PROTAC unleashed: Unveiling the synthetic approaches and potential therapeutic applications.

Proteolysis-Targeting Chimeras (PROTACs) are a novel class of bifunctional small molecules that alter protein levels by targeted degradation. This innovative approach uses the ubiquitin-proteasome system to selectively eradicate disease-associated proteins, providing a novel therapeutic strategy for a wide spectrum of diseases. This review delineates detailed synthetic approaches involved in PROTAC building blocks, including the ligand and protein binding parts, linker attached structural components of PROTACs and the actual PROTAC molecules. Furthermore, the recent advancements in PROTAC-mediated degradation of specific oncogenic and other disease-associated proteins, such as those involved in neurodegenerative, antiviral, and autoimmune diseases, were also discussed. Additionally, we described the current landscape of PROTAC clinical trials and highlighted key studies that underscore the translational potential of this emerging therapeutic modality. These findings demonstrate the versatility of PROTACs in modulating the levels of key proteins involved in various severe diseases.

Acute Chikungunya Virus Infection Triggers a Diverse Range of T Helper Lymphocyte Profiles.

Chikungunya virus (CHIKV) is an arbovirus causing acute febrile illness with severe joint pain, often leading to chronic arthralgia. This study investigated the adaptive immune responses during the early stages of symptomatic acute CHIKV infection, focusing on the transcription factors and cytokines linked to Th1, Th2, Th17, and Treg cells. Thirty-six individuals were enrolled: nine healthy controls and 27 CHIKV-positive patients confirmed by qRT-PCR. Blood samples were analyzed for the mRNA expression of transcription factors (Tbet, GATA3, FoxP3, STAT3, RORγt) and cytokines (IFN-γ, IL-4, IL-17, IL-22, TGF-ß, IL-10). The results showed the significant upregulation of Tbet, GATA3, FoxP3, STAT3, and RORγt in CHIKV-positive patients, with RORγt displaying the highest increase. Correspondingly, cytokines IFN-γ, IL-4, IL-17, and IL-22 were upregulated, while TGF-ß was downregulated. Principal component analysis (PCA) confirmed the distinct immune profiles between CHIKV-positive and healthy individuals. A correlation analysis indicated that higher Tbet expression correlated with a lower viral load, whereas FoxP3 and TGF-ß were associated with higher viral loads. Our study sheds light on the intricate immune responses during acute CHIKV infection, characterized by a mixed Th1, Th2, Th17, and Treg response profile. These results emphasize the complex interplay between different adaptive immune responses and how they may contribute to the pathogenesis of Chikungunya fever.

An in-depth exploration of snake venom-derived molecules for drug discovery in advancing antiviral therapeutics.

Snake venom is a cocktail and rich source of various bioactive compounds that have been extensively studied for their potential as pharmaceutical agents due to their diverse chemical structures and wide range of biological activities. In light of the emergency and the re-emergence of viral infectious diseases that threaten human health and economic systems, exploring new fertile and rich fields such as snake venom is an attractive path for anti-viral drug discovery, especially in the lack of effective vaccines. Although 85 % of reported antiviral molecules belong to the phospholipase A2 (PLA2) family, other protein families including L-amino acid oxidases (LAAO), disintegrins, metalloproteases (SVMPs), and cathelicidins have also shown antiviral activity. Thus, in this review, we have highlighted the antiviral properties of compounds derived from snake venom and their mechanisms of action against virus classes like HIV, Coronaviridae, Flaviviridae, and Paramyxoviridae. Although the initial research emphasis has been on Retroviridae (HIV) and Flaviviridae viruses, it is crucial to extend the exploration of the potential of these compounds to other viruses. The utilization of snake venom-derived compounds as antivirals shows significant promise for the development of novel therapeutics to address viral infections. However, a more in-depth investigation is necessary to fully assess the potential of these compounds against other viruses and unveil the mechanisms underlying their action.

Safety of remdesivir in the treatment of acute SARS-CoV-2 infection in pediatric patients.

