bluuebunny/crossref_metadata_2025 huggingface dataset:
An Adaptive Framework for Classification and Detection of Android Malware
Authors: Al Sharah Ashraf , Abu Alrub Yousef , Abu Owida Hamza , Abu Elsoud Esraa , Alshdaifat Nawaf , Khtatnaha Hamzah
Published: November 2024

The hardware and software of a computer are controlled by its operating system (OS), which performs essential tasks such as input and output processing, file and memory management, and the management of peripheral devices such as disk drives and printers. Application software refers to programs designed for specific purposes, these applications, often freely available and open source, contribute to the rising number of downloads. In the third quarter of 2022, combined downloads from the Apple App Store and Google Play Reached an estimated 35.3 billion. However, the prevalence of insecurity in these applications and technologies heightens the potential for cybercrimes. Protection against unauthorized intruders is crucial in identifying malicious applications. Machine learning (ML) serves as a promising avenue for detecting malware attacks, offering potential solutions to bolster cybersecurity measures. We propose a novel approach utilizing ML to enhance malware detection accuracy by segmenting datasets into distinct groups. Our research employs supervised ML techniques on the CICMaldroid2020 dataset, which includes comprehensive information such as intent actions, permissions, and sensitive APIs. The dataset was partitioned into four groups, each containing 150 features, and analyzed across four experiments to distinguish between attack and benign classes. Our proposed model demonstrated exceptional performance, with the random forest algorithm achieving an accuracy of 98.6% and a precision of 98.75%. These results highlight the effectiveness of our segmentation approach and its significant contribution to advancing malware detection in Android applications, offering a promising direction for future cybersecurity solutions.
Innovation and Optimization of Corporate Governance Models through Mobile Interactive Technology
Authors: Jia Meixia , Lyu Xiandong
Published: November 2024

In the context of rapid globalization and the swift advancement of information technology (IT), traditional corporate governance models are increasingly challenged by complex business environments and the growing demand for diversified collaboration. These models often exhibit rigidity and inefficiency. The introduction of mobile interactive technology offers new possibilities for the innovation and optimization of corporate governance models. Through mobile interactive networks, resource allocation and collaboration between departments can be conducted flexibly and efficiently. Existing research predominantly focuses on traditional management information systems and centralized resource allocation methods, which frequently overlook the diversity and dynamic nature of internal resource demands within enterprises, making them inadequate for addressing the needs of modern corporations. This study proposes a collaborative interaction mechanism for corporate governance based on matching theory, designing a system model that includes collaborative task demanders and executors, and the social utility of mobile interactive networks. The findings demonstrate that this mechanism enables efficient resource allocation and stable collaborative interactions in complex environments, providing both theoretical support and practical guidance for the innovation of corporate governance models.
The Impact of Mobile Payments on the Financial Management Efficiency of Small and Medium-Sized Enterprises
Authors: Li Qing
Published: November 2024

In recent years, mobile payment technology has rapidly developed and gradually become an integral part of daily economic activities, particularly within small and medium-sized enterprises (SMEs). The widespread adoption of mobile payments has significantly transformed financial management and operational models in SMEs. As vital drivers of economic growth, the financial management efficiency of SMEs is crucial to their survival and development. The extensive application of mobile payments not only facilitates daily transactions but also provides additional data and information support, potentially enhancing financial management efficiency. However, although numerous studies focus on the impact of mobile payments on business operations, there is a gap in the specific examination of their effect on the financial management efficiency of SMEs. Existing research predominantly addresses the impact of mobile payments on the overall market, lacking detailed analysis targeting SMEs. Furthermore, research methodologies often overlook the heterogeneity of user behavior on mobile payment platforms, failing to comprehensively reflect the differential impact on various types of enterprises. This study explores how user utility, user scale, platform profit, and equilibrium price affect the financial management efficiency of SMEs by constructing a theoretical model. It consists of two parts: initially defining financial management efficiency in SMEs and analyzing its influencing factors to update their financial management models. Subsequently, it examines the impact of mobile payment platforms on these factors through empirical analysis and model derivation. The findings demonstrate that mobile payment technology significantly enhances the financial management efficiency of SMEs, offering useful insights for business management and policy-making.
Les principes pédagogiques à l’œuvre dans les épistémologies pratiques de deux professeurs des écoles stagiaires
Authors: Musard Mathilde , Le Paven Maël , Le Borgne Philippe
Published: November 2024

Cet article s’intéresse à l’épistémologie pratique de deux professeurs des écoles stagiaires lors d’un dispositif d’analyse didactique des pratiques. Ceux-ci sont invités par les chercheurs à décrire et interpréter des extraits de leurs situations d’enseignement et d’apprentissage en mathématiques et en éducation physique et sportive lors d’entretiens d’auto-confrontation (EAC). Les résultats obtenus en croisant des données d’observation et d’entretien montrent que les professeurs sont attachés prioritairement à des principes pédagogiques pour définir et dévoluer les situations, réguler les apprentissages et institutionnaliser les savoirs. Les professeurs ont tendance à délaisser la question des contenus d’apprentissage. Nous discutons des modalités d’accès à ces principes pédagogiques, puis nous pointons quelques pistes sur la sensibilisation des professeurs débutants aux préoccupations didactiques et sur les caractéristiques des dispositifs de formation. Nous discutons la dimension générique de ces principes avant de décliner les conceptions des deux professeurs des écoles stagiaires (PES), contribuant ainsi à une ébauche de leurs épistémologies pratiques.
Environmentally Sustainable Production of Bacterial Nanocellulose in Waste-Based Cell Culture Media and Applications
Authors: Kobayashi Takaomi
Published: November 2024

