Orally bioavailable RORγ/DHODH dual host-targeting small molecules with broad-spectrum antiviral activity.

Host-directed antivirals (HDAs) represent an attractive treatment option and a strategy for pandemic preparedness, especially due to their potential broad-spectrum antiviral activity and high barrier to resistance development. Particularly, dual-targeting HDAs offer a promising approach for antiviral therapy by simultaneously disrupting multiple pathways essential for viral replication. Izumerogant (IMU-935) targets two host proteins, (i) the retinoic acid receptor-related orphan receptor γ isoform 1 (RORγ1), which modulates cellular cholesterol metabolism, and (ii) the enzyme dihydroorotate dehydrogenase (DHODH), which is involved in de novo pyrimidine synthesis. Here, we synthesized optimized derivatives of izumerogant and characterized their antiviral activity in comparison to a recently described structurally distinct RORγ/DHODH dual inhibitor. Cell culture-based infection models for enveloped and non-enveloped DNA and RNA viruses, as well as a retrovirus, demonstrated high potency and broad-spectrum activity against human viral pathogens for RORγ/DHODH dual inhibitors at nanomolar concentrations. Comparative analyses with equipotent single-target inhibitors in metabolite supplementation approaches revealed that the dual-targeting mode represents the mechanistic basis for the potent antiviral activity. For SARS-CoV-2, an optimized dual inhibitor completely blocked viral replication in human airway epithelial cells at 5 nM and displayed a synergistic drug interaction with the nucleoside analog molnupiravir. In a SARS-CoV-2 mouse model, treatment with a dual inhibitor alone, or in combination with molnupiravir, reduced the viral load by 7- and 58-fold, respectively. Considering the clinical safety, oral bioavailability, and tolerability of izumerogant in a recent Phase I study, izumerogant-like drugs represent potent dual-targeting antiviral HDAs with pronounced broad-spectrum activity for further clinical development.

Influence of AAV vector tropism on long-term expression and Fc-γ receptor binding of an antibody targeting SARS-CoV-2.

Long-acting passive immunization strategies are needed to protect immunosuppressed vulnerable groups from infectious diseases. To further explore this concept for COVID-19, we constructed Adeno-associated viral (AAV) vectors encoding the human variable regions of the SARS-CoV-2 neutralizing antibody, TRES6, fused to murine constant regions. An optimized vector construct was packaged in hepatotropic (AAV8) or myotropic (AAVMYO) AAV capsids and injected intravenously into syngeneic TRIANNI-mice. The highest TRES6 serum concentrations (511 µg/ml) were detected 24 weeks after injection of the myotropic vector particles and mean TRES6 serum concentrations remained above 100 µg/ml for at least one year. Anti-drug antibodies or TRES6-specific T cells were not detectable. After injection of the AAV8 particles, vector mRNA was detected in the liver, while the AAVMYO particles led to high vector mRNA levels in the heart and skeletal muscle. The analysis of the Fc-glycosylation pattern of the TRES6 serum antibodies revealed critical differences between the capsids that coincided with different binding activities to murine Fc-γ-receptors. Concomitantly, the vector-based immune prophylaxis led to protection against SARS-CoV-2 infection in K18-hACE2 mice. High and long-lasting expression levels, absence of anti-drug antibodies and favourable Fc-γ-receptor binding activities warrant further exploration of myotropic AAV vector-based delivery of antibodies and other biologicals.

Genetic barrier to resistance: a critical parameter for efficacy of neutralizing monoclonal antibodies against SARS-CoV-2 in a nonhuman primate model.

