Pseudomonas aeruginosa survives in epithelia by ExoS-mediated inhibition of autophagy and mTOR.

One major factor that contributes to the virulence of Pseudomonas aeruginosa is its ability to reside and replicate unchallenged inside airway epithelial cells. The mechanism by which P. aeruginosa escapes destruction by intracellular host defense mechanisms, such as autophagy, is not known. Here, we show that the type III secretion system effector protein ExoS facilitates P. aeruginosa survival in airway epithelial cells by inhibiting autophagy in host cells. Autophagy inhibition is independent of mTOR activity, as the latter is also inhibited by ExoS, albeit by a different mechanism. Deficiency of the critical autophagy gene Atg7 in airway epithelial cells, both in vitro and in mouse models, greatly enhances the survival of ExoS-deficient P. aeruginosa but does not affect the survival of ExoS-containing bacteria. The inhibitory effect of ExoS on autophagy and mTOR depends on the activity of its ADP-ribosyltransferase domain. Inhibition of mTOR is caused by ExoS-mediated ADP ribosylation of RAS, whereas autophagy inhibition is due to the suppression of autophagic Vps34 kinase activity.

A Recombinant Respiratory Syncytial Virus Vaccine Candidate Attenuated by a Low-Fusion F Protein Is Immunogenic and Protective against Challenge in Cotton Rats.

UNLABELLED: Although respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infections in infants, a safe and effective vaccine is not yet available. Live-attenuated vaccines (LAVs) are the most advanced vaccine candidates in RSV-naive infants. However, designing an LAV with appropriate attenuation yet sufficient immunogenicity has proven challenging. In this study, we implemented reverse genetics to address these obstacles with a multifaceted LAV design that combined the codon deoptimization of genes for nonstructural proteins NS1 and NS2 (dNS), deletion of the small hydrophobic protein (ΔSH) gene, and replacement of the wild-type fusion (F) protein gene with a low-fusion RSV subgroup B F consensus sequence of the Buenos Aires clade (BAF). This vaccine candidate, RSV-A2-dNS-ΔSH-BAF (DB1), was attenuated in two models of primary human airway epithelial cells and in the upper and lower airways of cotton rats. DB1 was also highly immunogenic in cotton rats and elicited broadly neutralizing antibodies against a diverse panel of recombinant RSV strains. When vaccinated cotton rats were challenged with wild-type RSV A, DB1 reduced viral titers in the upper and lower airways by 3.8 log10 total PFU and 2.7 log10 PFU/g of tissue, respectively, compared to those in unvaccinated animals (P < 0.0001). DB1 was thus attenuated, highly immunogenic, and protective against RSV challenge in cotton rats. DB1 is the first RSV LAV to incorporate a low-fusion F protein as a strategy to attenuate viral replication and preserve immunogenicity. IMPORTANCE: RSV is a leading cause of infant hospitalizations and deaths. The development of an effective vaccine for this high-risk population is therefore a public health priority. Although live-attenuated vaccines have been safely administered to RSV-naive infants, strategies to balance vaccine attenuation with immunogenicity have been elusive. In this study, we introduced a novel strategy to attenuate a recombinant RSV vaccine by incorporating a low-fusion, subgroup B F protein in the genetic background of codon-deoptimized nonstructural protein genes and a deleted small hydrophobic protein gene. The resultant vaccine candidate, DB1, was attenuated, highly immunogenic, and protective against RSV challenge in cotton rats.

Glycan variants of a respiratory syncytial virus antibody with enhanced effector function and in vivo efficacy.