Bacterial cellulose has attracted great attention due to the demand for eco–friendly materials and sustainable products. It possesses properties superior to those of plant cellulose and has potential uses in various applications. The wider application of bacterial cellulose depends on the practical considerations such as the scale–up capability and production costs. The high cost of bacterial cellulose production is the main drawback that hinders industrial implementation. The cost–competitiveness can be improved, and bacterial cellulose production can be maximized by the utilization of agricultural and industrial waste and by–products as bacterial cell culture media. This chapter provides an overview of cost–effective culture media for bacterial cellulose production by using agricultural waste as a primary nutrient source. The applications related to nanocellulose, and the future challenges are also described.
Assessing the Photocatalytic Performance of Carbon-Based Semiconductors in the Degradation of Pharmaceutical Wastes
Authors: Ismail Nursarah Sofea , Rithwan Ahmad Fadhil Bin , Thongmee Sirikanjana , Yusoff Siti Fairus Mohd , Adnan Rohana , Mohd Kaus Noor Haida
Published: November 2024

This review highlights recent research endeavors in the field, emphasizing the critical role of carbon-based photocatalysts for their superior performance under visible light irradiation and their capacity to efficiently photodegrade pharmaceutical contaminants. The integration of carbon materials with semiconductors has demonstrated remarkable synergistic effects, leading to enhanced photocatalytic efficiency. The exploration of carbon-based semiconductor materials represents a promising avenue for addressing the persistent issue of pharmaceutical waste in the environment, offering a greener and more efficient solution for water treatment.
Cultivating Computational Thinking Through STEM and AI in Early Education
Authors: Bataineh Ruba Fahmi , Bataineh Rula Fahmi
Published: November 2024

This chapter addresses the interconnectedness of computational thinking with STEM and AI and their potential to revamp early education. It also addresses practical approaches to teaching computational thinking through STEM and AI. The chapter also discusses the major challenges facing integrating computational thinking into academia, underscores the urgent need to redesign early education to equip learners with a future-oriented mindset, and emphasizes the need for fostering lifelong learning, creativity, and engagement while navigating the complexities of interactive technologies.
Integrating AI Into Early Childhood Education
Authors: Metaxoudis Eleftherios T.
Published: November 2024

This chapter offers a comprehensive exploration of integrating artificial intelligence (AI) into early childhood education, bridging theoretical foundations with practical applications. It begins by providing key educational theories, to establish a developmental framework for AI integration. The chapter then reviews current AI applications in early learning environments, analyzing their benefits and limitations through recent research and case studies. Practical examples demonstrate AI's role in personalized learning, language development, and social-emotional learning. Ethical considerations are also thoroughly discussed. The chapter concludes by discussing and providing actionable recommendations for educators, policymakers, and researchers to ensure ethical and effective AI integration in early childhood education. This chapter serves as an authoritative resource for stakeholders aiming to enhance early childhood education through AI.
Integration of Digital Competences in Preschool Education in the Czech Republic
Authors: Straková Nikola , Marinič Peter , Válek Jan
Published: November 2024

Due to the changing needs of the labour market, which are caused by the rapid development of digital technologies, the development of computational thinking and digital competences at the youngest children is already started in pre-school education. Teaching with digital technologies increases children's motivation to learn, develops creativity, independent thinking, the ability to solve problems, and has many other benefits, which children build on and develop further in primary schools and further education. The Czech Republic, with the support of the European Union, is actively implementing the digitization of education through curriculum reform and investments related to the implementation of the revised curriculum and equipping schools with advanced digital learning tools. The goal of digitization in education from pre-school education is to increase the number of IT professionals, improve advanced digital skills among the population and ensure long-term employment.
UTX-mediated conversion of CD8 progenitors to mediators in type 1 diabetes
Authors: Chen Ho-Chung , Zhang Shirley , Seyedsadr Maryam , McCarthy Ethan , Cheng Mandy , Ortega Jessica , Kohn Lisa , Lechner Melissa , Su Maureen
Published: November 2024

Type 1 Diabetes (T1D) is a CD8 T cell-mediated autoimmune disease characterized by the persistent destruction of pancreatic beta cells. Within the pancreatic lymph nodes (pLNs) where CD8 T cells are primed by antigen-presenting cells, a subset of CD8 T cell Tcf7+ progenitors with self-renewing stem-like properties was recently identified. These progenitors undergo differentiation and ultimately give rise to CD8 mediators, which enter the pancreas to cause beta cell destruction. Recent work has demonstrated that CD8 T cell differentiation from progenitors is subject to epigenetic regulation. Here, we identify the epigenetic regulator UTX (ubiquitously transcribed tetratricopeptide repeat, X chromosome) in promoting T1D by regulating the conversion of CD8 progenitors into mediator in pLNs. Non-obese diabetes (NOD) mice with T cell specific deletion of UTX displayed complete protection from T1D, which was associated with the accumulation of progenitor CD8 T cells and a reduced CD8 mediators in pLNs. Furthermore, knockout of UTX in human peripheral blood CD8 T cells demonstrated impaired differentiation into effector memory and terminally differentiated CD8 T cells. Treatment of NOD mice with GSK-J4, a histone demethylase inhibitor of JMJD3/UTX, delayed the spontaneous T1D development. Collectively, our findings show that UTX enhances the generation of CD8 mediators in pLNs, thereby facilitating T1D progression.
Investigating the contributions of microbial peptides in the induction of experimental autoimmune encephalomyelitis
Authors: Schwegman Lance , Valencia Mary Jo , Forsthuber Thomas
Published: November 2024