The potency of antibody neutralization in cell culture has been used as the key criterion for selection of antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) for clinical development. As other aspects may also influence the degree of protection in vivo, we compared the efficacy of two neutralizing monoclonal antibodies (TRES6 and 4C12) targeting different epitopes of the receptor binding domain (RBD) of SARS-CoV-2 in a prophylactic setting in rhesus monkeys. All four animals treated with TRES6 had reduced viral loads in the upper respiratory tract 2 days after naso-oropharyngeal challenge with the Alpha SARS-CoV-2 variant. Starting 2 days after challenge, mutations conferring resistance to TRES6 were dominant in two of the rhesus monkeys, with both animals failing to maintain reduced viral loads. Consistent with its lower serum neutralization titer at the day of challenge, prophylaxis with 4C12 tended to suppress viral load at day 2 less efficiently than TRES6. However, a week after challenge, mean viral loads in the lower respiratory tract in 4C12-treated animals were lower than in the TRES6 group and no mutations conferring resistance to 4C12 could be detected in viral isolates from nasal or throat swabs. Thus, genetic barrier to resistance seems to be a critical parameter for the efficacy of prophylaxis with monoclonal antibodies against SARS-CoV-2. Furthermore, comparison of antibody concentrations in respiratory secretions to those in serum shows reduced distribution of the 4C12 antibody into respiratory secretions and a delay in the appearance of antibodies in bronchoalveolar lavage fluid compared to their appearance in secretions of the upper respiratory tract.IMPORTANCEMonoclonal antibodies are a powerful tool for the prophylaxis and treatment of acute viral infections. Hence, they were one of the first therapeutic agents licensed for the treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Oftentimes, the main criterion for the selection of antibodies for clinical development is their potency of neutralization in cell culture. By comparing two antibodies targeting the Spike protein of SARS-CoV-2, we now observed that the antibody that neutralized SARS-CoV-2 more efficiently in cell culture suppressed viral load in challenged rhesus monkeys to a lesser extent. Extraordinary rapid emergence of mutants of the challenge virus, which had lost their sensitivity to the antibody, was identified as the major reason for the reduced efficacy of the antibody in rhesus monkeys. Therefore, the viral genetic barrier to resistance to antibodies also affects their efficacy.

Synthesis and Characterization of DHODH Inhibitors Based on the Vidofludimus Scaffold with Pronounced Anti-SARS-CoV-2 Activity.

New strategies for the rapid development of broad-spectrum antiviral therapies are urgently required for emerging and re-emerging viruses like the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Host-directed antivirals that target universal cellular metabolic pathways necessary for viral replication present a promising approach with broad-spectrum activity and low potential for development of viral resistance. Dihydroorotate dehydrogenase (DHODH) was identified as one of those universal host factors essential for the replication of many clinically relevant human pathogenic viruses. DHODH is the rate-limiting enzyme catalyzing the fourth step in the de novo pyrimidine synthesis. Therefore, it is also developed as a therapeutic target for many diseases relying on cellular pyrimidine resources, such as cancer, autoimmune diseases and viral or bacterial infection. Thus, several DHODH inhibitors, including vidofludimus calcium (VidoCa, IMU-838), are currently in development or have been investigated in clinical trials for the treatment of virus infections such as SARS-CoV-2-mediated coronavirus disease 19 (COVID-19). Here, we report the medicinal chemistry optimization of VidoCa that resulted in metabolically more stable derivatives with improved DHODH target inhibition in various mammalian species, which translated into improved efficacy against SARS-CoV-2.

Reading tea leaves worldwide: Decoupled drivers of initial litter decomposition mass-loss rate and stabilization.

The breakdown of plant material fuels soil functioning and biodiversity. Currently, process understanding of global decomposition patterns and the drivers of such patterns are hampered by the lack of coherent large-scale datasets. We buried 36,000 individual litterbags (tea bags) worldwide and found an overall negative correlation between initial mass-loss rates and stabilization factors of plant-derived carbon, using the Tea Bag Index (TBI). The stabilization factor quantifies the degree to which easy-to-degrade components accumulate during early-stage decomposition (e.g. by environmental limitations). However, agriculture and an interaction between moisture and temperature led to a decoupling between initial mass-loss rates and stabilization, notably in colder locations. Using TBI improved mass-loss estimates of natural litter compared to models that ignored stabilization. Ignoring the transformation of dead plant material to more recalcitrant substances during early-stage decomposition, and the environmental control of this transformation, could overestimate carbon losses during early decomposition in carbon cycle models.

TheraVision: Engineering platform technology for the development of oncolytic viruses based on herpes simplex virus type 1.

