Context-dependent role of group 3 innate lymphoid cells in mucosal protection.

How group 3 innate lymphoid cells (ILC3s) regulate mucosal protection in the presence of T cells remains poorly understood. Here, we examined ILC3 function in intestinal immunity using ILC3-deficient mice that maintain endogenous T cells, T helper 17 (TH17) cells, and secondary lymphoid organs. ILC3s were dispensable for generation of TH17 and TH22 cell responses to commensal and pathogenic bacteria, and absence of ILC3s did not affect IL-22 production by CD4 T cells before or during infection. However, despite the presence of IL-22-producing T cells, ILC3s and ILC3-derived IL-22 were required for maintaining homeostatic functions of the intestinal epithelium. T cell-sufficient, ILC3-deficient mice were capable of pathogen clearance and survived infection with a low dose of Citrobacter rodentium. However, ILC3s promoted pathogen tolerance at early time points of infection by activating tissue-protective immune pathways. Consequently, ILC3s were indispensable for survival after high-dose infection. Our results demonstrate a context-dependent role for ILC3s in immune-sufficient animals and provide a blueprint for uncoupling of ILC3 and TH17 cell functions.

A CAR enhancer increases the activity and persistence of CAR T cells.

Although chimeric antigen receptor (CAR) T cell therapies have demonstrated promising clinical outcomes, durable remissions remain limited. To extend the efficacy of CAR T cells, we develop a CAR enhancer (CAR-E), comprising a CAR T cell antigen fused to an immunomodulatory molecule. Here we demonstrate this strategy using B cell maturation antigen (BCMA) CAR T cells for the treatment of multiple myeloma, with a CAR-E consisting of the BCMA fused to a low-affinity interleukin 2 (IL-2). This selectively induces IL-2 signaling in CAR T cells upon antigen-CAR binding, enhancing T cell activation and antitumor activity while reducing IL-2-associated toxicities. We show that the BCMA CAR-E selectively binds CAR T cells and increases CAR T cell proliferation, clearance of tumor cells and development of memory CAR T cells. The memory cells retain the ability to re-expand upon restimulation, effectively controlling tumor growth upon rechallenge. Mechanistic studies reveal the involvement of both CAR and IL-2 receptor endodomains in the CAR-E mechanism of action. The CAR-E approach avoids the need for specific engineering and enables CAR T cell therapy with lower cell doses.

Microbiota metabolized Bile Acids accelerate Gastroesophageal Adenocarcinoma via FXR inhibition.

Background: The incidence of Barrett esophagus (BE) and Gastroesophageal Adenocarcinoma (GEAC) correlates with obesity and a diet rich in fat. Bile acids (BA) support fat digestion and undergo microbial metabolization in the gut. The farnesoid X receptor (FXR) is an important modulator of the BA homeostasis. The capacity of inhibiting cancer-related processes when activated, make FXR an appealing therapeutic target. In this work, we assess the role of diet on the microbiota-BA axis and evaluate the role of FXR in disease progression. Results: Here we show that high fat diet (HFD) accelerated tumorigenesis in L2-IL1B mice (BE- and GEAC- mouse model) while increasing BA levels and enriching gut microbiota that convert primary to secondary BA. While upregulated in BE, expression of FXR was downregulated in GEAC in mice and humans. In L2-IL1B mice, FXR knockout enhanced the dysplastic phenotype and increased Lgr5 progenitor cell numbers. Treatment of murine organoids and L2-IL1B mice with the FXR agonist obeticholic acid (OCA) deacelerated GEAC progression. Conclusion: We provide a novel concept of GEAC carcinogenesis being accelerated via the diet-microbiome-metabolome axis and FXR inhibition on progenitor cells. Further, FXR activation protected with OCA ameliorated the phenotype in vitro and in vivo, suggesting that FXR agonists have potential as differentiation therapy in GEAC prevention.

SARS-CoV-2 omicron BA.2.87.1 exhibits higher susceptibility to serum neutralization than EG.5.1 and JN.1.

