Safety and Efficacy of Dolutegravir in Treatment-Experienced Subjects With Raltegravir-Resistant HIV Type 1 Infection: 24-Week Results of the VIKING Study

Background. Dolutegravir (DTG; S/GSK1349572), a human immunodeficiency virus type 1 (HIV-1) integrase inhibitor, has limited cross-resistance to raltegravir (RAL) and elvitegravir in vitro. This phase IIb study assessed the activity of DTG in HIV-1–infected subjects with genotypic evidence of RAL resistance.Methods. Subjects received DTG 50 mg once daily (cohort I) or 50 mg twice daily (cohort II) while continuing a failing regimen (without RAL) through day 10, after which the background regimen was optimized, when feasible, for cohort I, and at least 1 fully active drug was mandated for cohort II. The primary efficacy end point was the proportion of subjects on day 11 in whom the plasma HIV-1 RNA load decreased by ≥0.7 log10 copies/mL from baseline or was <400 copies/mL.Results. A rapid antiviral response was observed. More subjects achieved the primary end point in cohort II (23 of 24 [96%]), compared with cohort I (21 of 27 [78%]) at day 11. At week 24, 41% and 75% of subjects had an HIV-1 RNA load of <50 copies/mL in cohorts I and II, respectively. Further integrase genotypic evolution was uncommon. Dolutegravir had a good, similar safety profile with each dosing regimen.Conclusion. Dolutegravir 50 mg twice daily with an optimized background provided greater and more durable benefit than the once-daily regimen. These data are the first clinical demonstration of the activity of any integrase inhibitor in subjects with HIV-1 resistant to RAL

Whole-genome sequencing reveals host factors underlying critical COVID-19

Critical COVID-19 is caused by immune-mediated inflammatory lung injury. Host genetic variation influences the development of illness requiring critical care1 or hospitalization2–4 after infection with SARS-CoV-2. The GenOMICC (Genetics of Mortality in Critical Care) study enables the comparison of genomes from individuals who are critically ill with those of population controls to find underlying disease mechanisms. Here we use whole-genome sequencing in 7,491 critically ill individuals compared with 48,400 controls to discover and replicate 23 independent variants that significantly predispose to critical COVID-19. We identify 16 new independent associations, including variants within genes that are involved in interferon signalling (IL10RB and PLSCR1), leucocyte differentiation (BCL11A) and blood-type antigen secretor status (FUT2). Using transcriptome-wide association and colocalization to infer the effect of gene expression on disease severity, we find evidence that implicates multiple genes—including reduced expression of a membrane flippase (ATP11A), and increased expression of a mucin (MUC1)—in critical disease. Mendelian randomization provides evidence in support of causal roles for myeloid cell adhesion molecules (SELE, ICAM5 and CD209) and the coagulation factor F8, all of which are potentially druggable targets. Our results are broadly consistent with a multi-component model of COVID-19 pathophysiology, in which at least two distinct mechanisms can predispose to life-threatening disease: failure to control viral replication; or an enhanced tendency towards pulmonary inflammation and intravascular coagulation. We show that comparison between cases of critical illness and population controls is highly efficient for the detection of therapeutically relevant mechanisms of disease

Colliding Worlds: Asteroid research and the legitimization of war in space

Accepted versio

Whole-genome sequencing reveals host factors underlying critical COVID-19

Critical COVID-19 is caused by immune-mediated inflammatory lung injury. Host genetic variation influences the development of illness requiring critical care1 or hospitalization2,3,4 after infection with SARS-CoV-2. The GenOMICC (Genetics of Mortality in Critical Care) study enables the comparison of genomes from individuals who are critically ill with those of population controls to find underlying disease mechanisms. Here we use whole-genome sequencing in 7,491 critically ill individuals compared with 48,400 controls to discover and replicate 23 independent variants that significantly predispose to critical COVID-19. We identify 16 new independent associations, including variants within genes that are involved in interferon signalling (IL10RB and PLSCR1), leucocyte differentiation (BCL11A) and blood-type antigen secretor status (FUT2). Using transcriptome-wide association and colocalization to infer the effect of gene expression on disease severity, we find evidence that implicates multiple genes—including reduced expression of a membrane flippase (ATP11A), and increased expression of a mucin (MUC1)—in critical disease. Mendelian randomization provides evidence in support of causal roles for myeloid cell adhesion molecules (SELE, ICAM5 and CD209) and the coagulation factor F8, all of which are potentially druggable targets. Our results are broadly consistent with a multi-component model of COVID-19 pathophysiology, in which at least two distinct mechanisms can predispose to life-threatening disease: failure to control viral replication; or an enhanced tendency towards pulmonary inflammation and intravascular coagulation. We show that comparison between cases of critical illness and population controls is highly efficient for the detection of therapeutically relevant mechanisms of disease

Role of Plasma Membrane Lipid Microdomains in Respiratory Syncytial Virus Filament Formation

The fusion protein (F) of respiratory syncytial virus (RSV) is the envelope glycoprotein responsible for the characteristic cytopathology of syncytium formation. RSV has been shown to bud from selective areas of the plasma membrane as pleomorphic virions, including both filamentous and round particles. With immunofluorescent microscopy, we demonstrated evidence of RSV filaments incorporating the fusion protein F and colocalizing with a lipid microdomain-specific fluorescent dye, 1,1-dihexadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate. Western blot analysis of Triton X-100 cold-extracted membrane fractions confirmed the presence of RSV proteins within the lipid microdomains. RSV proteins also colocalized with cellular proteins associated with lipid microdomains, caveolin-1, and CD44, as well as with RhoA, a small GTPase. ADP-ribosylation of RhoA by Clostridium botulinum exotoxin inactivated RhoA signaling and resulted in the absence of RSV-induced syncytia despite no significant change in viral titer. We demonstrated an overall decrease in both the number and length of the viral filaments and a shift in the localization of F to nonlipid microdomain regions of the membrane in the presence of C3 toxin. This suggests that the selective incorporation of RSV proteins into lipid microdomains during virus assembly may lead to critical interactions of F with cellular proteins, resulting in microvillus projections necessary for the formation of filamentous virus particles and syncytium formation. Thus, manipulation of membrane lipid microdomains may lead to alterations in the production of viral filaments and RSV pathogenesis and provide a new pharmacologic target for RSV therapy