Viruses are able to efficiently penetrate cells, multiply, and eventually kill infected cells, release tumor antigens, and activate the immune system. Therefore, viruses are highly attractive novel agents for cancer therapy. Clinical trials with first generations of oncolytic viruses (OVs) are very promising but show significant need for optimization. The aim of TheraVision was to establish a broadly applicable engineering platform technology for combinatorial oncolytic virus and immunotherapy. Through genetic engineering, an attenuated herpes simplex virus type 1 (HSV1) was generated that showed increased safety compared to the wild-type strain. To demonstrate the modularity and the facilitated generation of new OVs, two transgenes encoding retargeting as well as immunomodulating single-chain variable fragments (scFvs) were integrated into the platform vector. The resulting virus selectively infected epidermal growth factor receptor (EGFR)-expressing cells and produced a functional immune checkpoint inhibitor against programmed cell death protein 1 (PD-1). Thus, both viral-mediated oncolysis and immune-cell-mediated therapy were combined into a single viral vector. Safety and functionality of the armed OVs have been shown in novel preclinical models ranging from patient-derived organoids and tissue-engineered human in vitro 3D tumor models to complex humanized mouse models. Consequently, a novel and proprietary engineering platform vector based on HSV1 is available for the facilitated preclinical development of oncolytic virotherapy.

Mucosal immunization with a low-energy electron inactivated respiratory syncytial virus vaccine protects mice without Th2 immune bias.

The respiratory syncytial virus (RSV) is a leading cause of acute lower respiratory tract infections associated with numerous hospitalizations. Recently, intramuscular (i.m.) vaccines against RSV have been approved for elderly and pregnant women. Noninvasive mucosal vaccination, e.g., by inhalation, offers an alternative against respiratory pathogens like RSV. Effective mucosal vaccines induce local immune responses, potentially resulting in the efficient and fast elimination of respiratory viruses after natural infection. To investigate this immune response to an RSV challenge, low-energy electron inactivated RSV (LEEI-RSV) was formulated with phosphatidylcholine-liposomes (PC-LEEI-RSV) or 1,2-dioleoyl-3-trimethylammonium-propane and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DD-LEEI-RSV) for vaccination of mice intranasally. As controls, LEEI-RSV and formalin-inactivated-RSV (FI-RSV) were used via i.m. vaccination. The RSV-specific immunogenicity of the different vaccines and their protective efficacy were analyzed. RSV-specific IgA antibodies and a statistically significant reduction in viral load upon challenge were detected in mucosal DD-LEEI-RSV-vaccinated animals. Alhydrogel-adjuvanted LEEI-RSV i.m. showed a Th2-bias with enhanced IgE, eosinophils, and lung histopathology comparable to FI-RSV. These effects were absent when applying the mucosal vaccines highlighting the potential of DD-LEEI-RSV as an RSV vaccine candidate and the improved performance of this mucosal vaccine candidate.

Outcome of Epilepsy Surgery in MRI-Negative Patients Without Histopathologic Abnormalities in the Resected Tissue.

BACKGROUND AND OBJECTIVE: Patients with presumed nonlesional focal epilepsy-based on either MRI or histopathologic findings-have a lower success rate of epilepsy surgery compared with lesional patients. In this study, we aimed to characterize a large group of patients with focal epilepsy who underwent epilepsy surgery despite a normal MRI and had no lesion on histopathology. Determinants of their postoperative seizure outcomes were further studied. METHODS: We designed an observational multicenter cohort study of MRI-negative and histopathology-negative patients who were derived from the European Epilepsy Brain Bank and underwent epilepsy surgery between 2000 and 2012 in 34 epilepsy surgery centers within Europe. We collected data on clinical characteristics, presurgical assessment, including genetic testing, surgery characteristics, postoperative outcome, and treatment regimen. RESULTS: Of the 217 included patients, 40% were seizure-free (Engel I) 2 years after surgery and one-third of patients remained seizure-free after 5 years. Temporal lobe surgery (adjusted odds ratio [AOR]: 2.62; 95% CI 1.19-5.76), shorter epilepsy duration (AOR for duration: 0.94; 95% CI 0.89-0.99), and completely normal histopathologic findings-versus nonspecific reactive gliosis-(AOR: 4.69; 95% CI 1.79-11.27) were significantly associated with favorable seizure outcome at 2 years after surgery. Of patients who underwent invasive monitoring, only 35% reached seizure freedom at 2 years. Patients with parietal lobe resections had lowest seizure freedom rates (12.5%). Among temporal lobe surgery patients, there was a trend toward favorable outcome if hippocampectomy was part of the resection strategy (OR: 2.94; 95% CI 0.98-8.80). Genetic testing was only sporadically performed. DISCUSSION: This study shows that seizure freedom can be reached in 40% of nonlesional patients with both normal MRI and histopathology findings. In particular, nonlesional temporal lobe epilepsy should be regarded as a relatively favorable group, with almost half of patients achieving seizure freedom at 2 years after surgery-even more if the hippocampus is resected-compared with only 1 in 5 nonlesional patients who underwent extratemporal surgery. Patients with an electroclinically identified focus, who are nonlesional, will be a promising group for advanced molecular-genetic analysis of brain tissue specimens to identify new brain somatic epilepsy genes or epilepsy-associated molecular pathways.