As SARS-CoV-2 continues to spread and mutate, tracking the viral evolutionary trajectory and understanding the functional consequences of its mutations remain crucial. Here, we characterized the antibody evasion, ACE2 receptor engagement, and viral infectivity of the highly mutated SARS-CoV-2 Omicron subvariant BA.2.87.1. Compared with other Omicron subvariants, including EG.5.1 and the current predominant JN.1, BA.2.87.1 exhibits less immune evasion, reduced viral receptor engagement, and comparable infectivity in Calu-3 lung cells. Intriguingly, two large deletions (Δ15-26 and Δ136-146) in the N-terminal domain (NTD) of the spike protein facilitate subtly increased antibody evasion but significantly diminish viral infectivity. Collectively, our data support the announcement by the USA CDC that the public health risk posed by BA.2.87.1 appears to be low.

Stool Protein Mass Spectrometry Identifies Biomarkers for Early Detection of Diffuse-type Gastric Cancer.

There is a high unmet need for early detection approaches for diffuse gastric cancer (DGC). We examined whether the stool proteome of mouse models of gastric cancer (GC) and individuals with hereditary diffuse gastric cancer (HDGC) have utility as biomarkers for early detection. Proteomic mass spectrometry of the stool of a genetically engineered mouse model driven by oncogenic KrasG12D and loss of p53 and Cdh1 in gastric parietal cells [known as Triple Conditional (TCON) mice] identified differentially abundant proteins compared with littermate controls. Immunoblot assays validated a panel of proteins, including actinin alpha 4 (ACTN4), N-acylsphingosine amidohydrolase 2 (ASAH2), dipeptidyl peptidase 4 (DPP4), and valosin-containing protein (VCP), as enriched in TCON stool compared with littermate control stool. Immunofluorescence analysis of these proteins in TCON stomach sections revealed increased protein expression compared with littermate controls. Proteomic mass spectrometry of stool obtained from patients with HDGC with CDH1 mutations identified increased expression of ASAH2, DPP4, VCP, lactotransferrin (LTF), and tropomyosin-2 relative to stool from healthy sex- and age-matched donors. Chemical inhibition of ASAH2 using C6 urea ceramide was toxic to GC cell lines and GC patient-derived organoids. This toxicity was reversed by adding downstream products of the S1P synthesis pathway, which suggested a dependency on ASAH2 activity in GC. An exploratory analysis of the HDGC stool microbiome identified features that correlated with patient tumors. Herein, we provide evidence supporting the potential of analyzing stool biomarkers for the early detection of DGC. Prevention Relevance: This study highlights a novel panel of stool protein biomarkers that correlate with the presence of DGC and has potential use as early detection to improve clinical outcomes.

COVID-19 vaccines and adverse events of special interest: A multinational Global Vaccine Data Network (GVDN) cohort study of 99 million vaccinated individuals.

BACKGROUND: The Global COVID Vaccine Safety (GCoVS) Project, established in 2021 under the multinational Global Vaccine Data Network™ (GVDN®), facilitates comprehensive assessment of vaccine safety. This study aimed to evaluate the risk of adverse events of special interest (AESI) following COVID-19 vaccination from 10 sites across eight countries. METHODS: Using a common protocol, this observational cohort study compared observed with expected rates of 13 selected AESI across neurological, haematological, and cardiac outcomes. Expected rates were obtained by participating sites using pre-COVID-19 vaccination healthcare data stratified by age and sex. Observed rates were reported from the same healthcare datasets since COVID-19 vaccination program rollout. AESI occurring up to 42 days following vaccination with mRNA (BNT162b2 and mRNA-1273) and adenovirus-vector (ChAdOx1) vaccines were included in the primary analysis. Risks were assessed using observed versus expected (OE) ratios with 95 % confidence intervals. Prioritised potential safety signals were those with lower bound of the 95 % confidence interval (LBCI) greater than 1.5. RESULTS: Participants included 99,068,901 vaccinated individuals. In total, 183,559,462 doses of BNT162b2, 36,178,442 doses of mRNA-1273, and 23,093,399 doses of ChAdOx1 were administered across participating sites in the study period. Risk periods following homologous vaccination schedules contributed 23,168,335 person-years of follow-up. OE ratios with LBCI > 1.5 were observed for Guillain-Barré syndrome (2.49, 95 % CI: 2.15, 2.87) and cerebral venous sinus thrombosis (3.23, 95 % CI: 2.51, 4.09) following the first dose of ChAdOx1 vaccine. Acute disseminated encephalomyelitis showed an OE ratio of 3.78 (95 % CI: 1.52, 7.78) following the first dose of mRNA-1273 vaccine. The OE ratios for myocarditis and pericarditis following BNT162b2, mRNA-1273, and ChAdOx1 were significantly increased with LBCIs > 1.5. CONCLUSION: This multi-country analysis confirmed pre-established safety signals for myocarditis, pericarditis, Guillain-Barré syndrome, and cerebral venous sinus thrombosis. Other potential safety signals that require further investigation were identified.