Multiple sclerosis (MS) is an incurable neuroinflammatory autoimmune disease in which a patient’s immune system targets the myelin sheath of neurons. This impairs neuronal signaling to the periphery, often manifesting as severe pain, loss of vision, and partial paralysis. Several environmental and genetic risk factors, such as the DRB1*1501 allele, have been identified, but the exact etiology of this disease remains unclear. It is thought that MS is primarily mediated by myelin-reactive CD4+ T cells, but it remains unresolved how these cells are initially activated. Microbial peptides with amino acid sequences similar to myelin-derived peptides have previously been reported, however no pathogen has been firmly implicated in causing neuroinflammation via molecular mimicry. Previous work in our lab using the mouse model of MS, experimental autoimmune encephalomyelitis (EAE), suggests that the concurrent presence of low frequencies of CD4+ T cells that recognize distinct myelin-derived antigens can act in concert to trigger neuroinflammation. We therefore hypothesize that the induction of T cell immunity against multiple microbial antigens with sequence similarity to self-peptides may generate T cell populations that contribute to the induction of EAE in humanized mice. With the data from this study, we hope to clarify the effects of microbial encounters and provide new insights into the pathogenesis of MS.
Machine learning reveals UHRF-X as a biomarker for systemic lupus erythematosus and an antagonistic protein against UHRF1 E3 ligase
Authors: Chuang Huai-Chia , Tan Tse-Hua
Published: November 2024

Inflammatory T cells contribute to the pathogenesis of the autoimmune diseases such as systemic lupus erythematosus (SLE). Analysis of the T-cell transcriptomics data of two independent SLE patient cohorts by three machine learning models revealed UHRF-X as a novel SLE biomarker. T-cell-specific UHRF-X transgenic mice manifested the induction of IL-17A and autoimmune inflammation. Mechanistically, UHRF-X prevented UHRF1-induced Lys48-linked ubiquitination and degradation of MAP4K3 (GLK), which is a kinase known to induce IL-17A. Consistently, IL-17A induction and autoimmune phenotypes of UHRF-X transgenic mice were obliterated by MAP4K3 knockout. Furthermore, UHRF-X protein was indeed overexpressed in peripheral blood T cells of SLE patients. Collectively, UHRF-X overexpression in T cells inhibits the E3 ligase function of UHRF1 and induces IL-17A-mediated autoimmune responses. This report unveils a novel gene that can block UHRF1 function, leading to autoimmune disease.
Heterogeneity of B cells in the immunoregulatory niche at the cribriform plate during EAE
Authors: Kovacs Kristof , Laaker Collin , Fabry Zsuzsanna
Published: November 2024

B cells may play regulatory roles in multiple sclerosis (MS) patients and experimental autoimmune encephalomyelitis (EAE) mice. Recently, B cell development was revealed to occur in the meninges, inducing B cell tolerance toward central nervous system (CNS) antigens. Our group recently showed that neuroinflammation in EAE induces an immune regulatory niche at the cribriform plate (CP) meningeal lymphatics. Here we characterize the phenotype and regulatory function of B cells at the CP in a mouse model of CNS autoimmunity.

Using immunohistochemistry and cytofluorimetry, we found that the number of B cells increases during EAE at the CP niche. The majority of B cells form doublets, triplets and larger aggregates with dendritic cells (DC) at the CP. Single-cell RNA-sequencing of DC-interactor B cells showed that B cells undergo maturation at the CP. We identified upregulated genes indicative of different maturation stages characteristic of common lymphoid progenitors (Ly6D), early pro-B cells (Vpreb3, FOXO1), immature (BAFF-R) and mature (CD20) B cells. MetaScape analysis confirmed that B cells differentiate and become activated at the CP. We hypothesize that interaction with B cells induces a tolerogenic state in DCs, promoting the B cell-driven regulation of CNS autoimmunity.

In conclusion, we propose that B cells contribute to the formation of the CP immunoregulatory niche. This holds promise in elucidating potential implications of B cell immunomodulatory therapies in MS.
Autoreactive IgA antibodies are high prevalent and associated with clinical manifestations in systemic lupus erythematosus, systemic scleroderma and idiopathic inflammatory myositis
Authors: Li Quanzhen , Wei Junxia , Luo Hui , Zhu Honglin , Zhu Chengsong
Published: November 2024

Systemic lupus erythematosus (SLE), systemic scleroderma (SSc) and idiopathic inflammatory myositis (IIM) are autoimmune diseases characterized by a broad range of autoreactive antibodies (AutoAbs). In this study, we measured the serum IgA AutoAbs against 120 self-antigens in a cohort of 128 SLE, 73 SSc, 75 IIM and 140 healthy controls (HC), using OmicsArray™ antigen microarrays. We found IgA AutoAbs are highly prevalent in SLE and SSc compared with IIM and NC. 58.4% SLE and 41.7% SSc have 5 or more IgA AutoAbs compared with 16% in IIM and 14.1% in NC. 9 IgA AutoAbs most significantly elevated in SLE were anti-DNA antigens including dsDNA (54.7%), ssDNA (51.1%), and chromatin (46%). 32 IgA AutoAbs were significantly increased in SSC with highest reactivity to Scl-70 (55.6%). IIM showed lower IgA prevalence in general with highest in anti- Jo-1 (20%), anti-muscarinic receptor (8%) and anti-B2-microglubulin (6.7%). Significant correlation was observed between IgA and IgG isotypes on most AutoAbs. 3 IgA autoAbs (anit-gDNA, anti-C1q and anti-dsDNA) positively correlated with SLEDAI Score, but only anti-C1q was significantly elevated in lupus nephritis. 14 anti-nuclear IgA AutoAbs negatively correlated with serum complement C3 and/or C4 levels. Our study indicated that autoreactive IgA against nuclear components, in parallel with IgG, are highly prevalent in SLE and SSC patients. IgA AutoAbs could be used as biomarkers for diagnosis and prognosis of systemic autoimmune diseases.
The Effect of Weight Cycling on Myelopoiesis
Authors: Martinez-Navarrete Wenceslao , Kalev-Altman Rotem , Guha-Ray Aleepta , Wiseman Destini , Weinstock Ada
Published: November 2024