Immunological Quantitation of the Glycation Site Lysine-414 in Serum Albumin in Human Plasma Samples by Indirect ELISA Using Highly Specific Monoclonal Antibodies.

Diabetes mellitus, a metabolic disorder that is characterized by elevated blood glucose levels, is common throughout the world and its prevalence is steadily increasing. Early diagnosis and treatment are important to prevent acute complications and life-threatening long-term organ damage. Glycation sites in human serum albumin (HSA) are considered to be promising biomarkers of systemic glycemic status. This work aimed to develop a sensitive and clinically applicable ELISA for the quantification of glycation site Lys414 in HSA (HSAK414 ). The monoclonal antibodies (mAbs) were generated by immunizing mice with a glycated peptide. The established indirect ELISA based on mAb 50D8 (IgG1 isotype) yielded a limit of detection of 0.39 nmol/g HSA for HSAK414 with a linear dynamic range from 0.50 to 6.25 nmol/g glycated HSA. The inter- and intra-day assays with coefficients of variation less than 20 % indicated good assay performance and precision. Assay evaluation was based on plasma samples from diabetic and non-diabetic subjects with known HSAK414 glycation levels previously determined by LC-MS. Both data sets correlated very well. In conclusion, the generated mAb 50D8 and the established ELISA could be a valuable tool for the rapid quantitation of glycation site HSAK414 in plasma samples to evaluate its clinical relevance.

CD44v6 specific CAR-NK cells for targeted immunotherapy of head and neck squamous cell carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is a major challenge for current therapies. CAR-T cells have shown promising results in blood cancers, however, their effectiveness against solid tumors remains a hurdle. Recently, CD44v6-directed CAR-T cells demonstrated efficacy in controlling tumor growth in multiple myeloma and solid tumors such as HNSCC, lung and ovarian adenocarcinomas. Apart from CAR-T cells, CAR-NK cells offer a safe and allogenic alternative to autologous CAR-T cell therapy. In this paper, we investigated the capacity of CAR-NK cells redirected against CD44v6 to execute cytotoxicity against HNSCC. Anti-CD44v6 CAR-NK cells were generated from healthy donor peripheral blood-derived NK cells using gamma retroviral vectors (gRVs). The NK cell transduction was optimized by exploring virus envelope proteins derived from the baboon endogenous virus envelope (BaEV), feline leukemia virus (FeLV, termed RD114-TR) and gibbon ape leukemia virus (GaLV), respectively. BaEV pseudotyped gRVs induced the highest transduction rate compared to RD114-TR and GaLV envelopes as measured by EGFP and surface CAR expression of transduced NK cells. CAR-NK cells showed a two- to threefold increase in killing efficacy against various HNSCC cell lines compared to unmodified, cytokine-expanded primary NK cells. Anti-CD44v6 CAR-NK cells were effective in eliminating tumor cell lines with high and low CD44v6 expression levels. Overall, the improved cytotoxicity of CAR-NK cells holds promise for a therapeutic option for the treatment of HNSCC. However, further preclinical trials are necessary to test in vivo efficacy and safety, as well to optimize the treatment regimen of anti-CD44v6 CAR-NK cells against solid tumors.

Immunization with different recombinant West Nile virus envelope proteins induces varying levels of serological cross-reactivity and protection from infection.

Introduction: West Nile Virus (WNV) is a zoonotic flavivirus transmitted by mosquitoes. Especially in the elderly or in immunocompromised individuals an infection with WNV can lead to severe neurological symptoms. To date, no human vaccine against WNV is available. The Envelope (E) protein, located at the surface of flaviviruses, is involved in the invasion into host cells and is the major target for neutralizing antibodies and therefore central to vaccine development. Due to their close genetic and structural relationship, flaviviruses share highly conserved epitopes, such as the fusion loop domain (FL) in the E protein, that are recognized by cross-reactive antibodies. These antibodies can lead to enhancement of infection with heterologous flaviviruses, which is a major concern for potential vaccines in areas with co-circulation of different flaviviruses, e.g. Dengue or Zika viruses. Material: To reduce the potential of inducing cross-reactive antibodies, we performed an immunization study in mice using WNV E proteins with either wild type sequence or a mutated FL, and WNV E domain III which does not contain the FL at all. Results and discussion: Our data show that all antigens induce high levels of WNV-binding antibodies. However, the level of protection against WNV varied, with the wildtype E protein inducing full, the other antigens only partial protection. On the other hand, serological cross-reactivity to heterologous flaviviruses was significantly reduced after immunization with the mutated E protein or domain III as compared to the wild type version. These results have indications for choosing antigens with the optimal specificity and efficacy in WNV vaccine development.

