PUBLICATIONS

BACKGROUND: Taiwan experienced a major dengue outbreak in 2023 following the relaxation of COVID-19 border controls. The contributing factors remained unclear. This study investigated potential virological, immunological, and clinical drivers.

METHODS: We retrospectively analyzed laboratory-confirmed dengue virus (DENV) infections at a tertiary care hospital in southern Taiwan. Serotypes were identified by qRT-PCR. Viral origins were assessed through phylogenetic and envelope (E) gene amino acid analyses. Clinical features of DENV-1 and DENV-2 cases were compared. Neutralization and antibody-dependent enhancement (ADE) were evaluated using PRNT and ADE assays.

RESULTS: DENV-1 and DENV-2 were identified as the predominant circulating serotypes. Clinical analysis revealed that DENV-2 infection was significantly associated with older age, diabetes mellitus, hypertension, and elevated hematocrit levels (p < 0.05), and these associations remained statistically significant in multivariate analysis. Phylogenetic analysis showed that DENV-1 isolates belonged to genotypes I and IV, while DENV-2 strains were of the cosmopolitan genotype. These viruses clustered closely with strains from Southeast Asia. Amino acid analysis indicated that DENV-1 strains exhibited 2-10 substitutions relative to 2014 isolates, while DENV-2 strains closely matched those from 2015. Sera from the 2014-2015 outbreaks demonstrated potent homotypic but limited heterotypic neutralization. ADE was observed in heterotypic infection contexts.

CONCLUSIONS: The 2023 dengue outbreak in Taiwan was driven by co-circulation of DENV-1 and DENV-2, limited heterotypic immunity, and ADE. These findings highlight the importance of integrated virological surveillance, genotype monitoring, and immunological assessment to inform dengue control strategies in non-endemic regions experiencing imported viral threats.

BACKGROUND: Strengthening mRNA vaccine development in LMICs is essential for enhancing global pandemic preparedness. This study evaluated the safety and immunogenicity of Comvigen, a bivalent SARS-CoV-2 vaccine, in comparison to the Comirnaty bivalent vaccine (Comirnaty).

METHODS: This phase II, randomised, open-label, non-inferiority trial was conducted in Thailand across four centres. Participants (n = 450) were randomly assigned (2:1) to receive either Comvigen (50 μg) or Comirnaty (30 μg), using block randomisation (size = 9). Eligible participants had completed at least 2 doses of any approved COVID-19 vaccine, with the last mRNA-vaccine dose given over 3 months before enrolment. The non-inferiority margin of a geometric mean ratio (GMR) of 0.67. The primary immunogenicity endpoint was pseudovirus neutralisation titres (psVNT-50) against SARS-CoV-2 wild-type and Omicron BA.4/BA.5 at Day 29. Safety outcomes included local and systemic adverse reactions up to six months post-vaccination. Immunogenicity analyses were conducted on the Per-Protocol (PP) population and the modified Intent-to-Treat (mITT) population; safety analyses included all participants. Laboratory personnel were blinded to vaccine assignment (ClinicalTrials.gov: NCT05930730).

FINDINGS: Between October and November 2023, 450 participants were enrolled (median age of 36 years, IQR 30-45). At day 29, the geometric mean titre (GMT) of psVNT-50 against wild-type virus increased from 475.9 to 2062.9 for Comvigen and from 458.8 to 1905.1 for Comirnaty (GMR 1.1, 95% CI: 1.0-1.2), meeting non-inferiority criteria. Against Omicron BA.4/BA.5, GMTs were 3909.8 for Comvigen and 3288.6 for Comirnaty (GMR 1.2, 95% 1.0-1.4). Local and systemic reactions were more frequent with Comvigen (91% vs. 78%, p = 0.0002, 79% vs. 70%, p = 0.028) but were mild or moderate and transient with no difference in fever (6% vs. 5%, p = 0.84).

INTERPRETATION: Comvigen demonstrated non-inferiority immunogenicity to Comirnaty and had a comparable safety profile, supporting mRNA vaccine development for global access and pandemic preparedness.