BACKGROUND: Transaminase and creatinine elevations have been well described in adults treated with remdesivir for COVID-19. It is hypothesized that a similar safety profile exists in children with COVID-19 treated with remdesivir, but available data are limited, especially in children < 12 months. The primary aim of this study was to determine the prevalence and timing of elevations in transaminases and creatinine in children with COVID-19 who were treated with remdesivir. METHODS: This was a retrospective, observational cohort study including all pediatric patients admitted to a single, freestanding children's hospital who were positive for COVID-19 and received at least 1 dose of remdesivir between 1/1/2020 and 5/31/2022. Available baseline and peak transaminase and creatinine concentrations were evaluated. Multivariable logistic regression analysis was performed to identify risk factors for transaminase elevation. RESULTS: A total of 180 patients met inclusion criteria. Creatinine elevation of any grade was noted in 16% and remained elevated only in those with underlying chronic kidney disease. Transaminase elevation of any grade was noted in 58% of patients and remained elevated in only 1%. Older age and critical respiratory disease were associated with higher risk of significant transaminase elevation, whereas non-Hispanic ethnicity was strongly associated with protection against significant transaminase elevation. CONCLUSIONS: In our cohort of hospitalized children with COVID-19 who were treated with remdesivir, most patients experienced only mild transaminitis and normal creatinine concentrations. A limited number of patients experienced laboratory abnormalities which were transient, suggesting a favorable safety profile for remdesivir use in pediatrics.

Post-Chimeric Antigen Receptor T-Cell Therapy Hepatitis B Virus Reactivation After 23 Months of Entecavir Prophylaxis.

Hepatitis B virus (HBV) reactivation can occur in immunosuppressed patients. Specifically, HBV reactivation after chimeric antigen receptor T-cell (CAR T-cell) therapy is a known complication with few case reports and specific treatment guidelines. Our patient experienced HBV reactivation 27 months after CAR T-cell therapy even with 23 months of entecavir prophylaxis. This unique case highlights the need for further investigation into the risk of HBV reactivation after CAR T-cell therapy and the proper HBV prophylaxis during and after CAR T-cell therapy.

A brief review on the lessons learned from COVID-19 on drug discovery and research.

COVID-19 pandemic has taught many lessons regarding drug discovery and development. This review covers these aspects of drug discovery and research for COVID-19 which might be used as a tool for future. It summarizes the positives such as progresses in antiviral drug discovery, drug repurposing, adaptations of clinical trial and its regulations, as well as the negative points such as the need to develop more collaboration among stakeholders and future directions. It also discusses the benefits and limitations of finding new indications for existing drugs, and the lessons learned regarding rigorous and robust clinical trials, pharmacokinetic/pharmacodynamic modelling, as well as combination therapy. The pandemic has also revealed some gaps regarding global collaboration and coordination, data sharing and transparency and equitable distribution. Finally, the review enumerates the future directions and implications of drug discovery and research for COVID-19 and other infectious diseases such as preparedness and resilience, interdisciplinary and integrative approaches, diversity and inclusion, and personalized and precision medicine.

Dual Role of Extracellular Vesicles as Orchestrators of Emerging and Reemerging Virus Infections.

Current decade witnessed the emergence and re-emergence of many viruses, which affected public health significantly. Viruses mainly utilize host cell machinery to promote its growth, and spread of these diseases. Numerous factors influence virus-host cell interactions, of which extracellular vesicles play an important role, where they transfer information both locally and distally by enclosing viral and host-derived proteins and RNAs as their cargo. Thus, they play a dual role in mediating virus infections by promoting virus dissemination and evoking immune responses in host organisms. Moreover, it acts as a double-edged sword during these infections. Advances in extracellular vesicles regulating emerging and reemerging virus infections, particularly in the context of SARS-CoV-2, Dengue, Ebola, Zika, Chikungunya, West Nile, and Japanese Encephalitis viruses are discussed in this review.

Targeting stress induction of GRP78 by cardiac glycoside oleandrin dually suppresses cancer and COVID-19.