Drug repurposing for the treatment of COVID-19: Targeting nafamostat to the lungs by a liposomal delivery system.

Despite tremendous global efforts since the beginning of the COVID-19 pandemic, still only a limited number of prophylactic and therapeutic options are available. Although vaccination is the most effective measure in preventing morbidity and mortality, there is a need for safe and effective post-infection treatment medication. In this study, we explored a pipeline of 21 potential candidates, examined in the Calu-3 cell line for their antiviral efficacy, for drug repurposing. Ralimetinib and nafamostat, clinically used drugs, have emerged as attractive candidates. Due to the inherent limitations of the selected drugs, we formulated targeted liposomes suitable for both systemic and intranasal administration. Non-targeted and targeted nafamostat liposomes (LipNaf) decorated with an Apolipoprotein B peptide (ApoB-P) as a specific lung-targeting ligand were successfully developed. The developed liposomal formulations of nafamostat were found to possess favorable physicochemical properties including nano size (119-147 nm), long-term stability of the normally rapidly degrading compound in aqueous solution, negligible leakage from the liposomes upon storage, and a neutral surface charge with low polydispersity index (PDI). Both nafamostat and ralimetinib liposomes showed good cellular uptake and lack of cytotoxicity, and non-targeted LipNaf demonstrated enhanced accumulation in the lungs following intranasal (IN) administration in non-infected mice. LipNaf retained its anti-SARS-CoV 2 activity in Calu 3 cells with only a modest decrease, exhibiting complete inhibition at concentrations >100 nM. IN, but not intraperitoneal (IP) treatment with targeted LipNaf resulted in a trend to reduced viral load in the lungs of K18-hACE2 mice compared to targeted empty Lip. Nevertheless, upon removal of outlier data, a statistically significant 1.9-fold reduction in viral load was achieved. This observation further highlights the importance of a targeted delivery into the respiratory tract. In summary, we were able to demonstrate a proof-of-concept of drug repurposing by liposomal formulations with anti-SARS-CoV-2 activity. The biodistribution and bioactivity studies with LipNaf suggest an IN or inhalation route of administration for optimal therapeutic efficacy.

Mucosal Application of a Low-Energy Electron Inactivated Respiratory Syncytial Virus Vaccine Shows Protective Efficacy in an Animal Model.

Respiratory syncytial virus (RSV) is a leading cause of acute lower respiratory tract infections in the elderly and in children, associated with pediatric hospitalizations. Recently, first vaccines have been approved for people over 60 years of age applied by intramuscular injection. However, a vaccination route via mucosal application holds great potential in the protection against respiratory pathogens like RSV. Mucosal vaccines induce local immune responses, resulting in a fast and efficient elimination of respiratory viruses after natural infection. Therefore, a low-energy electron irradiated RSV (LEEI-RSV) formulated with phosphatidylcholine-liposomes (PC-LEEI-RSV) was tested ex vivo in precision cut lung slices (PCLSs) for adverse effects. The immunogenicity and protective efficacy in vivo were analyzed in an RSV challenge model after intranasal vaccination using a homologous prime-boost immunization regimen. No side effects of PC-LEEI-RSV in PCLS and an efficient antibody induction in vivo could be observed. In contrast to unformulated LEEI-RSV, the mucosal vaccination of mice with PC formulated LEEI-RSV showed a statistically significant reduction in viral load after challenge. These results are a proof-of-principle for the use of LEEI-inactivated viruses formulated with liposomes to be administered intranasally to induce a mucosal immunity that could also be adapted for other respiratory viruses.

Detection and attribution of an anomaly in terrestrial photosynthesis in Europe during the COVID-19 lockdown.