FUNDING: Covid-19 Pandemic Emergency Fund granted by Thailand's National Economic and Social Development Council provided major funding. Supplementary funding was provided by National Vaccine Institute (NVI), Thailand; Center of Excellence in Vaccine Research and Development (Chula VRC), Faculty of Medicine, Chulalongkorn University; Chulalongkorn University Second Century Fund (C2F); BioNet-Asia and Public Donation through Covid-19 vaccine development fund of the Faculty of Medicine, Chulalongkorn University and the Thai Red Cross Society, Thailand.

Severe diseases like severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreak urge efficient discovery of drugs for emergent needs and precision medicine. This study demonstrates an interdisciplinary solution by identification of a chief anti-SARS-CoV-2 small molecule, 2,3,5,8-tetrahydroxy-6-methylnaphthalene-1,4-dione (TMD) from a Taiwanese traditional medicine, Antrodia cinnamomea. TMD was found to inhibit the key viral replication enzymes, 3-chymotrypsin-like protease (3CLpro) and papain-like protease (PLpro), by structure-based prediction analysis using the Explore program of BT&D2 drug targeting system. Subsequently, TMD was total-synthesized and shown to inhibit Omicron BA.2 and BA.5 variants. The enzymatic inhibition kinetics studies revealed the competitive inhibition and dual-target nature of TMD and its potential capability to target the proteases in mixed-type mode, consistent with the in silico mechanistic analysis. The predicted less off-target side effects of TMD also agreed with the animal toxicity test result of no apparent toxicity. This study provides new insight into anti-viral mechanism of Antrodia cinnamomea and a new lead drug with improved pharmacokinetics to combat SARS-CoV-2 infection.

This study explores immune responses in mild Omicron-era COVID-19 breakthrough cases, focusing on cytokine dysregulation, antibody dynamics, and Long COVID. Samples from 114 mild COVID-19 patients across multiple waves were analyzed at three timepoints (T1: 2-4 weeks, T2: 3-4 months, T3: 6-8 months post-infection). Persistent IP-10 elevation up to 8 months suggests prolonged low-grade immune activation. Hybrid immunity from Omicron breakthrough infections provided broad cross-variant antibody recognition but showed declining neutralization over time. Among vaccination regimens, mRNA-inclusive combinations were associated with lower Long COVID scores. CoV-229E antibody levels correlated with Long COVID scores. These findings underscore the need for extended monitoring of mild COVID-19 cases and highlight the potential of mRNA vaccines in reducing post-COVID-19 complications. Insights into immune alterations and vaccine effects can inform the development of future vaccination strategies and approaches for managing post-COVID-19 conditions.

Galectins are a family of β-galactosides-binding protein, crucial regulators of host-virus interactions. They achieve this by recognizing specific glycan patterns on viral surfaces or mediating interactions with intracellular viral or host proteins, subsequently influencing the critical phases of the viral life cycle, such as attachment, replication, immune evasion, and reactivation. Furthermore, galectins modulate host immune responses, shaping the progression and outcomes of viral infections. This review comprehensively examines the roles of both endogenous and exogenous galectins in viral infections, noting that only a few galectins, including Galectin-1, -3, -4, -7, -8, and - 9, Have been identified as key players in viral infection. Notably, Galectin-1, -3, and - 9 play diverse functions in both DNA and RNA viral infection. Emerging evidence highlights the potential of Galectin-4 and -8 as intracellular sensors and modulators of viral pathogenesis. Endogenous galectins, produced by host cells, act through both glycan-dependent and glycan-independent mechanisms, influencing viral processes and immune responses. Exogenous galectins, which are secreted by other cells or administered as recombinant proteins, can either enhance or counteract the actions of endogenous galectins. The functions of galectins are virus-specific and context-dependent, serving as either promoters or inhibitors of viral replication and reactivation. Dysregulation of galectin expression is often linked to disease progression, highlighting their potential as diagnostic and prognostic biomarkers, as well as therapeutic targets. The important and varied roles that galectins play in viral infections are highlighted in this review, which also provides fresh insights into host-pathogen interactions and the development of antiviral tactics.