BACKGROUND: Despite recent therapeutic advances, combating cancer resistance remains a formidable challenge. The 78-kilodalton glucose-regulated protein (GRP78), a key stress-inducible endoplasmic reticulum (ER) chaperone, plays a crucial role in both cancer cell survival and stress adaptation. GRP78 is also upregulated during SARS-CoV-2 infection and acts as a critical host factor. Recently, we discovered cardiac glycosides (CGs) as novel suppressors of GRP78 stress induction through a high-throughput screen of clinically relevant compound libraries. This study aims to test the possibility that agents capable of blocking stress induction of GRP78 could dually suppress cancer and COVID-19. RESULTS: Here we report that oleandrin (OLN), is the most potent among the CGs in inhibiting acute stress induction of total GRP78, which also results in reduced cell surface and nuclear forms of GRP78 in stressed cells. The inhibition of stress induction of GRP78 is at the post-transcriptional level, independent of protein degradation and autophagy and may involve translational control as OLN blocks stress-induced loading of ribosomes onto GRP78 mRNAs. Moreover, the human Na+/K+-ATPase α3 isoform is critical for OLN suppression of GRP78 stress induction. OLN, in nanomolar range, enhances apoptosis, sensitizes colorectal cancer cells to chemotherapeutic agents, and reduces the viability of patient-derived colon cancer organoids. Likewise, OLN, suppresses GRP78 expression and impedes tumor growth in an orthotopic breast cancer xenograft model. Furthermore, OLN blocks infection by SARS-CoV-2 and its variants and enhances existing anti-viral therapies. Notably, GRP78 overexpression mitigates OLN-mediated cancer cell apoptotic onset and suppression of virus release. CONCLUSION: Our findings validate GRP78 as a target of OLN anti-cancer and anti-viral activities. These proof-of-principle studies support further investigation of OLN as a readily accessible compound to dually combat cancer and COVID-19.

Clinical rebound after treatment with nirmatrelvir/ritonavir in COVID-19.

BACKGROUND: Nirmatrelvir/ritonavir (NM/r) is a safe and effective oral antiviral therapeutic used for treatment of mild-to-moderate COVID-19. Case reports described a clinical rebound syndrome whereby individuals experience a relapse of symptoms shortly after completing successful treatment. There is a lack of information on frequency of COVID-19 rebound after NM/r in routine clinical care, contributing factors, and clinical outcomes. METHODS: We reviewed electronic medical records to verify COVID-19 diagnosis, symptoms, and treatment with NM/r from January-June 2022. We defined COVID-19 clinical rebound as clear improvement in symptoms followed by recurrence or worsening of symptoms within 30 days of a five-day course of NM/r. RESULTS: We studied 268 adults with median age 57 (IQR 47, 68), 80% White race, 85% non-Hispanic ethnicity, 55% female, 80% vaccinated and boosted against SARS-CoV-2, and 68% with any co-morbidity. Sixteen (6.0%) of studied patients were determined to have COVID-19 clinical rebound. The median time from starting NM/r to rebound was 11 days (IQR 9, 13). Notable demographic and clinical factors with higher proportion (not statistically significant) among COVID-19 rebound patients were female sex (75% rebound vs. 54.5% no rebound), Black race (12.5% rebound vs. 4.9% no rebound), presence of at least one co-morbidity (81.3% rebound vs. 67.5% no rebound), and lack of prior SARS-CoV-2 infection (100% rebound vs. 92.9% no rebound). Only one patient (6.25%) was hospitalized after COVID-19 rebound. CONCLUSIONS: COVID-19 clinical rebound after treatment with NM/r is mild with favorable outcomes and more common than previously reported from real-world clinical care studies.

SARS-CoV-2 papain-like protease inhibits ISGylation of the viral nucleocapsid protein to evade host anti-viral immunity.

A severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection causes mild-to-severe respiratory symptoms, including acute respiratory distress. Despite remarkable efforts to investigate the virological and pathological impacts of SARS-CoV-2, many of the characteristics of SARS-CoV-2 infection still remain unknown. The interferon-inducible ubiquitin-like protein ISG15 is covalently conjugated to several viral proteins to suppress their functions. It was reported that SARS-CoV-2 utilizes its papain-like protease (PLpro) to impede ISG15 conjugation, ISGylation. However, the role of ISGylation in SARS-CoV-2 infection remains unclear. We aimed to elucidate the role of ISGylation in SARS-CoV-2 replication. We observed that the SARS-CoV-2 nucleocapsid protein is a target protein for the HERC5 E3 ligase-mediated ISGylation in cultured cells. Site-directed mutagenesis reveals that the residue K374 within the C-terminal spacer B-N3 (SB/N3) domain is required for nucleocapsid-ISGylation, alongside conserved lysine residue in MERS-CoV (K372) and SARS-CoV (K375). We also observed that the nucleocapsid-ISGylation results in the disruption of nucleocapsid oligomerization, thereby inhibiting viral replication. Knockdown of ISG15 mRNA enhanced SARS-CoV-2 replication in the SARS-CoV-2 reporter replicon cells, while exogenous expression of ISGylation components partially hampered SARS-CoV-2 replication. Taken together, these results suggest that SARS-CoV-2 PLpro inhibits ISGylation of the nucleocapsid protein to promote viral replication by evading ISGylation-mediated disruption of the nucleocapsid oligomerization.IMPORTANCEISG15 is an interferon-inducible ubiquitin-like protein that is covalently conjugated to the viral protein via specific Lys residues and suppresses viral functions and viral propagation in many viruses. However, the role of ISGylation in SARS-CoV-2 infection remains largely unclear. Here, we demonstrated that the SARS-CoV-2 nucleocapsid protein is a target protein for the HERC5 E3 ligase-mediated ISGylation. We also found that the residue K374 within the C-terminal spacer B-N3 (SB/N3) domain is required for nucleocapsid-ISGylation. We obtained evidence suggesting that nucleocapsid-ISGylation results in the disruption of nucleocapsid-oligomerization, thereby suppressing SARS-CoV-2 replication. We discovered that SARS-CoV-2 papain-like protease inhibits ISG15 conjugation of nucleocapsid protein via its de-conjugating enzyme activity. The present study may contribute to gaining new insight into the roles of ISGylation-mediated anti-viral function in SARS-CoV-2 infection and may lead to the development of more potent and selective inhibitors targeted to SARS-CoV-2 nucleocapsid protein.

Crystal structure of NYN domain of Human KHNYN in complex with single strand RNA.

KHNYN protein with a KH-like domain and a NYN endoribonuclease domain interacts with Zinc-finger antiviral protein (ZAP). ZAP isoforms recognize viral or cellular RNAs and recruit KHNYN to form the ZAP: KHNYN complex. Although the structures of several PIN/NYN domains have been determined, the precise substrate RNA binding mode remains poorly understood. This study presents the crystal structure of a complex of the NYN domain of KHNYN and a 7mer RNA from interferon lambda3 (IFNL3). Our structural analysis reveals that NYN domain of human KHNYN shares structural similarities with other NYN domains of ZC3H12àC proteins. The RNA is bound in the central groove region of the protein, facilitated by interactions including coordination by two Mg2+ ions, hydrophobic interactions, and hydrogen bonds. In the observed RNA-protein complex, the U5, A6, and U7 bases are stacked on top of one another, while U3 and U4 bases adopt an "open" conformation (as opposed to base-stacked), forming a U-shaped overall structure. Mutagenesis studies underscore the significance of residues involved in RNA binding for RNase activity. Interestingly, NYN domain of human KHNYN forms a head-to-tail dimer in the crystal, a structural feature also observed in other homologous PIN/NYN proteins, with a residue from the symmetry mate contributing to hydrophobic interactions with the bound RNA.

Direct-acting antivirals for RSV treatment, a review.

Respiratory syncytial virus (RSV) causes respiratory disease and complications in infants, the elderly and the immunocompromised. While three vaccines and two prophylactic monoclonal antibodies are now available, only one antiviral, ribavirin, is currently approved for treatment. This review aims to summarize the current state of treatments directly targeting RSV. Two major viral processes are attractive for RSV-specific antiviral drug discovery and development as they play essential roles in the viral cycle: the entry/fusion process carried out by the fusion protein and the replication/transcription process carried out by the polymerase complex constituted of the L, P, N and M2-1 proteins. For each viral target resistance mutations to small molecules of different chemotypes seem to delineate definite binding pockets in the fusion proteins and in the large proteins. Elucidating the mechanism of action of these inhibitors thus helps to understand how the fusion and polymerase complexes execute their functions. While many inhibitors have been studied, few are currently in clinical development for RSV treatment: one is in phase III, three in phase II and two in phase I. Progression was halted for many others because of strategic decisions, low enrollment, safety, but also lack of efficacy. Lessons can be learnt from the halted programs to increase the success rate of the treatments currently in development.

COVID-19 clinical rebound after treatment with nirmatrelvir/ritonavir.