Carbon dioxide (CO2) uptake by plant photosynthesis, referred to as gross primary production (GPP) at the ecosystem level, is sensitive to environmental factors, including pollutant exposure, pollutant uptake, and changes in the scattering of solar shortwave irradiance (SWin) - the energy source for photosynthesis. The 2020 spring lockdown due to COVID-19 resulted in improved air quality and atmospheric transparency, providing a unique opportunity to assess the impact of air pollutants on terrestrial ecosystem functioning. However, detecting these effects can be challenging as GPP is influenced by other meteorological drivers and management practices. Based on data collected from 44 European ecosystem-scale CO2 flux monitoring stations, we observed significant changes in spring GPP at 34 sites during 2020 compared to 2015-2019. Among these, 14 sites showed an increase in GPP associated with higher SWin, 10 sites had lower GPP linked to atmospheric and soil dryness, and seven sites were subjected to management practices. The remaining three sites exhibited varying dynamics, with one experiencing colder and rainier weather resulting in lower GPP, and two showing higher GPP associated with earlier spring melts. Analysis using the regional atmospheric chemical transport model (LOTOS-EUROS) indicated that the ozone (O3) concentration remained relatively unchanged at the research sites, making it unlikely that O3 exposure was the dominant factor driving the primary production anomaly. In contrast, SWin increased by 9.4 % at 36 sites, suggesting enhanced GPP possibly due to reduced aerosol optical depth and cloudiness. Our findings indicate that air pollution and cloudiness may weaken the terrestrial carbon sink by up to 16 %. Accurate and continuous ground-based observations are crucial for detecting and attributing subtle changes in terrestrial ecosystem functioning in response to environmental and anthropogenic drivers.

Identification and evaluation of a novel tribenzamide derivative as an inhibitor targeting the entry of the respiratory syncytial virus.

Human respiratory syncytial virus (RSV) is the leading cause of severe lower respiratory tract infections in infants, the elderly, and the immunocompromised, yet no licensed vaccine and only limited therapeutic options for prevention and treatment are available, which poses a global health challenge and emphasizes the urgent medical need for novel antiviral agents. In the current study, a novel potent small molecule inhibitor of RSV was identified by performing a screening and structure optimization campaign, wherein a naturally occurring dicaffeoylquinic acid (DCQA) compound served as a chemical starting point. The reported benzamide derivative inhibitor, designated as 2f, was selected for its improved stability and potent antiviral activity from a series of investigated structurally related compounds. 2f was well tolerated by cells and able to inhibit RSV infection with a half maximal inhibitory concentration (IC50) of 35 nM and a favorable selectivity index (SI) of 3742. Although the exact molecular target for 2f is not known, in vitro mechanism of action investigations revealed that the compound inhibits the early stage of infection by interacting with RSV virion and interferes primarily with the attachment and potentially with the virus-cell fusion step. Moreover, intranasal administration of 2f to mice simultaneously or prior to intranasal infection with RSV significantly decreased viral load in the lungs, pointing to the in vivo potential of the compound. Our results suggest that 2f is a viable candidate for further preclinical development and evaluation as an antiviral agent against RSV infections.

Joint optimization of land carbon uptake and albedo can help achieve moderate instantaneous and long-term cooling effects.

Both carbon dioxide uptake and albedo of the land surface affect global climate. However, climate change mitigation by increasing carbon uptake can cause a warming trade-off by decreasing albedo, with most research focusing on afforestation and its interaction with snow. Here, we present carbon uptake and albedo observations from 176 globally distributed flux stations. We demonstrate a gradual decline in maximum achievable annual albedo as carbon uptake increases, even within subgroups of non-forest and snow-free ecosystems. Based on a paired-site permutation approach, we quantify the likely impact of land use on carbon uptake and albedo. Shifting to the maximum attainable carbon uptake at each site would likely cause moderate net global warming for the first approximately 20 years, followed by a strong cooling effect. A balanced policy co-optimizing carbon uptake and albedo is possible that avoids warming on any timescale, but results in a weaker long-term cooling effect.

Advanced Molecular Tweezers with Lipid Anchors against SARS-CoV-2 and Other Respiratory Viruses.