Monkeypox virus (Mpox) has been recognized for causing distinct skin lesions and is primarily transmitted through skin and sexual contact. To date, the transmissibility and pathogenesis of the Mpox virus in distal human lung has never been completely explored. Here the transmission pathways and Mpox tropism on patient-derived air-liquid epithelium (ALE) model fabricated using isolated primary human alveolar epithelial cells (hAECs) were investigated. hAECs were cultured and exposed to the Mpox virus clade IIb isolated from patient. DNA, proteins, and the tropism were elucidated using polymerase chain reaction (PCR), Western blot and high-content fluorescent imaging. Transmission electron microscopy (TEM) was employed to systematically observe the cellular distribution of viral particles. Viral titers were determined by TCID50 assay. Innate immune response and inflammatory mediators were measured using Milliplex® multiplex and ELISA analysis. Pathology at alveolar barrier integrity was determined using transepithelial electrical resistance (TEER) analysis. The study included mock-infected cells as control. Mpox virus significantly infected 42.82% of total hAEC populations. The prominent observed pathology included a significant reduction in TEER values, loss of tight junction protein, presence of tunneling nanotubes (TNTs) and syncytium morphology. Four stages of Mpox biogenesis were clearly observed without significant activation of IL-6, MIP1alpha, TNF-α, and Galectin-9, although IL-1β were subtly promoted. The developed patient-derived ALE is a versatile model for Mpox virus clade IIb infection reflecting respiratory transmission competence of the Mpox. Postinfection lung pathogenesis demonstrated alveolar barrier damage without significant inflammation, raising concerns about possible immune evasion by the virus.

In our study, Prodrug nucleoTide (ProTide) technology was applied to cordycepin to enhance its antiviral activities and metabolic stability. Using cordycepin as starting material, we developed a synthetic method to access a series of stereospecific-phosphoramidate derivatives with various ester groups. We also successfully synthesized halogenated cordycepin derivatives via stannylation. Our 17 ProTide-cordycepin derivatives were pharmacologically evaluated for their antiviral activities. Phosphorus diastereomers 22 S and 22 R showed moderate inhibitory activity against corona and influenza viruses, while these compounds and derivatives (25 S, 27 S, and 27 R) demonstrated promising antiviral efficacy against dengue virus. Pharmacological screening indicated that Sp-isomers generally exhibited slightly greater inhibitory activity than their Rp-isomer counterparts against the dengue virus. The selected ProTides were assessed for their metabolic mechanism and stability via carboxypeptidase and microsomes. The hydrolysis rate of the Rp-isomers was observed to be slightly higher than that of the Sp-isomers, and the addition of a fluorine group also modestly increased this rate and fluorinated 39 S extended its half-life compared to nonfluorinated counterparts. These findings suggested not only structure-activity relationships of cordycepin ProTide but also the comprehensive synthetic route to access cordycepin derivatives for further antiviral development.

Establishing a drug-screening platform is critical for the discovery of potential antiviral agents against SARS-CoV-2. In this study, we developed a platform based on human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) to investigate SARS-CoV-2 infectivity, with the aim of evaluating potential antiviral agents for anti-SARS-CoV-2 activity and cardiotoxicity. Cultured myocytes of iPSC-CMs and immortalized human cardiomyocyte cell line (AC-16) were primarily characterized for the expression of cardiac markers and host receptors of SARS-CoV-2. An infectivity model for the wild-type SARS-CoV-2 strain was then established. Infection modeling involved inoculating cells with SARS-CoV-2 at varying multiplicities of infection (MOIs) and then quantifying infection using immunofluorescence and plaque assays. Only iPSC-CMs, not AC16 cells, expressed angiotensin-converting enzyme 2 (ACE-2), and quantitative assays confirmed the dose-dependent infection of iPSC-CMs by SARS-CoV-2, unlike the uninfectable AC16 cells lacking the expression of ACE2. Cytotoxicity was evaluated using MTT assays across a concentration range. An assessment of the plant-derived compound panduratin A (panA) showed cytotoxicity at higher doses (50% cytotoxic concentration (CC50) 10.09 μM) but promising antiviral activity against SARS-CoV-2 (50% inhibition concentration (IC50) 0.8-1.6 μM), suppressing infection at concentrations 10 times lower than its CC50. Plaque assays also showed decreased viral production following panA treatment. Overall, by modeling cardiac-specific infectivity, this iPSC-cardiomyocyte platform enables the reliable quantitative screening of compound cytotoxicity alongside antiviral efficacy. By combining disease pathogenesis and pharmacology, this system can facilitate the evaluation of potential novel therapeutics, such as panA, for drug discovery applications.