Respiratory syncytial virus (RSV) can cause devastating lower respiratory tract infections in preterm infants or when other serious health problems are present. Immunoprophylaxis with palivizumab (Synagis), a humanized IgG1 mAb, is the current standard of care for preventing RSV infection in at-risk neonates. We have explored the contribution of effector function to palivizumab efficacy using a plant-based expression system to produce palivizumab N-glycan structure variants with high homogeneity on different antibody isotypes. We compared these isotype and N-glycoform variants with commercially available palivizumab with respect to both in vitro receptor and C1q binding and in vivo efficacy. Whereas the affinity for antigen and neutralization activity of each variant were indistinguishable from those of palivizumab, their Fcγ receptor binding profiles were very different, which was reflected in either a reduced or enhanced ability to influence the RSV lung titer in challenged cotton rats. Enhanced Fcγ receptor binding was associated with reduced viral lung titers compared with palivizumab, whereas abrogation of receptor binding led to a drastic reduction in efficacy. The results support the hypotheses that classic antibody neutralization is a minor component of efficacy by palivizumab in the cotton rat and that antibody-dependent cell-mediated cytotoxicity activity can significantly enhance the efficacy of this antiviral mAb.

Generation and Characterization of ALX-0171, a Potent Novel Therapeutic Nanobody for the Treatment of Respiratory Syncytial Virus Infection.

Respiratory syncytial virus (RSV) is an important causative agent of lower respiratory tract infections in infants and elderly individuals. Its fusion (F) protein is critical for virus infection. It is targeted by several investigational antivirals and by palivizumab, a humanized monoclonal antibody used prophylactically in infants considered at high risk of severe RSV disease. ALX-0171 is a trimeric Nanobody that binds the antigenic site II of RSV F protein with subnanomolar affinity. ALX-0171 demonstrated in vitro neutralization superior to that of palivizumab against prototypic RSV subtype A and B strains. Moreover, ALX-0171 completely blocked replication to below the limit of detection for 87% of the viruses tested, whereas palivizumab did so for 18% of the viruses tested at a fixed concentration. Importantly, ALX-0171 was highly effective in reducing both nasal and lung RSV titers when delivered prophylactically or therapeutically directly to the lungs of cotton rats. ALX-0171 represents a potent novel antiviral compound with significant potential to treat RSV-mediated disease.

GS-5806 Inhibits a Broad Range of Respiratory Syncytial Virus Clinical Isolates by Blocking the Virus-Cell Fusion Process.

Respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract infections in infants and young children. In addition, RSV causes significant morbidity and mortality in hospitalized elderly and immunocompromised patients. Currently, only palivizumab, a monoclonal antibody against the RSV fusion (F) protein, and inhaled ribavirin are approved for the prophylactic and therapeutic treatment of RSV, respectively. Therefore, there is a clinical need for safe and effective therapeutic agents for RSV infections. GS-5806, discovered via chemical optimization of a hit from a high-throughput antiviral-screening campaign, selectively inhibits a diverse set of 75 RSV subtype A and B clinical isolates (mean 50% effective concentration [EC50] = 0.43 nM). The compound maintained potency in primary human airway epithelial cells and exhibited low cytotoxicity in human cell lines and primary cell cultures (selectivity > 23,000-fold). Time-of-addition and temperature shift studies demonstrated that GS-5806 does not block RSV attachment to cells but interferes with virus entry. Follow-up experiments showed potent inhibition of RSV F-mediated cell-to-cell fusion. RSV A and B variants resistant to GS-5806, due to mutations in F protein (RSV A, L138F or F140L/N517I, and RSV B, F488L or F488S), were isolated and showed cross-resistance to other RSV fusion inhibitors, such as VP-14637, but remained fully sensitive to palivizumab and ribavirin. In summary, GS-5806 is a potent and selective RSV fusion inhibitor with antiviral activity against a diverse set of RSV clinical isolates. The compound is currently under clinical investigation for the treatment of RSV infection in pediatric, immunocompromised, and elderly patients.

Development of an adenovirus-based respiratory syncytial virus vaccine: preclinical evaluation of efficacy, immunogenicity, and enhanced disease in a cotton rat model.