The COVID-19 pandemic caused by SARS-CoV-2 presents a global health emergency. Therapeutic options against SARS-CoV-2 are still very limited but urgently required. Molecular tweezers are supramolecular agents that destabilize the envelope of viruses resulting in a loss of viral infectivity. Here, we show that first-generation tweezers, CLR01 and CLR05, disrupt the SARS-CoV-2 envelope and abrogate viral infectivity. To increase the antiviral activity, a series of 34 advanced molecular tweezers were synthesized by insertion of aliphatic or aromatic ester groups on the phosphate moieties of the parent molecule CLR01. A structure-activity relationship study enabled the identification of tweezers with a markedly enhanced ability to destroy lipid bilayers and to suppress SARS-CoV-2 infection. Selected tweezer derivatives retain activity in airway mucus and inactivate the SARS-CoV-2 wildtype and variants of concern as well as respiratory syncytial, influenza, and measles viruses. Moreover, inhibitory activity of advanced tweezers against respiratory syncytial virus and SARS-CoV-2 was confirmed in mice. Thus, potentiated tweezers are broad-spectrum antiviral agents with great prospects for clinical development to combat highly pathogenic viruses.

Macromolecular Viral Entry Inhibitors as Broad-Spectrum First-Line Antivirals with Activity against SARS-CoV-2.

Inhibitors of viral cell entry based on poly(styrene sulfonate) and its core-shell nanoformulations based on gold nanoparticles are investigated against a panel of viruses, including clinical isolates of SARS-CoV-2. Macromolecular inhibitors are shown to exhibit the highly sought-after broad-spectrum antiviral activity, which covers most analyzed enveloped viruses and all of the variants of concern for SARS-CoV-2 tested. The inhibitory activity is quantified in vitro in appropriate cell culture models and for respiratory viral pathogens (respiratory syncytial virus and SARS-CoV-2) in mice. Results of this study comprise a significant step along the translational path of macromolecular inhibitors of virus cell entry, specifically against enveloped respiratory viruses.

Inhibitors of dihydroorotate dehydrogenase cooperate with molnupiravir and N4-hydroxycytidine to suppress SARS-CoV-2 replication.

The nucleoside analog N4-hydroxycytidine (NHC) is the active metabolite of the prodrug molnupiravir, which has been approved for the treatment of COVID-19. SARS-CoV-2 incorporates NHC into its RNA, resulting in defective virus genomes. Likewise, inhibitors of dihydroorotate dehydrogenase (DHODH) reduce virus yield upon infection, by suppressing the cellular synthesis of pyrimidines. Here, we show that NHC and DHODH inhibitors strongly synergize in the inhibition of SARS-CoV-2 replication in vitro. We propose that the lack of available pyrimidine nucleotides upon DHODH inhibition increases the incorporation of NHC into nascent viral RNA. This concept is supported by the rescue of virus replication upon addition of pyrimidine nucleosides to the media. DHODH inhibitors increased the antiviral efficiency of molnupiravir not only in organoids of human lung, but also in Syrian Gold hamsters and in K18-hACE2 mice. Combining molnupiravir with DHODH inhibitors may thus improve available therapy options for COVID-19.

Low-Energy Electron Irradiation of Tick-Borne Encephalitis Virus Provides a Protective Inactivated Vaccine.

Tick-borne encephalitis virus (TBEV) is a zoonotic flavivirus which is endemic in many European and Asian countries. Humans can get infected with TBEV usually via ticks, and possible symptoms of the infection range from fever to severe neurological complications such as encephalitis. Vaccines to protect against TBEV-induced disease are widely used and most of them consist of whole viruses, which are inactivated by formaldehyde. Although this production process is well established, it has several drawbacks, including the usage of hazardous chemicals, the long inactivation times required and the potential modification of antigens by formaldehyde. As an alternative to chemical treatment, low-energy electron irradiation (LEEI) is known to efficiently inactivate pathogens by predominantly damaging nucleic acids. In contrast to other methods of ionizing radiation, LEEI does not require substantial shielding constructions and can be used in standard laboratories. Here, we have analyzed the potential of LEEI to generate a TBEV vaccine and immunized mice with three doses of irradiated or chemically inactivated TBEV. LEEI-inactivated TBEV induced binding antibodies of higher titer compared to the formaldehyde-inactivated virus. This was also observed for the avidity of the antibodies measured after the second dose. After viral challenge, the mice immunized with LEEI- or formaldehyde-inactivated TBEV were completely protected from disease and had no detectable virus in the central nervous system. Taken together, the results indicate that LEEI could be an alternative to chemical inactivation for the production of a TBEV vaccine.