Background: Nirmatrelvir/ritonavir (NM/r) is a safe and effective oral antiviral therapeutic used for treatment of mild-to-moderate COVID-19. Case reports described a clinical rebound syndrome whereby individuals experience a relapse of symptoms shortly after completing successful treatment. There is a lack of information on frequency of COVID-19 rebound after NM/r in routine clinical care, contributing factors, and clinical outcomes. Methods: We reviewed electronic medical records to verify COVID-19 diagnosis, symptoms, and treatment with NM/r from January-June 2022. We defined COVID-19 clinical rebound as clear improvement in symptoms followed by recurrence or worsening of symptoms within 30 days of a five-day course of NM/r. Results: We studied 268 adults with median age 57 (IQR 47, 68), 80% White race, 85% non-Hispanic ethnicity, 55% female, 80% vaccinated and boosted against SARS-CoV-2, and 68% with any co-morbidity. Sixteen (6.0%) of studied patients were determined to have COVID-19 clinical rebound. The median time from starting NM/r to rebound was 11 days (IQR 9, 13). Notable demographic and clinical factors with higher proportion (not statistically significant) among COVID-19 rebound patients were female sex (75% rebound vs 54.5% no rebound), Black race (12.5% rebound vs 4.9% no rebound), presence of at least one co-morbidity (81.3% rebound vs 67.5% no rebound), and lack of prior SARS-CoV-2 infection (100% rebound vs 92.9% no rebound). Only one patient (6.25%) was hospitalized after COVID-19 rebound. Conclusions: COVID-19 clinical rebound after treatment with NM/r is mild with favorable outcomes and more common than previously reported from real-world clinical care studies.

Owner experience and veterinary involvement with unlicensed GS-441524 treatment of feline infectious peritonitis: a prospective cohort study.

Introduction: Feline Infectious Peritonitis (FIP) has historically been a fatal coronavirus disease in cats. In recent years, the therapeutic agent GS-441524, developed by Gilead Sciences, was found to be a successful treatment for FIP in most patients in clinical trials. However, this particular drug has remained stalled in the therapeutic pipeline, leaving patients and cat owners without a licensed medication. In the meantime, online social media platforms began to emerge, connecting cat owners with a community of citizen non-veterinary professionals sourcing unlicensed GS-441524. Methods: This study prospectively followed participants (N = 141) that successfully completed 12 weeks of treatment, capturing their treatment experiences with self-administered GS-441524-like medication. A one-time survey was administered to enrolled participants with mixed format of questions (open-ended and multiple-choice) asking about treatment administration techniques, observed side effects of GS-441524, accrued cost, veterinarian involvement, impact on the cat-human bond, and social media usage. Results: Our results show cat owners experienced a shift in treatment modality from injectable GS-441524 to pill formulation across the treatment period. The average total cost of medication has decreased since 2021 to approximately USD 3100, and participants reported the human-animal bond being affected negatively. Additionally, there was an increased trend in veterinarian awareness of GS-441524-like therapeutics and monitoring of clients undergoing treatment. Social media usage was reported as being important at the beginning of treatment to establish treatment administration but lessened by the end of treatment. Discussion: This study is the first detailed, prospective account of owner experiences with unlicensed GS-441524, raising an important discussion surrounding citizen veterinary medicine.

The safety of antivirals and neutralising monoclonal antibodies used in prehospital treatment of Covid-19.