UNLABELLED: The lack of a vaccine against respiratory syncytial virus (RSV) is a challenging and serious gap in preventive medicine. Herein, we characterize the immunogenicity of an adenovirus serotype 5-based RSV vaccine encoding the fusion (F) protein (Ad5.RSV-F) and the protection provided following immunization with Ad5.RSV-F and assess its potential for producing enhanced disease in a cotton rat (CR) model. Animals were immunized intranasally (i.n.) and/or intramuscularly (i.m.) and subsequently challenged with RSV/A/Tracy (i.n.) to assess protection. Robust immune responses were seen in CRs vaccinated with Ad5.RSV-F given i.m. or i.n., and these responses correlated with reduced replication of the virus in noses and lungs after challenge. Neutralizing antibody responses following immunization with a single dose of Ad5.RSV-F at 1 × 10(11) viral particles (v.p.) elicited antibody titers 64- to 256-fold greater than those seen after natural infection. CRs boosted with Ad5.RSV-F i.n. 28 days after an i.m. dose also had significant increases in neutralizing antibody titers. Antibody affinity for different F-protein antigenic sites revealed substantial differences between antibodies elicited by Ad5.RSV-F and those seen after RSV infection; differences in antibody profiles were also seen between CRs given Ad5.RSV-F i.m. and CRs given Ad5.RSV-F i.n. Ad5.RSV-F priming did not result in enhanced disease following live-virus challenge, in contrast to the histopathology seen in CRs given the formalin-inactivated RSV/A/Burnett vaccine. IMPORTANCE: Respiratory syncytial virus (RSV) is the most common cause of acute lower respiratory infection in infants and young children and a serious health threat in the immunocompromised and the elderly. Infection severity increased in children in an immunization trial, hampering the over 4-decade-long quest for a successful RSV vaccine. In this study, we show that a genetically engineered RSV-F-encoding adenoviral vector provides protective immunity against RSV challenge without enhanced lung disease in cotton rats (CRs). CRs were vaccinated under a number of different regimens, and the immunity induced by the recombinant adenoviral RSV vaccine administered by use of an intramuscular prime-intranasal boost regimen may provide the best protection for young infants and children at risk of RSV infection, since this population is naive to adenoviral preformed immunity. Overall, this report describes a potential RSV vaccine candidate that merits further evaluation in a phase I clinical study in humans.

WFDC1 is a key modulator of inflammatory and wound repair responses.

WFDC1/ps20 is a whey acidic protein four-disulfide core member that exhibits diverse growth and immune-associated functions in vitro. In vivo functions are unknown, although WFDC1 is lower in reactive stroma. A Wfdc1-null mouse was generated to assess core functions. Wfdc1-null mice exhibited normal developmental and adult phenotypes. However, homeostasis challenges affected inflammatory and repair processes. Wfdc1-null mice infected with influenza A exhibited 2.75-log-fold lower viral titer relative to control mice. Wfdc1-null infected lungs exhibited elevated macrophages and deposition of osteopontin, a potent macrophage chemokine. In wounding studies, Wfdc1-null mice exhibited an elevated rate of skin closure, and this too was associated with elevated deposition of osteopontin and macrophage recruitment. Wfdc1-null fibroblasts exhibited impaired spheroid formation, elevated adhesion to fibronectin, and an increased rate of wound closure in vitro. This was reversed by neutralizing antibody to osteopontin. Osteopontin mRNA and cleaved protein was up-regulated in Wfdc1-null cells treated with lipopolysaccharide or polyinosinic-polycytidylic acid coordinate with constitutively active matrix metallopeptidase-9 (MMP-9), a protease that cleaves osteopontin. These data suggest that WFDC1/ps20 modulates core host response mechanisms, in part, via regulation of osteopontin and MMP-9 activity. Release from WFDC1 regulation is likely a key component of inflammatory and repair response mechanisms, and involves the processing of elevated osteopontin by activated MMP-9, and subsequent macrophage recruitment.

In-vivo and human evidence for potential efficacy of therapeutic polyclonal RSV neutralizing antibodies for palivizumab-resistant RSV infections.