Modified Vaccinia Virus Ankara Immunization Protects against Lethal Challenge with Recombinant Vaccinia Virus Expressing Murine Interleukin-4

Recent events have raised concern over the use of pathogens, including variola virus, as biological weapons. Vaccination with Dryvax is associated with serious side effects and is contraindicated for many people, and the development of a safer effective smallpox vaccine is necessary. We evaluated an attenuated vaccinia virus, modified vaccinia virus Ankara (MVA), by use of a murine model to determine its efficacy against an intradermal (i.d.) or intranasal (i.n.) challenge with vaccinia virus (vSC8) or a recombinant vaccinia virus expressing murine interleukin-4 that exhibits enhanced virulence (vSC8-mIL4). After an i.d. challenge, 15 of 16 mice who were inoculated with phosphate-buffered saline developed lesions, one dose of intramuscularly administered MVA was partially protective (3 of 16 mice developed lesions), and the administration of two or three doses of MVA was completely protective (0 of 16 mice developed lesions). In unimmunized mice, an i.n. challenge with vSC8 caused a significant but self-limited illness, while vSC8-mIL4 resulted in lethal infections. Immunization with one or two doses of MVA prevented illness and reduced virus titers in mice who were challenged with either vSC8 or vSC8-mIL4. MVA induced a dose-related neutralizing antibody and vaccinia virus-specific CD8(+)-T-cell response. Mice immunized with MVA were fully protected from a low-dose vSC8-mIL4 challenge despite a depletion of CD4(+) cells, CD8(+) cells, or both T-cell subsets or an antibody deficiency. CD4(+)- or CD8(+)-T-cell depletion reduced the protection against a high-dose vSC8-mIL4 challenge, and the depletion of both T-cell subsets was associated with severe illness and higher vaccinia virus titers. Thus, MVA induces broad humoral and cellular immune responses that can independently protect against a molecularly modified lethal poxvirus challenge in mice. These data support the continued development of MVA as an alternative candidate vaccine for smallpox

RhoA Signaling Is Required for Respiratory Syncytial Virus-Induced Syncytium Formation and Filamentous Virion Morphology

Respiratory syncytial virus (RSV) is an important human pathogen that can cause severe and life-threatening respiratory infections in infants, the elderly, and immunocompromised adults. RSV infection of HEp-2 cells induces the activation of RhoA, a small GTPase. We therefore asked whether RhoA signaling is important for RSV replication or syncytium formation. The treatment of HEp-2 cells with Clostridium botulinum C3, an enzyme that ADP-ribosylates and specifically inactivates RhoA, inhibited RSV-induced syncytium formation and cell-to-cell fusion, although similar levels of PFU were released into the medium and viral protein expression levels were equivalent. Treatment with another inhibitor of RhoA signaling, the Rho kinase inhibitor Y-27632, yielded similar results. Scanning electron microscopy of C3-treated infected cells showed reduced numbers of single blunted filaments, in contrast to the large clumps of long filaments in untreated infected cells. These data suggest that RhoA signaling is associated with filamentous virus morphology, cell-to-cell fusion, and syncytium formation but is dispensable for the efficient infection and production of infectious virus in vitro. Next, we developed a semiquantitative method to measure spherical and filamentous virus particles by using sucrose gradient velocity sedimentation. Fluorescence and transmission electron microscopy confirmed the separation of spherical and filamentous forms of infectious virus into two identifiable peaks. The C3 treatment of RSV-infected cells resulted in a shift to relatively more spherical virions than those from untreated cells. These data suggest that viral filamentous protuberances characteristic of RSV infection are associated with RhoA signaling, are important for filamentous virion morphology, and may play a role in initiating cell-to-cell fusion

Leukotriene B4 amplifies NF-κB activation in mouse macrophages by reducing SOCS1 inhibition of MyD88 expression

Activation of NF-κB and 5-lipoxygenase–mediated (5-LO–mediated) biosynthesis of the lipid mediator leukotriene B4 (LTB4) are pivotal components of host defense and inflammatory responses. However, the role of LTB4 in mediating innate immune responses elicited by specific TLR ligands and cytokines is unknown. Here we have shown that responses dependent on MyD88 (an adaptor protein that mediates signaling through all of the known TLRs, except TLR3, as well as IL-1β and IL-18) are reduced in mice lacking either 5-LO or the LTB4 receptor BTL1, and that macrophages from these mice are impaired in MyD88-dependent activation of NF-κB. This macrophage defect was associated with lower basal and inducible expression of MyD88 and reflected impaired activation of STAT1 and overexpression of the STAT1 inhibitor SOCS1. Expression of MyD88 and responsiveness to the TLR4 ligand LPS were decreased by Stat1 siRNA silencing in WT macrophages and restored by Socs1 siRNA in 5-LO–deficient macrophages. These results uncover a pivotal role in macrophages for the GPCR BLT1 in regulating activation of NF-κB through Stat1-dependent expression of MyD88