OBJECTIVE: This proof-of-principle pharmacovigilance study used Electronic Health Record (EHR) data to examine the safety of sotrovimab, paxlovid and molnupiravir in prehospital treatment of Covid-19. METHOD: With NHS England approval, we conducted an observational cohort study using OpenSAFELY-TPP, a secure software-platform which executes analyses across EHRs for 24 million people in England. High-risk individuals with Covid-19 eligible for prehospital treatment were included. Adverse events (AEs) were categorised into events in the drug's Summary of Product Characteristics (SmPC), drug-reactions and immune-mediated. Cox models compared risk across treatments. A pre-pandemic record analysis was performed for comparative purposes. RESULTS: Between 2021-2023, 37,449 patients received sotrovimab, paxlovid or molnupiravir whilst 109,647 patients made up an eligible-but-untreated population. The 28-day rates of AEs were low: SmPC 0.34 per 1000 patient-years (95% CI 0.32-0.36); drug-reactions 0.01 (95% CI 0.01-0.02) and immune-mediated 0.03 (95% CI 0.03-0.04), and similar or lower than the pre-pandemic period. Compared with the eligible but untreated population, sotrovimab and paxlovid associated with a risk of SmPC AE [adjHR 1.36 (95% CI 1.15-1.62) and 1.28 (95% CI 1.05-1.55), respectively], whilst sotrovimab associated with a risk of drug-reactions [adjHR 2.95 (95% CI 1.56-5.55)] and immune-mediated events [adjHR 3.22 (95% CI 1.86-5.57)]. CONCLUSION: Sotrovimab, paxlovid and molnupiravir demonstrate acceptable safety profiles. Although the risk of AEs was greatest with sotrovimab, event rates were lower than comparative pre-pandemic period.

The antiherpetic and anti-inflammatory activity of the frog-derived peptide Hylin-a1.

AIM: The high incidence of virus-related infections and the large diffusion of drug-resistant pathogens stimulate the search and identification of new antiviral agents with a broad spectrum of action. Antivirals can be designed to act on a single target by interfering with a specific step in the viral lifecycle. On the contrary, antiviral peptides (AVPs) are known for acting on a wide range of viruses, with a diversified mechanism of action targeting virus and/or host cell. In the present study, we evaluated the antiviral potential of the peptide Hylin-a1 secreted by the frog Hypsiobas albopunctatus against members of the Herpesviridae family. METHODS AND RESULTS: The inhibitory capacity of the peptide was evaluated in vitro by plaque assays in order to understand the possible mechanism of action. The results were also confirmed by real-time PCR and Western blot evaluating the expression of viral genes. Hylin-a1 acts to block the herpetic infection interfering at the early stages of both herpes simplex virus type 1 (HSV-1) and type 2 infection. Its mechanism is mainly directed on the membrane, probably by damaging the viral envelope. The same effect was also observed against HSV-1 strains resistant to acyclovir. CONCLUSIONS: The data presented in this study, such as the increased activity of the peptide when combined to acyclovir, a weak hemolytic profile, an anti-inflammatory effect, and a tolerable half-life in serum, indicates Hylin-a1 as a novel antiherpetic molecule with promising potential in the clinical setting.

ELISpots for Detecting Antigen-Specific Memory B Cells in Infection or Autoimmunity.

Enzyme-Linked Immunosorbent Spot assay (ELISpot) is an immunoassay used to quantify individual protein-specific secreting cells. Proteins secreted by cells cultured in ELISpot plates (96- or 8-well format) bind to a specific antigen bound to a PVDF membrane at the bottom of the well. A detection antibody followed by an enzymatic reaction is used to identify secreted protein bound to the membrane coated antigen. This reaction results in distinct "spots" on the membrane corresponding to individual protein secreting cells. While the design is similar to an ELISA, ELISpots quantify the number and relative amount of secreted protein on a single cell level, as opposed to an ELISA that reveals the concentration of secreted proteins from a population of cells. The sensitivity, robustness, and diversity of different antigens used by ELISpots have led to an array of research applications such as measuring cytokines from cytotoxic T cells in cancer and quantifying antibody specificity from B cells following vaccinations. Improvements have been made to assays measuring cytokines and antibodies on a single cell basis, such as intracellular flow cytometry. Yet the ability of an ELISpot to evaluate the quantity and quality of protein secretion on an individual cell basis remains unmatched. Here, we describe the use of a modified ELISpot assay to detect antigen-specific memory B cells in the setting of a viral infection and autoimmunity.