The ongoing COVID-19 pandemic has triggered extensive research, mainly focused on identifying effective therapeutic agents, specifically those targeting highly pathogenic SARS-CoV-2 variants. This study aimed to investigate the in vivo antiviral efficacy and anti-inflammatory activity of herbal extracts derived from Andrographis paniculata and Boesenbergia rotunda, using a Golden Syrian hamster model infected with Delta, a representative variant associated with severe COVID-19. Hamsters were intranasally inoculated with the SARS-CoV-2 Delta variant and orally administered either vehicle control, B. rotunda, or A. paniculata extract at a dosage of 1000 mg/kg/day. Euthanasia was conducted on days 1, 3, and 7 post-inoculation, with 4 animals per group. The results demonstrated that oral administration of A. paniculata extract significantly alleviated both lethality and infection severity compared with the vehicle control and B. rotunda extract. However, neither extract exhibited direct antiviral activity in terms of reducing viral load in the lungs. Nonetheless, A. paniculata extract treatment significantly reduced IL-6 protein levels in the lung tissue (7278 ± 868.4 pg/g tissue) compared to the control (12,495 ± 1118 pg/g tissue), indicating there was a decrease in local inflammation. This finding is evidenced by the ability of A. paniculata extract to reduce histological lesions in the lungs of infected hamsters. Furthermore, both extracts significantly decreased IL-6 and IP-10 mRNA expression in peripheral blood mononuclear cells of infected hamsters compared to the control group, suggesting systemic anti-inflammatory effects occurred. In conclusion, A. paniculata extract's potential therapeutic application for SARS-CoV-2 arises from its observed capacity to lessen inflammatory cytokine concentrations and mitigate lung pathology.

BACKGROUND: Andrographolide is a medicinal compound which possesses anti-SARS-CoV-2 activity. A number of cellular targets of andrographolide have been identified by target predictions and computational studies.

PURPOSE: However, a potential cellular target of andrographolide has never been explored in SARS-CoV-2 infected lung epithelial cells. We aimed to identify cellular pathways involved in andrographolide-mediated anti-SARS-CoV-2 activity.

METHODS: The viral infection was determined by immunofluorescence staining, enzyme-linked immunosorbent assay and focus-forming assay. Proteomic analysis was employed to identify cellular pathways and key proteins controlled by andrographolide in the human lung epithelial cells Calu-3 infected by SARS-CoV-2. Immunofluorescence staining was used to test protein expression and localization. Western blot and realtime PCR were utilized to elucidate gene expression. Cellular glutathione level was examined by a reduced/oxidized glutathione assay. An ectopic gene expression was delivered by plasmid transfection.

RESULTS: Gene ontology analysis indicates that proteins involved in nuclear factor erythroid 2-related factor 2 (NRF2)-regulated pathways were differentially expressed by andrographolide. Notably, andrographolide increased expression and nuclear localization of the transcription factor NRF2. In addition, transcriptional expression of GCLC and glutamate-cysteine ligase modifier subunit (GCLM), which are NRF2 target genes, were induced by andrographolide. We further find that infection of SARS-CoV-2 resulted in a reduction of glutathione level in Calu-3; the effect that was rescued by andrographolide. Moreover, andrographolide also induced expression of the glutathione producing enzyme GCLC in SARS-CoV-2 infected lung epithelial cells. Importantly, an ectopic over-expression of GCLC or treatment of N-acetyl-L-cysteine in Calu-3 cells led to a decrease in SARS-CoV-2 infection.

CONCLUSION: Collectively, our findings suggest the interplay between GCLC-mediated glutathione biogenesis induced by andrographolide and the anti-SARS-CoV-2 activity. The glutathione biogenesis and recycling pathways should be further exploited as a targeted therapy against SARS-CoV-2 infection.