large nanobody library, four mesothelin-specific nanobodies were selected and genetically fused to a truncated Pseudomonas exotoxin (PE24B). Various optimizations, including the incorporation of furin cleavage sites, maltose-binding protein tags, and tobacco etch virus protease cleavage sites, were implemented to improve protein expression, solubility, and purification. The RITs were successfully overexpressed in Escherichia coli, achieving high solubility and purity post-purification. In vitro cytotoxicity assays on gastric carcinoma cell lines NCI-N87 and AGS revealed that Meso(Nb2)-PE24B demonstrated the highest cytotoxic efficacy, warranting further characterization. This RIT also displayed selective binding to human and monkey mesothelins but not to mouse mesothelin. The competitive binding assays between different RIT constructs revealed significant alterations in IC50 values, emphasizing the importance of nanobody specificity. Finally, a modification in the endoplasmic reticulum retention signal at the C-terminus further augmented its cytotoxic activity. Our findings offer valuable insights into the design and optimization of RITs, showcasing the potential of Meso(Nb2)-PE24B as a promising therapeutic candidate for targeted cancer treatment.
Subject(s)
Antineoplastic Agents , Bacterial Toxins , Immunotoxins , Neoplasms , Single-Domain Antibodies , Animals , Mice , Humans , Exotoxins/genetics , Exotoxins/pharmacology , Exotoxins/chemistry , Immunotoxins/genetics , Immunotoxins/pharmacology , Immunotoxins/chemistry , Mesothelin , Single-Domain Antibodies/genetics , Single-Domain Antibodies/pharmacology , Bacterial Toxins/genetics , Bacterial Toxins/chemistry , Bacterial Toxins/metabolism , Catalytic Domain , Cell Line, Tumor , ADP Ribose Transferases/genetics , ADP Ribose Transferases/chemistry , ADP Ribose Transferases/metabolism , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/pharmacology , Recombinant Fusion Proteins/metabolism , Neoplasms/drug therapyABSTRACT
The E6 and E7 proteins of specific subtypes of human papillomavirus (HPV), including HPV 16 and 18, are highly associated with cervical cancer as they modulate cell cycle regulation. The aim of this study was to investigate the potential antitumor effects of a messenger RNA-HPV therapeutic vaccine (mHTV) containing nononcogenic E6 and E7 proteins. To achieve this, C57BL/6j mice were injected with the vaccine via both intramuscular and subcutaneous routes, and the resulting effects were evaluated. mHTV immunization markedly induced robust T cell-mediated immune responses and significantly suppressed tumor growth in both subcutaneous and orthotopic tumor-implanted mouse model, with a significant infiltration of immune cells into tumor tissues. Tumor retransplantation at day 62 postprimary vaccination completely halted progression in all mHTV-treated mice. Furthermore, tumor expansion was significantly reduced upon TC-1 transplantation 160 days after the last immunization. Immunization of rhesus monkeys with mHTV elicited promising immune responses. The immunogenicity of mHTV in nonhuman primates provides strong evidence for clinical application against HPV-related cancers in humans. All data suggest that mHTV can be used as both a therapeutic and prophylactic vaccine.