BACKGROUND: Monoclonal antibody (palivizumab), intravenous immune globulin (IGIV), or respiratory syncytial virus (RSV)-polyclonal-hyperimmune-globulin (RSV-IG as Respigam®, RI-001, RI-002) are used with ribavirin in RSV-infected immunocompromised patients, with debated efficacy. Palivizumab-resistance (PR) can arise during treatment of persistent infections in this population. RSV-IG may confer benefit in PR-RSV infection. METHODS: RSV-IG [RI-001] was provided for an immunocompromised infant with RSV-pneumonitis refractory to ribavirin and palivizumab. RSV-neutralizing antibody, respiratory RSV load (qPCR), and F-gene-sequence-detection of PR was determined. Prophylactic RSV-IG [RI-002] or palivizumab was administered in a cotton-rat model infected with wild-type and PR-RSV. Lung RSV load and neutralizing antibody were measured. RESULTS: As protective RI-001-neutralizing antibody titers waned in the infant, a subpopulation of PR-escape mutants were detected with a fatal RSV-burden in the lungs. In PR-RSV-infected cotton rats, prophylactic RI-002 reduced RSV-load in the lungs (2.45 vs 0.28 log10 PFU/g lung-tissue reduction, respectively, p < 0.05) and provided protective RSV-neutralizing antibody. CONCLUSIONS: RSV-IG and ribavirin use in immunocompromised patients requires further study.

Adult Memory T Cell Responses to the Respiratory Syncytial Virus Fusion Protein During a Single RSV Season (2018-2019).

Respiratory Syncytial Virus (RSV) is ubiquitous and re-infection with both subtypes (RSV/A and RSV/B) is common. The fusion (F) protein of RSV is antigenically conserved, induces neutralizing antibodies, and is a primary target of vaccine development. Insight into the breadth and durability of RSV-specific adaptive immune response, particularly to the F protein, may shed light on susceptibility to re-infection. We prospectively enrolled healthy adult subjects (n = 19) and collected serum and peripheral blood mononuclear cells (PBMCs) during the 2018-2019 RSV season. Previously, we described their RSV-specific antibody responses and identified three distinct antibody kinetic profiles associated with infection status: uninfected (n = 12), acutely infected (n = 4), and recently infected (n = 3). In this study, we measured the longevity of RSV-specific memory T cell responses to the F protein following natural RSV infection. We stimulated PBMCs with overlapping 15-mer peptide libraries spanning the F protein derived from either RSV/A or RSV/B and found that memory T cell responses mimic the antibody responses for all three groups. The uninfected group had stable, robust memory T cell responses and polyfunctionality. The acutely infected group had reduced polyfunctionality of memory T cell response at enrollment compared to the uninfected group, but these returned to comparable levels by end-of-season. The recently infected group, who were unable to maintain high levels of RSV-specific antibody following infection, similarly had decreased memory T cell responses and polyfunctionality during the RSV season. We observed subtype-specific differences in memory T cell responses and polyfunctionality, with RSV/A stimulating stronger memory T cell responses with higher polyfunctionality even though RSV/B was the dominant subtype in circulation. A subset of individuals demonstrated an overall deficiency in the generation of a durable RSV-specific adaptive immune response. Because memory T cell polyfunctionality may be associated with protection against re-infection, this latter group would likely be at greater risk of re-infection. Overall, these results expand our understanding of the longevity of the adaptive immune response to the RSV fusion protein and should be considered in future vaccine development efforts.

Antibody responses of healthy adults to the p27 peptide of respiratory syncytial virus fusion protein.

The respiratory syncytial virus (RSV) fusion (F) protein undergoes two furin-cleavage events to become fusion competent, resulting in the release of a twenty-seven amino acid peptide (p27). Recent studies indicate that the p27 region of the F protein was an immunodominant antigen in young children. In this study, we evaluated the kinetics of the serum antibody response to the p27 peptide following natural RSV reinfection in adults. Nineteen healthy adults under sixty-five years of age were enrolled during the 2018-2019 RSV season in Houston, TX. Blood was collected at three study visits and RSV infection status was defined by changes in neutralizing antibody resulting in three groups: uninfected (n = 12), acutely infected (n = 4), and recently infected (n = 3). Serum IgG and IgA antibodies against RSV/A and RSV/B p27 peptides were measured by enzyme-linked immunosorbent assays, and serum p27-like antibodies were detected by a p27 competitive antibody assay. Anti-p27 antibodies were detected in all subjects at each study visit. The measured IgG and IgA anti-p27 antibody levels followed the same pattern as other RSV site-specific and neutralizing antibody responses described for this cohort previously: the uninfected group had stable responses for the duration of the study period, the acutely infected group had a significant increase following RSV infection, and the recently infected group had a decrease in anti-p27 antibody during the study period. These results indicate that antibodies to the p27 region of the F protein are generated following natural RSV reinfection and suggest that some of the F protein is potentially in a partially cleaved state on the surface of virions, expanding on the previous assumption that all of p27 is post-translationally released and not present on mature F. Additionally, antibody responses were significantly lower (1.4-1.5-fold) toward RSV/B than to RSV/A p27 at each study visit, despite being an RSV/B dominant outbreak. Understanding the mechanism for the differences in the magnitude of the RSV/A and RSV/B p27 antibody response may enhance our understanding of the intracellular processing of the F protein.