Inflammation is a hallmark of obesity and is associated with many complications, with myeloid cells being major culprits. Weight loss improves obesity-related comorbidities, however, subsequent weight regain, known as weight cycling (WC), has been linked to worsening of obesity-related diseases. Therefore, it is critical to understand how WC impacts inflammation. We hypothesize that mice undergoing WC will have greater myeloid cell production with heightened inflammatory capacity. To test this hypothesis, we characterized immune cell populations in bone marrow (BM) and blood using multi-parametric flow cytometry in mice undergoing acute and long-term WC compared to non-WC mice. Our results indicate WC increases the frequency of inflammatory Ly6Chi monocytes and lowers pro-resolving Ly6Clo monocytes in the BM. Additionally, WC increases circulating monocytes. The increase in monocytes are coupled with expansion of their bone marrow progenitors, suggesting that WC heightens myelopoiesis. We will further examine inflammatory capacity by stimulating bone marrow derived macrophages with lipopolysaccharide and measuring cytokine secretion. Furthermore, hematopoietic progenitor differentiation will be tested through colony forming assays. Combined RNA/ATAC-seq will be used to understand the transcriptomic and epigenetic changes occurring in progenitors with WC. This study will provide novel insight into how WC worsens disease, and the basic mechanisms regulating myelopoiesis.
Excessive expression of TIM-3 on myeloid cells leads to inflammation in the lungs
Authors: Kim Ki Sun , Lee Chanju , Kim Hyung-Seok , Gu Su Jeong , Yoon Hee Jung , Won Su Bin , Kane Lawrence P , Park Eun Jung
Published: November 2024

T cell immunoglobulin and mucin-containing molecule 3 (TIM-3) has increasingly shown to exhibit unique characteristics and functions that differ according to the specific cell type and cell status. However, at present, little is known about the cell-specific properties of TIM-3 in pathophysiological conditions. Here, we show that TIM-3 on myeloid cells exerts essential roles in modulating inflammation and abnormality of lungs. We found that myeloid cell-specific TIM-3 knock-in (FSF-TIM3/LysM-Cre+) mice have a characteristic appearance of low body weight and short lifespans compared to WT mice. Interestingly, whole-body PET/CT analysis revealed that FSF-TIM3/LysM-Cre+ mice showed considerably high [18F]FDG uptake in the areas of lungs. Given that FSF-TIM3/LysM-Cre+ mice displayed abnormal features in lung areas, we next carefully examined the lungs. We observed that the lungs of FSF-TIM3/LysM-Cre+ mice were enlarged and histopathological examination showed features of disease-associated pathology compared to WT mice. Similar features were observed in a different myeloid cell-specific TIM-3 knock-in mouse (FSF-TIM3/Cx3cr1-Cre+). In addition, inflammation-related cytokines and the number of myeloid innate immune cells increased in the lungs of FSF-TIM3/LysM-Cre+ mice. Collectively, these findings suggest that TIM-3 is an important regulator of myeloid cells in the lungs and maybe a relevant therapeutic strategy against inflammation-associated lung disease.
Plasmacytoid dendritic cells are central regulators of the humoral ChAdOx1 nCoV-19 vaccine response in a human tonsil organoid model
Authors: Provine Nicholas , Pudjohartono Maria , Powell Kate , Ferry Helen , Barnes Ellie , Klenerman Paul
Published: November 2024

Studying human vaccine responses is limited by the difficulty of access to secondary lymphoid tissues. To overcome this, we have optimized a human tonsil organoid model to investigate the processes that regulate the humoral responses to the ChAdOx1 nCoV-19 vaccine.

Stimulation of tonsil cultures with ChAdOx1 nCoV-19 induced broad innate immune activation and cytokine secretion within 24 hours. This induced (global and antigen-specific) B cell activation and spike-specific antibody secretion over the following 14-day culture period. Stimulation with an irrelevant ChAdOx1 GFP vector did not induce spike-specific antibody secretion.

Plasmacytoid dendritic cells (pDCs) showed the highest rate of transduction and activation. Critically, pDC-depletion decreased both innate and humoral responses. IFN-⍺ blockade phenocopied pDC depletion while supplementing IFN-⍺ to pDC-depleted cultures rescued humoral responses. IL-6 was also critical for optimal humoral responses, and its production was modulated in an IFN-⍺-dependent manner even in the absence of pDCs, suggesting cross-talk of multiple key signaling pathways.