Sangju Cold Granule exerts anti-viral and anti-inflammatory activities against influenza A virus in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Sangju Cold Granule (SJCG) is a classical traditional Chinese medicine (TCM) prescription described in "Item Differentiation of Warm Febrile Diseases". Historically, SJCG was employed to treat respiratory illnesses. Despite its popular usage, the alleviating effect of SJCG on influenza A virus infection and its mechanisms have not been fully elucidated. AIM OF THE STUDY: Influenza is a severe respiratory disease that threatens human health. This study aims to assess the therapeutic potential of SJCG and the possible molecular mechanism underlying its activity against influenza A virus in vitro and in vivo. MATERIALS AND METHODS: Ultrahigh-performance liquid chromatography (UPLC)-Q-Exactive was used to identify the components of SJCG. The 50% cytotoxic concentration of SJCG in MDCK and A549 cells were determined using the CCK-8 assay. The activity of SJCG against influenza A virus H1N1 was evaluated in vitro using plaque reduction and progeny virus titer reduction assays. RT-qPCR was performed to obtain the expression levels of inflammatory mediators and the transcriptional regulation of RIG-I and MDA5 in H1N1-infected A549 cells. Then, the mechanism of SJCG effect on viral replication and inflammation was further explored by measuring the expressions of proteins of the RIG-I/NF-kB/IFN(I/III) signaling pathway by Western blot. The impact of SJCG was explored in vivo in an intranasally H1N1-infected BALB/c mouse pneumonia model treated with varying doses of SJCG. The protective role of SJCG in this model was evaluated by survival, body weight monitoring, lung viral titers, lung index, lung histological changes, lung inflammatory mediators, and peripheral blood leukocyte count. RESULTS: The main SJCG chemical constituents were flavonoids, carbohydrates and glycosides, amino acids, peptides, and derivatives, organic acids and derivatives, alkaloids, fatty acyls, and terpenes. The CC50 of SJCG were 24.43 mg/mL on MDCK cells and 20.54 mg/mL on A549 cells, respectively. In vitro, SJCG significantly inhibited H1N1 replication and reduced the production of TNF-α, IFN-ß, IL-6, IL-8, IL-13, IP-10, RANTES, TRAIL, and SOCS1 in infected A549 cells. Intracellularly, SJCG reduced the expression of RIG-I, MDA5, P-NF-κB P65 (P-P65), P-IκBα, P-STAT1, P-STAT2, and IRF9. In vivo, SJCG enhanced the survival rate and decreased body weight loss in H1N1-infected mice. Mice with H1N1-induced pneumonia treated with SJCG showed a lower lung viral load and lung index than untreated mice. SJCG effectively alleviated lung damage and reduced the levels of TNF-α, IFN-ß, IL-6, IP-10, RANTES, and SOCS1 in lung tissue. Moreover, SJCG significantly ameliorated H1N1-induced leukocyte changes in peripheral blood. CONCLUSIONS: SJCG significantly reduced influenza A virus and virus-mediated inflammation through inhibiting the RIG-I/NF-kB/IFN(I/III) signaling pathway. Thus, SJCG could provide an effective TCM for influenza treatment.

Self-inhibiting percolation and viral spreading in epithelial tissue.

SARS-CoV-2 induces delayed type-I/III interferon production, allowing it to escape the early innate immune response. The delay has been attributed to a deficiency in the ability of cells to sense viral replication upon infection, which in turn hampers activation of the antiviral state in bystander cells. Here, we introduce a cellular automaton model to investigate the spatiotemporal spreading of viral infection as a function of virus and host-dependent parameters. The model suggests that the considerable person-to-person heterogeneity in SARS-CoV-2 infections is a consequence of high sensitivity to slight variations in biological parameters near a critical threshold. It further suggests that within-host viral proliferation can be curtailed by the presence of remarkably few cells that are primed for IFN production. Thus, the observed heterogeneity in defense readiness of cells reflects a remarkably cost-efficient strategy for protection.

In vitro evaluation of antiviral activity in carnivorous plant species.

Sarracenia purpurea is a carnivorous plant historically used to treat smallpox infections. Our previous data found S. purpurea had broad spectrum antiviral activity in vitro. S. purpurea is one of several hundred identified carnivorous species of plants. Carnivorous plants have evolved through convergent evolution in at least ten independent events, usually in response to harsh environments where nutrition from prey is required for growth. These prey are known vectors of plant viruses that might introduce novel biotic stressors and defense pathways in carnivorous plants. This study evaluated the antiviral activity of several non-carnivorous and carnivorous plants from four evolutionarily distinct clades. Results demonstrated that carnivorous plants have evolved antiviral activity, a trait that is not present in related species of non-carnivorous plants. The antiviral trait may be due to the plant-prey relationship with insect vectors and an evolutionary need for carnivorous plants to have more robust antiviral defense systems.