A prospective surveillance study on the kinetics of the humoral immune response to the respiratory syncytial virus fusion protein in adults in Houston, Texas.

Respiratory syncytial virus (RSV)-specific serum antibody has been correlated to protection of infection and reduction of severe disease, but reinfection is still frequent. In this study, we evaluated RSV-specific serum antibody activity following natural RSV re-infection to examine the longevity of the humoral immune response in adults. Nineteen healthy adult volunteers under sixty-five years of age were enrolled during the 2018-2019 RSV season in Houston, TX. Blood was collected at three study visits. The kinetics of RSV-neutralizing, RSV F site-specific competitive, and RSV-binding antibodies in serum samples were measured by microneutralization and enzyme-linked immunosorbent assays. Three distinct profiles of RSV-specific antibody kinetics were identified that were consistent with RSV infection status: uninfected, acutely infected, and recently infected. The uninfected group had stable antibody titers for the duration of the study period (185 days). The acutely infected group had lower antibody responses at the beginning of the study, supporting a correlate of infection, followed by a significant antibody response after infection that was maintained for at least 125 days. Unlike the acutely infected group, the recently infected group had a significant precipitous decrease in RSV antibody in only 60 days. This study is the first, to our knowledge, to describe this abrupt loss of RSV-specific antibody in detail. This rapid decline of antibody may present an obstacle for the development of vaccines with lasting protection against RSV, and perhaps other respiratory pathogens. Neutralizing antibody responses were greater to prototypic than contemporaneous RSV strains, regardless of infection status, indicating that original antigenic sin may impact the humoral immune response to new or emerging RSV strains.

Electronic cigarettes disrupt lung lipid homeostasis and innate immunity independent of nicotine.

Electronic nicotine delivery systems (ENDS) or e-cigarettes have emerged as a popular recreational tool among adolescents and adults. Although the use of ENDS is often promoted as a safer alternative to conventional cigarettes, few comprehensive studies have assessed the long-term effects of vaporized nicotine and its associated solvents, propylene glycol (PG) and vegetable glycerin (VG). Here, we show that compared with smoke exposure, mice receiving ENDS vapor for 4 months failed to develop pulmonary inflammation or emphysema. However, ENDS exposure, independent of nicotine, altered lung lipid homeostasis in alveolar macrophages and epithelial cells. Comprehensive lipidomic and structural analyses of the lungs revealed aberrant phospholipids in alveolar macrophages and increased surfactant-associated phospholipids in the airway. In addition to ENDS-induced lipid deposition, chronic ENDS vapor exposure downregulated innate immunity against viral pathogens in resident macrophages. Moreover, independent of nicotine, ENDS-exposed mice infected with influenza demonstrated enhanced lung inflammation and tissue damage. Together, our findings reveal that chronic e-cigarette vapor aberrantly alters the physiology of lung epithelial cells and resident immune cells and promotes poor response to infectious challenge. Notably, alterations in lipid homeostasis and immune impairment are independent of nicotine, thereby warranting more extensive investigations of the vehicle solvents used in e-cigarettes.

MegaRibavirin aerosol for the treatment of influenza A virus infections in mice.