Overall, this model reveals a critical role for pDCs in inducing humoral responses to ChAdOx1 nCoV-19, primarily through IFN-⍺. IFN-⍺ appears to act directly and indirectly through IFN-induced IL-6 secretion to augment the humoral response. This model has the potential for studying other vaccine platforms.
Comparing the antibody repertoire of XBB1.5 booster immunization with mRNA or protein-based platforms
Authors: Lobo Igor , Cohn Hallie , Bozarth Bailey , Fantin Raianna , Rao Vishal , Bushfield Kaitlyn , Nardulli Jessica , Srivastava Komal , Clark Jordan , Study Group PVI , Krammer Florian , Simon Viviana , Coelho Camila
Published: November 2024

The SARS-CoV-2 spike (S) protein is an essential target for neutralizing antibodies, accumulating mutations, and generating variants of concern (VOC). The Omicron variant is a VOC that continues to evolve into antibody-resistant subvariants, such as XBB1.5. This led to the approval of an updated mRNA vaccine and a protein-based vaccine to target the XBB1.5 strain. However, whether these vaccines can elicit functional monoclonal antibodies specifically against this strain and how the different vaccine platforms (mRNA vs protein) shape the antibody repertoire is unknown. Here, we analyzed the antibody repertoire of five humans who either received the updated mRNA or the protein-based vaccine. We used the Immcantation framework to analyze the VDJ sequences. We assigned V, D, and J genes using IgBlast according to the Change-O pipeline. Clonal groups were defined using the SCOPer package v.1.3.0, and then we calculated somatic hypermutation (SHM) for the BCR sequences. Firstly, we reconstructed germline V and J sequences. The total number of mutations was found for the V gene using SHazaM v.1.2.0, and clonal lineage trees were built using the dowser package v.2.0. We are currently expressing human monoclonal antibodies from the participants and will assess binding, neutralization, and affinity. Our results will compare the antibody activity elicited by vaccination between protein-based (Novavax) and mRNA (Pfizer/Moderna) in humans receiving the XBB1.5 booster.
The breadth and neutralizing activity of SARS-CoV-2 antibodies among children 6 months to 4 years of age following vaccination with mRNA-1273 or BNT162b2
Authors: Williams Caitlin , Dalapat Trisha , Giorgi Elena , Hurst Jillian , Herbek Savannah , Chen Jui-Lin , Kosman Christina , Rotta Alexandre , Turner Nicholas , Pulido Natalie , Aquino Jhoanna , Pfeiffer Trevor , Rodriguez Javier , Permar Sallie , Fouda Genevieve , Kelly Matthew
Published: November 2024

Body: The mRNA-based COVID-19 vaccines approved for use in children less than 5 years of age have differences that could affect vaccine immunogenicity and effectiveness. We directly compared the adaptive immune responses to mRNA-1273 or BNT162b2 in children under the age of 5.

Methods: We enrolled children 6 months to 4 years of age who completed primary series vaccination with monovalent mRNA-1273 or BNT162b2 vaccines between July and December 2022. Serum was collected from children one month after completion of the primary vaccine series. Children receiving either vaccine series developed robust and broad serum SARS-CoV-2-specific binding and neutralizing antibodies, including to Omicron subvariants. Children with a prior history of SARS-CoV-2 infection or who had detectable serum SARS-CoV-2 nucleocapsid antibodies at enrollment developed significantly higher antibody binding responses and neutralization. No differences in vaccine-induced antibody responses were observed between children 6-23 months of age and 24-59 months of age.

Conclusions: Monovalent mRNA-1273 and BNT162b2 elicit similar antibody responses one month after vaccination in young children irrespective of age. Prior infection significantly enhanced the strength of antibody responses to Omicron subvariants. These findings demonstrate that both mRNA-based COVID-19 vaccines induce immune correlates of protection from severe COVID-19 among young children.
Optimization of an anti-fentanyl monoclonal antibody to improve stability
Authors: Hoppe Brooke , Hicks Dustin , Baehr Carly , Kang Jason , Pravetoni Marco , Rodarte Justas , Pancera Marie
Published: November 2024

Fentanyl and other synthetic opioids continue to be the primary contributors to overdose deaths despite widespread availability of FDA-approved reversal agents, highlighting the need for novel complementary therapeutics to address this epidemic. Monoclonal antibodies (mAbs) targeting fentanyl and related analogues have been shown to be promising candidates to help treat opioid overdose in preclinical models. Maintaining formulation stability is critical to the clinical utility of these immunotherapeutics. Previously, the structure of a lead anti-fentanyl mAb in complex with fentanyl was solved through X-ray crystallography, identifying a residue near the binding site that was liable to oxidation. Using a structure-guided approach, the identified amino acid residue was mutated to residues that were expected to have reduced oxidative liability. Stability of the resulting mAbs was quantified using differential scanning fluorimetry, and affinity of the mAbs for fentanyl was assessed via biolayer interferometry (BLI) and competitive enzyme-linked immunosorbent assays (ELISA). Replacing the oxidation-liable residue improved antibody stability as evidenced by an increase in Fab melting temperature. One specific mutation resulted in a 40-fold increase in affinity measured against a fentanyl hapten by BLI, and a 4-fold increase in affinity for free fentanyl by competitive ELISA. Future studies will seek to validate the improved mAb affinity in vivo relative to the parent mAb.
Elucidating the role of macrophage galactose-type lectin 1 (MGL-1) in the pathogenesis and resolution of SARS-CoV2
Authors: Perez Jaid , Puebla-Clark Lucinda , Huante Matt , Orozco Maria , Naqvi Kubra , Martinez-Martinez Yazmin , Menachery Vineet , Rajsbaum Ricardo , Stephens Robin , Endsley Mark
Published: November 2024