Bacterial genome engineering using CRISPR-associated transposases.

Clustered regularly interspaced short palindromic repeats (CRISPR)-associated transposases have the potential to transform the technology landscape for kilobase-scale genome engineering, by virtue of their ability to integrate large genetic payloads with high accuracy, easy programmability and no requirement for homologous recombination machinery. These transposons encode efficient, CRISPR RNA-guided transposases that execute genomic insertions in Escherichia coli at efficiencies approaching ~100%. Moreover, they generate multiplexed edits when programmed with multiple guides, and function robustly in diverse Gram-negative bacterial species. Here we present a detailed protocol for engineering bacterial genomes using CRISPR-associated transposase (CAST) systems, including guidelines on the available vectors, customization of guide RNAs and DNA payloads, selection of common delivery methods, and genotypic analysis of integration events. We further describe a computational CRISPR RNA design algorithm to avoid potential off-targets, and a CRISPR array cloning pipeline for performing multiplexed DNA insertions. The method presented here allows the isolation of clonal strains containing a novel genomic integration event of interest within 1-2 weeks using available plasmid constructs and standard molecular biology techniques.

High-throughput transcriptomics of 409 bacteria-drug pairs reveals drivers of gut microbiota perturbation.

Many drugs can perturb the gut microbiome, potentially leading to negative health consequences. However, mechanisms of most microorganism-drug responses have not been elucidated at the genetic level. Using high-throughput bacterial transcriptomics, we systematically characterized the gene expression profiles of prevalent human gut bacteria exposed to the most frequently prescribed orally administered pharmaceuticals. Across >400 drug-microorganism pairs, significant and reproducible transcriptional responses were observed, including pathways involved in multidrug resistance, metabolite transport, tartrate metabolism and riboflavin biosynthesis. Importantly, we discovered that statin-mediated upregulation of the AcrAB-TolC efflux pump in Bacteroidales species enhances microbial sensitivity to vitamin A and secondary bile acids. Moreover, gut bacteria carrying acrAB-tolC genes are depleted in patients taking simvastatin, suggesting that drug-efflux interactions generate collateral toxicity that depletes pump-containing microorganisms from patient microbiomes. This study provides a resource to further understand the drivers of drug-mediated microbiota shifts for better informed clinical interventions.

Alarming antibody evasion properties of rising SARS-CoV-2 BQ and XBB subvariants

The SARS-CoV-2 Omicron variant continues to evolve, with new BQ and XBB subvariants now rapidly expanding in Europe/US and Asia, respectively. As these new subvariants have additional spike mutations, they may possess altered antibody evasion properties. Here, we report that neutralization of BQ.1, BQ.1.1, XBB, and XBB.1 by sera from vaccinees and infected persons was markedly impaired, including sera from individuals who were boosted with a WA1/BA.5 bivalent mRNA vaccine. Compared to the ancestral strain D614G, serum neutralizing titers against BQ and XBB subvariants were lower by 13-81-fold and 66-155-fold, respectively, far beyond what had been observed to date. A panel of monoclonal antibodies capable of neutralizing the original Omicron variant, including those with Emergency Use Authorization, were largely inactive against these new subvariants. The spike mutations that conferred antibody resistance were individually studied and structurally explained. Finally, the ACE2-binding affinities of the spike proteins of these novel subvariants were found to be similar to those of their predecessors. Taken together, our findings indicate that BQ and XBB subvariants present serious threats to the efficacy of current COVID-19 vaccines, render inactive all authorized monoclonal antibodies, and may have gained dominance in the population because of their advantage in evading antibodies.