While newer neuraminidase inhibitors have been used recently to treat influenza A and B virus infections, emergence of drug resistance poses potential problems. Previous ribavirin aerosol treatments of influenza were effective and drug resistance was not observed. To make ribavirin aerosol treatment a quicker process and limited to once or twice daily treatments, a MegaRibavirin formulation (100 mg of ribavirin/mL) was developed that when used with the Aerotech II nebulizer was effective in preventing death in a lethal influenza A virus mouse model. Aerosol generated using the Aerotech II nebulizer flowing at 10 L of air/min produced aerosol droplets that contained 2.3 mg of ribavirin/L with a mass median aerodynamic diameter of 1.8 microm. Using this system for treatment, a single daily 30-min exposure on days 1-4 produced a survival rate of greater than 90%. Delaying the start of aerosol treatment for 48 or 72 h and treating once daily for 30 min for two days (days 2-3 and 3-4, respectively) still significantly increased the number of survivors and mean time to death. For the treatment of influenza in general and for pandemic avian influenza, the MegaRibavirin-Aerotech II method of aerosol treatment allows for short treatment periods, minimizes environmental issues and costs less.

Respiratory syncytial virus fusion nanoparticle vaccine immune responses target multiple neutralizing epitopes that contribute to protection against wild-type and palivizumab-resistant mutant virus challenge.

Human respiratory syncytial virus (RSV) is the leading cause of severe lower respiratory tract infections in newborns, young children, elderly, and immune-compromised. The RSV fusion (F) glycoprotein is a major focus of vaccine development and the target of palivizumab (Synagis®) which is licensed as an immuno-prophylactic for use in newborn children at high risk of infection. However, clinical use of a narrowly targeted monoclonal antibodies leads to the generation of escape mutant strains that are fully resistant to neutralization by the antibody. Herein, we evaluated the RSV F nanoparticle vaccine (RSV F vaccine), produced as near-full-length, pre-fusogenic F trimers that form stable protein-detergent nanoparticles. The RSV F vaccine induces polyclonal antibodies that bind to antigenic site II as well as other epitopes known to be broadly neutralizing. Cotton rats immunized with the RSV F vaccine produced antibodies that were both neutralizing and protected against wild-type RSV infection, as well as against a palivizumab-resistant mutant virus. Use of aluminum phosphate adjuvant with the RSV F vaccine increased site II antibody avidity 100 to 1000-fold, which correlated with enhanced protection against challenge. The breadth of the vaccine-induced antibody response was demonstrated using competitive binding with monoclonal antibodies targeting antigenic sites Ø, II, IV, and VIII found on pre-fusion and post-fusion conformations of RSV F. In summary, we found the RSV F vaccine induced antibodies that bind to conserved epitopes including those defined as pre-fusion F specific; that use of adjuvant increased antibody avidity that correlated with enhanced protection in the cotton rat challenge model; and the polyclonal, high-avidity antibodies neutralized and protected against both wild-type and palivizumab-resistant mutant virus. These data support the ongoing clinical development of the aluminum phosphate adjuvanted RSV F nanoparticle vaccine.

Combined aerosolized Toll-like receptor ligands are an effective therapeutic agent against influenza pneumonia when co-administered with oseltamivir.

Influenza pneumonia remains a common and debilitating viral infection despite vaccination programs and antiviral agents developed for prophylaxis and treatment. The neuraminidase inhibitor oseltamivir is frequently prescribed for established influenza A virus infections, but the emergence of neuraminidase inhibitor resistant viruses, a brief therapeutic window and competing diagnoses complicate its use. PUL-042 is a clinical stage, aerosol drug comprised of synthetic ligands for Toll-like receptor (TLR) 2/6 and TLR 9. This host-targeted, innate immune stimulant broadly protects against bacterial, fungal and viral pneumonias, including those caused by influenza, when given prophylactically to animals. This study evaluated the therapeutic antiviral effects of PUL-042 against established influenza A pneumonia, when given alone or in combination with oseltamivir. Mice were treated with PUL-042 aerosol, oseltamivir or both at varying time points before or after challenge with influenza pneumonia. Treating established, otherwise lethal influenza A pneumonia (>1 LD100) with multiple inhaled doses of PUL-042 aerosol plus oral oseltamivir resulted in greater mouse survival than treatment with either drug alone. Single agent PUL-042 also protected mice against established infections following challenges with lower viral inocula (approximately 1 LD20). Aerosolized oseltamivir further enhanced survival when co-delivered with PUL-042 aerosol. The prophylactic and therapeutic benefits of PUL-042 were similar against multiple strains of influenza virus. In vitro influenza challenge of human HBEC3kt lung epithelial cells revealed PUL-042-induced protection against infection that was comparable to that observed in vivo. These studies offer new insights into means to protect susceptible populations against influenza A pneumonia.