The C-type lectin receptor (CLR) family plays an important role in mediating the innate immune responses to some viruses. The Macrophage galactose-type lectin (MGL), or CLEC10, is a CLR that binds SARS-CoV-2 Spike. This binding event is dispensable for virus entry but has been implicated in an increased inflammatory response. Downstream immune responses relevant to host protection against SARS-CoV-2 are not well described. We investigated the role of MGL in disease pathogenesis by utilizing MGL-1 deficient mice (KO) and wild type (WT) control mice. Male and female mice were intranasally infected with the mouse-adapted SARS-CoV-2 strain CMA3p20 for 3 and 7-days. Lung tissues were assessed for viral load, pathology, and changes in leukocyte populations. Viral load was similar in mouse strains at 3 days post-infection (dpi), while virus persisted in MGL-1-/-, but not WT, at 7 days. This observed difference was more pronounced in female mice. Histology analysis indicated no difference in lung pathology between wild type and MGL deficient mice at 3 dpi. Immunohistochemistry and flow cytometric analysis of lung tissue for CD3 also indicated similar numbers and distribution of T cell populations. By using full spectrum flow cytometry, preliminary clustering analysis shows an increase in activated B cells and plasma cells in MGL-1-/- compared to WT mice. We are exploring the implications of altered B cell populations by investigating changes in antibody titers and isotypes.
B and T cell repertoire and gene expression differences in measles infection and vaccination revealed by longitudinal single-cell RNA/VDJ sequencing
Authors: Chan Kwan Pui , Rubens Jessica , Bartlett Maggie , Brockhurst Jacqueline , Ghimire Shristi , Griffin Diane
Published: November 2024

While wild-type measles virus (WT MeV) was demonstrated to induce immune amnesia, it paradoxically still elicited high titers of measles-specific antibodies longitudinally; and much higher than the live-attenuated measles vaccine (LAMV). In order to further understand the longitudinal B and T repertoires, clonotype evolution and gene expression difference in WT MeV and LAMV, we conducted longitudinal single-cell RNA/VDJ sequencing of peripheral blood mononuclear cells (PBMC) from WT MeV and LAMV-infected rhesus macaques (RMs) (n = 3 in each WT MeV and LAMV groups, with 4 timepoints at D0, D11, D42/56, D168/176).

Firstly, B cell subtypes (e.g. naïve, memory, activated and antibody-secreting B cells) were successfully annotated in PBMC based on key gene markers and immunogenetic information. Furthermore, preliminary analysis of 1 WT MeV and 1 LAMV revealed interesting differences in the B cell repertoire, including a more diverse usage of IGHV genes, greater isotype switching from IgD/M to IgGs, higher level of somatic hypermutation and later B cell clonal expansion peak in LAMV than WT-MeV infected RMs longitudinally. Interestingly, there was greater T cell clonotype expansion in the WT MeV than LAMV RMs.

Our study highlights the differential humoral and adaptive immune responses between LAMV and WT MeV infections, uncovering important immunogenetic understanding in natural infection and vaccination.
Electronic health record data shows immunosuppressive conditions are associated with poor outcomes in patients hospitalized for COVID-19
Authors: Guggilla Vijeeth , Pacheco Jennifer , Carvalho Alexandre , Pawlowski Anna , Whitmer Grant , Achenbach Chad , Walunas Theresa
Published: November 2024

We identified 10,713 adults hospitalized for COVID-19 from 03/2020 to 05/2022 in electronic health record data from Northwestern Medicine. Based on extracted diagnosis codes, we identified 214 of these adults with primary immunodeficiency (PI), 669 with history of solid organ transplant, and 183 with HIV. Assessed COVID-19 outcomes included mortality, a seven-point ordinal severity scale (1 = Death) based on maximum oxygen requirements during care, and hospitalization length. Patients with PI had a significantly higher mean hospitalization length of 11.8 days and a significantly lower mean maximum severity of 3.3 compared to 7.8 days and 3.9 in immunocompetent patients when controlling for obesity and T2DM. These effects were not significant when controlling for age. Patients with transplant had a significantly higher mean hospitalization length of 15.8 days when controlling for obesity, T2DM, and age, and a significantly lower mean maximum severity of 3.2 when controlling for obesity. Significantly higher mortalities were observed in PI and transplant (RR = 1.5, 95% CI 1.2 to 2.0; RR = 1.5, 95% CI 1.3 to 1.7), but these effects were not significant when controlling for age. There were no significant differences in any outcomes for patients with HIV. Our results suggest patients with PI or transplant hospitalized for COVID-19 may have worse clinical outcomes compared to immunocompetent patients. Meanwhile, patients with HIV had similar outcomes to immunocompetent patients.
Increased interleukin-6 correlates with LAMP1 expression in NK cells and hospital stay of patients with SARS-COV-2 pneumonia.
Authors: OSUNA ESPINOZA KENIA , Mejía Torres Manuel , Camacho Ortiz Adrian , Perez Alba Eduardo , Salinas Carmona Mario , Rosas Taraco Adrian
Published: November 2024