Antibody Evasion Properties of SARS-CoV-2 Omicron Sublineages

The identification of the Omicron variant (B.1.1.529.1 or BA.1) of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) in Botswana in November 20211 immediately raised alarms due to the sheer number of mutations in the spike glycoprotein that could lead to striking antibody evasion. We2 and others3-6 recently reported results in this Journal confirming such a concern. Continuing surveillance of Omicron evolution has since revealed the rise in prevalence of two sublineages, BA.1 with an R346K mutation (BA.1+R346K) and B.1.1.529.2 (BA.2), with the latter containing 8 unique spike mutations while lacking 13 spike mutations found in BA.1. We therefore extended our studies to include antigenic characterization of these new sublineages. Polyclonal sera from patients infected by wild-type SARS-CoV-2 or recipients of current mRNA vaccines showed a substantial loss in neutralizing activity against both BA.1+R346K and BA.2, with drops comparable to that already reported for BA.12,3,5,6. These findings indicate that these three sublineages of Omicron are antigenically equidistant from the wild-type SARS-CoV-2 and thus similarly threaten the efficacies of current vaccines. BA.2 also exhibited marked resistance to 17 of 19 neutralizing monoclonal antibodies tested, including S309 (sotrovimab)7, which had retained appreciable activity against BA.1 and BA.1+R346K2-4,6. This new finding shows that no presently approved or authorized monoclonal antibody therapy could adequately cover all sublineages of the Omicron variant.

Structural basis for antibody resistance to SARS-CoV-2 omicron variant

The recently reported B.1.1.529 Omicron variant of SARS-CoV-2 includes 34 mutations in the spike protein relative to the Wuhan strain that initiated the COVID-19 pandemic, including 15 mutations in the receptor binding domain (RBD). Functional studies have shown omicron to substantially escape the activity of many SARS-CoV-2-neutralizing antibodies. Here we report a 3.1 [A] resolution cryo-electron microscopy (cryo-EM) structure of the Omicron spike protein ectodomain. The structure depicts a spike that is exclusively in the 1-RBD-up conformation with increased mobility and inter-protomer asymmetry. Many mutations cause steric clashes and/or altered interactions at antibody binding surfaces, whereas others mediate changes of the spike structure in local regions to interfere with antibody recognition. Overall, the structure of the omicron spike reveals how mutations alter its conformation and explains its extraordinary ability to evade neutralizing antibodies. HighlightsO_LISARS-CoV-2 omicron spike exclusively adopts 1-RBD-up conformation C_LIO_LIOmicron substitutions alter conformation and mobility of RBD C_LIO_LIA subset of omicron mutations change the local conformation of spike C_LIO_LIThe structure reveals the basis of antibody neutralization escape C_LI

Striking Antibody Evasion Manifested by the Omicron Variant of SARS-CoV-2

The Omicron (B.1.1.529) variant of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) was only recently detected in southern Africa, but its subsequent spread has been extensive, both regionally and globally1. It is expected to become dominant in the coming weeks2, probably due to enhanced transmissibility. A striking feature of this variant is the large number of spike mutations3 that pose a threat to the efficacy of current COVID-19 (coronavirus disease 2019) vaccines and antibody therapies4. This concern is amplified by the findings from our study. We found B.1.1.529 to be markedly resistant to neutralization by serum not only from convalescent patients, but also from individuals vaccinated with one of the four widely used COVID-19 vaccines. Even serum from persons vaccinated and boosted with mRNA-based vaccines exhibited substantially diminished neutralizing activity against B.1.1.529. By evaluating a panel of monoclonal antibodies to all known epitope clusters on the spike protein, we noted that the activity of 17 of the 19 antibodies tested were either abolished or impaired, including ones currently authorized or approved for use in patients. In addition, we also identified four new spike mutations (S371L, N440K, G446S, and Q493R) that confer greater antibody resistance to B.1.1.529. The Omicron variant presents a serious threat to many existing COVID-19 vaccines and therapies, compelling the development of new interventions that anticipate the evolutionary trajectory of SARS-CoV-2.

Contact investigation in the prevention of mother-to-child transmission of HIV comparing urban and rural outcomes in Jamaica / Investigación de contactos en la prevención de la transmisión del VIH de madre a hijo comparando los resultados urbanos y rurales en Jamaica