Correction: Repurposed Transcriptomic Data Reveal Small Viral RNA Produced by Influenza Virus during Infection in Mice.

[This corrects the article DOI: 10.1371/journal.pone.0165729.].

AIMp1 Potentiates TH1 Polarization and Is Critical for Effective Antitumor and Antiviral Immunity.

Dendritic cells (DCs) must integrate a broad array of environmental cues to exact control over downstream immune responses including TH polarization. The multienzyme aminoacyl-tRNA synthetase complex component AIMp1/p43 responds to cellular stress and exerts pro-inflammatory functions; however, a role for DC-expressed AIMp1 in TH polarization has not previously been shown. Here, we demonstrate that the absence of AIMp1 in bone marrow-derived DC (BMDC) significantly impairs cytokine and costimulatory molecule expression, p38 MAPK signaling, and TH1 polarization of cocultured T-cells while significantly dysregulating immune-related gene expression. These deficits resulted in significantly compromised BMDC vaccine-mediated protection against melanoma. AIMp1 within the host was also critical for innate and adaptive antiviral immunity against influenza virus infection in vivo. Cancer patients with AIMp1 expression levels in the highest tertiles exhibited a 70% survival advantage at 15-year postdiagnosis as determined by bioinformatics analysis of nearly 9,000 primary human tumor samples in The Cancer Genome Atlas database. These data establish the importance of AIMp1 for the effective governance of antitumor and antiviral immune responses.

Repurposed Transcriptomic Data Reveal Small Viral RNA Produced by Influenza Virus during Infection in Mice.

Influenza virus, a highly infectious ssRNA virus, replicates in the nucleus of host cells. This unusual feature brings the possibility that the virus may hijack host small noncoding RNA metabolism. Influenza small viral RNA production has been examined in vitro but has not yet been studied in an in vivo setting. We assessed small RNA species from influenza virus during mouse infection by mining publicly available mouse small RNA transcriptome data. We uncovered 26 nt reads corresponding to svRNA, a small viral RNA previously detected in vitro that regulates the transition from transcription to replication during infection, and found a strong positive correlation between svRNA production and host susceptibility to influenza virus infection. We also detected significant overrepresentation of a non-coding 23 nt sequence that we speculate may behave like a miRNA and work with influenza protein NS1 to prevent the transcription and maturation of interferon-stimulated mRNAs.

A live RSV vaccine with engineered thermostability is immunogenic in cotton rats despite high attenuation.

Respiratory syncytial virus (RSV) is a leading cause of infant hospitalization and there remains no pediatric vaccine. RSV live-attenuated vaccines (LAVs) have a history of safe testing in infants; however, achieving an effective balance of attenuation and immunogenicity has proven challenging. Here we seek to engineer an RSV LAV with enhanced immunogenicity. Genetic mapping identifies strain line 19 fusion (F) protein residues that correlate with pre-fusion antigen maintenance by ELISA and thermal stability of infectivity in live RSV. We generate a LAV candidate named OE4 which expresses line 19F and is attenuated by codon-deoptimization of non-structural (NS1 and NS2) genes, deletion of the small hydrophobic (SH) gene, codon-deoptimization of the attachment (G) gene and ablation of the secreted form of G. OE4 (RSV-A2-dNS1-dNS2-ΔSH-dGm-Gsnull-line19F) exhibits elevated pre-fusion antigen levels, thermal stability, immunogenicity, and efficacy despite heavy attenuation in the upper and lower airways of cotton rats.