During viral infections, NK cells play an important role in virus clearance. In the course of COVID-19 pandemic, the immunologic response to viral infections gains a main interest. Despite the extensive evidence, the functional status of cytotoxic cells has remained a controversial area of interest. Here, we provide a detailed comparative analysis of immune population counts and activation markers expression (LAMP1, IFNγ, KIR2DL1 and granzyme B) in NK cells from blood by flow cytometry, and serum cytokine concentration by LUMINEX from patients with severe COVID-19 and healthy participants. A significant decrease in total lymphocyte was observed in COVID-19 patients, mainly by T-CD4+, T-CD8+ and NK cells. The expression of the activation marker LAMP1 on NK cells was significantly higher in COVID-19 patients compared with the control group (P<0.05), and its expression was related to their clinical outcome. Also, IL-6 concentration was significantly higher in COVID-19 patients, correlated negatively with LAMP1 expression (r2=-0.693, P<0.05) and positively with hospital stay (r2=0.752, P<0.01). In conclusion, we show evidence of increased expression of the activation marker LAMP1 in NK cells, and high levels of IL-6 in COVID-19 patients, as well as, their correlation with the clinical outcome and hospital stay of the patients.
High sensitive detection and validation of neuronal specific autoantibodies in post COVID-19 patients with neurocognitive disorders
Authors: Li Quanzhen , Wei Junxia , Niu Luming , Yang Shuwei , Liang Chen
Published: November 2024

COVID-19 has affected tens of million patients worldwide and a significant portion of COVID-19 patients experience post-COVID syndrome characterized by a variety of persistent, debilitating symptoms including profound neurological symptoms. Studies have demonstrated the association between autoantibodies and the severity of COVID-19. but the role of autoantibodies in post-COVID-19 patients with neurological disorders has not yet well established. We screened the levels of anti-neuronal antibodies in 120 serum and CSF samples from 60 post-COVID-19 patients using an OmicsArray™ antigen panel containing 83 neuronal specific antigens (NSA). We identified 7 anti-NSA antibodies which were significantly elevated in serum of at least 1 post-COVID-19 patient with neurocognitive symptoms, including anti-CASPR2, anti-GAD65, anti-RCVRN, anti-mGluR2, anti-MuSK, anti-CSPG4 and anti-ELAVL2 (HuB). 3 anti-NSA antibodies (anti-MOG, anti-GAD65 and anti-mGluR2) were also detected positive in CSF samples of neurocognitive patients. Pre-incubation of the patient serum samples with the purified antigens inhibited the binding of the antibody with the specific NSA on array, validated the specificity of antigen array results. We further confirmed that anti-MuSK antibody in patient serum was targeting the extracellular domain of the MuSK protein using peptide array. Our results suggest anti-NSA antibodies may play a role in post-COVID neurological syndrom and warrant further study.
Epigenetic drug screen of natural killer cells identified compounds controlling immune training and tolerance
Authors: Geanes Eric , Fraley Elizabeth , Pierce Stephen , McLennan Rebecca , Bradley Todd
Published: November 2024

Natural killer (NK) cells are cytotoxic innate lymphocytes that provide defense against pathogens and malignancy. New evidence identified that NK cells are capable of memory-like immune responses in certain settings. This innate immune memory is thought to be imprinted through epigenetic modifications. However, precise epigenic pathways or strategies to modulate long-term NK cell activity are not well defined. In this study, we performed a screen of an epigenetic library containing 160 drug compounds with well-characterized epigenetic regulatory mechanisms, to identify drugs and pathways that could train or tolerize NK cell activation. One compound that we identified was the H3K27 methyltransferase Ezh2, which regulated NK cell lineage commitment from bone marrow hematopoietic stem cells and altered the phenotype of differentiated NK cells to increased cytotoxicity. This outcome provides an option to train or induce stronger cytotoxic effects on NK cells. In contrast, another compound we identified was the histone deacetylase inhibitor, Givinostat, which did not alter NK cell commitment from bone marrow stem cells, but significantly decreased NK cell function in the periphery resulting in a more tolerant innate immune state to secondary NK cell stimulation. Identification and exploration of these epigenome altering drugs may offer further insight into downstream pathways of NK cell function or provide novel therapies for tolerizing or priming innate immune responses.
Regulation and activity of Runx2 in CD8+ T cells responding to infection promotes long-term survival
Authors: Michaels-Foster Mary , Berg Leslie
Published: November 2024

Constitutive expression of Runx family proteins in CD8 T cells and maintenance of accessibility surrounding Runx motifs in regulatory regions of lineage-specific loci suggest that they may play a role in establishing and maintaining the epigenetic state of CD8 T cells. Additionally, Runx proteins possess a highly homologous DNA binding domain, leading not only to redundancies, but also potential competition between Runx proteins for the same genomic binding sites. While Runx1 and Runx3 have well known roles in the development of CD8 T cells, the function of Runx2 is not well established. To further explore the function of Runx2, our lab utilizes a model of retroviral overexpression and CRISPR knockout of Runx proteins in OT-1 CD8 T cells. Adoptive transfer of Runx2 overexpressing or knockout cells followed by ovalbumin-expressing Listeria monocytogenes (LmOva) infection suggests that Runx2 activity is important for the expression and maintenance of genes needed for long-term survival and function, such as Tcf7 and Il7ra. Data analyzed from the literature and chromatin analyses from our lab suggest that Runx proteins may compete for binding sites and drive opposing transcriptional and epigenetic profiles in CD8 T cells after infection. Preliminary data also give insight into the mechanisms controlling Runx2 expression, suggesting that the signals received during priming may affect the balance of Runx protein activity.
Mechanisms by which Lactoferrin-derived chimeric peptide (LFch) enhances TGFβ1-induced Treg Differentiation
Authors: Kim Pyeung-Hyeun , Jang Young-Saeng , Park SunHee
Published: November 2024