BACKGROUND AND PURPOSE: The contact investigators played a significant role in the decline of infectious syphilis in Jamaica and are likely important players in the prevention of mother-to-child transmission (PMTCT) in the HIV programme. A brief evaluation was done comparing the outcomes of contact investigation in Kingston and St Andrew (KSA) with that of the contact investigation in two rural parishes? METHODS: The interview and field records for the seropositive antenatal clinic attendees for the period October 2004 to September 2005, in urban KSA, were compared with those for rural Clarendon and Portland. RESULTS: HIV seropositive pregnant women (n = 88) were notified and/or referred to the parish contact investigators: 36 in KSA, 9 in Portland and 43 in Clarendon. The time from test date to interview date was almost twice as long for KSA (mean 27 days) than Portland (mean 15.7 days) and thrice that of Clarendon (mean 9 days). Mean disposition (case closure) times were for KSA: 19 days; Portland: 28 days and Clarendon: 15 days. Only 40% of the contacts were located for KSA and 48% of these tested positive for HIV. For Portland, 73% were located and 8% tested positive. For Clarendon, 45% were located and 35% of these tested positive. CONCLUSIONES: On site same day HIV rapid testing is not always available so the contact investigator is an essential member of the pMTCT team in Jamaica. One of the programme outcomes (time to interview) was longer in the urban than the rural parishes while others (time to resolution of the case and percentage of contacts located and tested) had no consistent urban-rural differences.

ANTECEDENTES Y PROPÓSITO: Los investigadores de contactos desempeñaron un papel significativo en el disminución de la sífilis infecciosa en Jamaica, y son probablemente agentes importantes en la prevención de la transmisión del VIH de madre a hijo dentro del programa de VIH. Se realizó una breve evaluación comparando los resultados de las investigaciones de contactos en Kingston y Saint Andrew (KSA) con los de la investigación de contactos en dos provincias rurales. MÉTODOS: Los datos de entrevistas y de campo de los asistentes seropositivas a la clínica de atención prenatal para el período comprendido desde octubre de 2004 a septiembre de 2005 en el perímetro urbano KSA, fueron comparados con los de las rurales Clarendon y Pórtland. RESULTADOS: Las mujeres embarazadas VIH seropositivas (n = 88) fueron notificadas y/o referidas a los investigadores de contactos de las provincias: 36 en KSA, 9 en Portland y 43 en Clarendon. El tiempo de la fecha de prueba a la fecha de la entrevista fue casi el doble para KSA (promedio 27 días) en comparación con Pórtland (promedio 15.7 días) y tres veces mayor que el de Clarendon (promedio 9 días). Los tiempos de disposición promedio (cierre de caso) fueron como sigue: KSA, 19 días; Port-land, 28 días; y Clarendon, 15 días. Sólo el 40% de los contactos fueron localizados para KSA y el 48% de estos resultaron VIH positivos a las pruebas. Para Pórtland, 73% fueron localizados y 8% resultaron positivos. Para Clarendon, 45% fueron localizados y 35% de estos resultaron positivos. CONCLUSIONES: No siempre hay pruebas de VIH rápidas disponibles para su realización en el mismo lugar el mismo día, de manera que el investigador de contactos es un miembro esencial del team PMTCT en Jamaica. Uno de los resultados del programa (tiempo de entrevista) tuvo mayor duración en las provincias urbanas que en las rurales, en tanto que otros (tiempo de solución del caso y porcentaje de contactos localizados y sometidos a prueba) no mostraron diferencias consistentes urbano-rurales.

Paediatric head injury - abstract

All paediactric head injury cases admitted to the Port-of-Spain General Hospital, Trinidad in the 12 months from August 1981- July 1982 were studied. There were 306 cases of which 187 (61 percent) were males and 119 (39 percent) females. Causes of injury were falls 221 (72 percent), road traffic accidents 71 (23 percent) other accidents 12(3 percent) and child abuse (2 cases). The commonest clinical features were loss of consciousness 124 (40 percent), nausea and vomiting 114 (37 percent), scalp haematomas 102 (33 percent), scalp lacerations 57 (19 percent), headaches 39 (13 percent), bleeding from ear or nose 19 (6 percent), seizures 18 (6 percent), focal deficits 6 (2 percent). Skull fractures were present in 29 (9 percent). Surgical interventions was necessary in 9 cases. Elevation of compound depressed fractures 3, craniotomy for evacuation of acute subdural haematomas 1 and multiple exploratory burr holes 5 showing cerebral contusions 3, and cerebral oedema 2 cases. Mortality was only 3, all with brain stem injuries. Regarding mobidity, 2 patients had hemiparesis 1 abnormal mental state and 5 continued treatment for seizures. These figures compare well with studies done in UK the United States and Canada regarding sex incidence, causes of injury and symptomatology; however, they differ considerably from the adult head injury in Trinidad where males account for 90 percent, road traffic accidents 59 percent, fights 27 percent, and other causes only 14 percent. These figures also indicate that paediatric head injury demands vigorous treatment despite the initial findings, as the result may be much better than initially predicted (AU)