We recently reported that LF substantially promoted Foxp3 expression by activated CD4+T cells and this activity was further enhanced by TGF-β1. We also found that the lactoferrin chimera (LFch, synthetic LF-derived chimeric peptides) dramatically enhanced TGFβ1-induced Foxp3 expression, although LFch itself did not stimulate Foxp3 expression. In the present study, we explored the mechanism by which LFch boosts TGFβ1-induced Treg differentiation. We applied biotinylated LFch (LFch-biotin) to elucidate this issue. Through the receptor ELISA and co-immunoprecipitation experiments, we observed that LFch-biotin directly binds to TβRIII and PI3K. Further, it also binds and diminishes phosphorylated-PI3K which is associated with CD28. As expected, LFch-biotin boosted TGFβ1-induced Foxp3 expression. The same was true when PI3K inhibitor was treated. Taken together, these results suggest that LFch upregulates TGFβ1-induced Treg differentiation by diminishing CD28 signal intensity through the complex of LFch/TβRIII/PI3K/CD28.
Targeting the SIRT1 pathway to modulate inflammation in a murine model of Kawasaki Disease vasculitis
Authors: Atici Asli , Jena Prasant , Gomez Angela , T. Carvalho Thacyana , Crother Timothy , Noval Rivas Magali
Published: November 2024

Kawasaki Disease (KD) is an acute febrile systemic vasculitis and the leading cause of acquired heart disease among children. Overactivation of the NLRP3 inflammasome and IL-1b production are critical for the development of cardiovascular lesions during KD. Sirtuin 1 (SIRT1), a nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylase, is crucial for responding to cellular stress, and regulates NLRP3 activation as well as clearance of damaged mitochondria in cardiovascular diseases. However, its role in KD pathophysiology remains unknown. Here, we used the Lactobacillus casei cell wall extract (LCWE) murine model of KD to determine the role of SIRT1 on the development of cardiovascular lesions. We show that Sirt1 expression is downregulated in LCWE-injected mice with a significant reduction of nicotinamide, a product resulting from NAD+ degradation by SIRT1. In addition, supplementation of mice with NAD+ precursors significantly reduced the development of LCWE-induced cardiovascular lesions. Furthermore, KD development was less severe in SIRT1 overexpressing mice but was exacerbated with the specific deletion of Sirt1 in vascular smooth muscle cells. Treatment of LCWE-stimulated coronary smooth muscle cells with a SIRT1 activator favored mitophagy by inhibiting ribosomal S6 phosphorylation. These results indicate an impaired NAD+/SIRT1 axis during LCWE-induced KD vasculitis, which could potentially be targeted to reduce cardiovascular lesions in KD.
Homogeneous bioluminescent immunoassays detect profound synergistic regulation of IL-8 and MCP-1 release in cultured human monocytes
Authors: Lazar Dan , Ames Cassidy , Sequeira Paula , Kupcho Kevin , Mamott Daniel , O'Brien Martha , Mahan Sarah , Cali James
Published: November 2024

We have developed sensitive bioluminescent immunoassays for IL-8 and MCP-1 and applied these homogeneous assays to demonstrate synergistic regulation of their release in a culture model of human monocytes. IL-8 and MCP-1, key proinflammatory chemokines, play important roles in neutrophil and monocyte recruitment. Employing Lumit™ technology, we labeled candidate monoclonal antibodies for these chemokines with NanoBiT luciferase subunits (SmBiT or LgBiT), identifying pairs that produce strong luminescent signals upon target binding. These assays exhibited outstanding sensitivity (IL-8 at 0.4 pg/mL, MCP-1 at 0.8 pg/mL) and a broad linear range over three logs of analyte concentration, with high specificity against a panel of cytokines. Applying these assays to THP-1 monocytes in 96-well format, we observed lipopolysaccharide (LPS) induced a dose-dependent IL-8 release (max 1532 pg/mL), whereas IFN-γ triggered minimal release (169 pg/mL). However, LPS and IFN-γ together significantly increased IL-8 levels (8554 pg/mL). More profound synergy was observed for MCP-1 release; LPS or IFN-γ alone elicited modest responses (23 and 48 pg/mL, respectively), but their combination led to a dramatic increase (5902 pg/mL). This work establishes rapid, simple luminescent immunoassays for IL-8 and MCP-1 detection directly in cell culture and shows application of these assays to demonstrate synergistic regulation of chemokine release with implications for modulation of inflammatory responses.
Utilizing a surgical resection model for evaluating mRNA-based multi-antigen targeting CAR-T cell therapy against solid tumors
Authors: Dagher Oula , Posey Avery
Published: November 2024

Chimeric antigen receptor (CAR)-T cell therapies have demonstrated considerable remissions against hematological malignancies but only transient remission against glioblastoma. CAR-T cell engineering relies on viral transduction or nonviral delivery such as mRNA incorporation. However, current in vivo models are inadequate for mRNA-based CAR-T cell evaluation due to short-term CAR expression. Here, we investigated multi-antigen targeting mRNA-based CAR-T cells locally injected in a xenograft mouse model after surgical resection of primary tumor. NSG mice, subcutaneously inoculated with glioma cells, were divided into groups receiving either mock T or CAR-T cells. A resection cutoff of minimal residual disease was used to subdivide mice based on resection intensity (desired or undesired). Anti-tumor cytotoxicity was both resection- and dose-dependent. Notably, >75% of mice undergoing undesired resection succumbed 49 days post-resection. However, CAR-T cell treated mice undergoing desired resection demonstrated complete or partial remissions along with significantly higher survival rates. Treatment of resection models with multivariant mRNA-based CAR-T cells demonstrates the translational utility of nonviral CAR-T cell approaches, building on previous reports targeting multiple antigens to offset solid tumor evasion. To sum up, our xenograft resection/CAR-T cell model is better tailored for evaluation of mRNA-based CAR-T cell therapies against solid tumors than existing models.