4E10-Resistant HIV-1 Isolated from Four Subjects with Rare Membrane-Proximal External Region Polymorphisms

Human antibody 4E10 targets the highly conserved membrane-proximal external region (MPER) of the HIV-1 transmembrane glycoprotein, gp41, and has extraordinarily broad neutralizing activity. It is considered by many to be a prototype for vaccine development. In this study, we describe four subjects infected with viruses carrying rare MPER polymorphisms associated with resistance to 4E10 neutralization. In one case resistant virus carrying a W680G substitution was transmitted from mother to infant. We used site-directed mutagenesis to demonstrate that the W680G substitution is necessary for conferring the 4E10-resistant phenotype, but that it is not sufficient to transfer the phenotype to a 4E10-sensitive Env. Our third subject carried Envs with a W680R substitution causing variable resistance to 4E10, indicating that residues outside the MPER are required to confer the phenotype. A fourth subject possessed a F673L substitution previously associated with 4E10 resistance. For all three subjects with W680 polymorphisms, we observed additional residues in the MPER that co-varied with position 680 and preserved charged distributions across this region. Our data provide important caveats for vaccine development targeting the MPER. Naturally occurring Env variants described in our study also represent unique tools for probing the structure-function of HIV-1 envelope

Effects of a Topically Applied Bioadhesive Berry Gel on Loss of Heterozygosity Indices in Premalignant Oral Lesions

The aim of this study was to assess the effects of topical application of a 10% (w/w) freeze-dried black raspberry (FBR) gel on oral intraepithelial neoplasia (IEN) variables that included histologic diagnoses and loss of heterozygosity (LOH) indices. Microsatellite instability and/or LOH at tumor suppressor gene – associated chromosomal loci have been associated with a higher risk for oral IEN progression to oral squamous cell carcinoma. Previously, our laboratories have shown that FBRs are well tolerated and possess potent antioxidant, apoptotic, and differentiation-inducing properties

A cluster randomised feasibility study of an adolescent incentive intervention to increase uptake of HPV vaccination.

BACKGROUND: Uptake of human papillomavirus (HPV) vaccination is suboptimal among some groups. We aimed to determine the feasibility of undertaking a cluster randomised controlled trial (RCT) of incentives to improve HPV vaccination uptake by increasing consent form return. METHODS: An equal-allocation, two-arm cluster RCT design was used. We invited 60 London schools to participate. Those agreeing were randomised to either a standard invitation or incentive intervention arm, in which Year 8 girls had the chance to win a £50 shopping voucher if they returned a vaccination consent form, regardless of whether consent was provided. We collected data on school and parent participation rates and questionnaire response rates. Analyses were descriptive. RESULTS: Six schools completed the trial and only 3% of parents opted out. The response rate was 70% for the girls' questionnaire and 17% for the parents'. In the intervention arm, 87% of girls returned a consent form compared with 67% in the standard invitation arm. The proportion of girls whose parents gave consent for vaccination was higher in the intervention arm (76%) than the standard invitation arm (61%). CONCLUSIONS: An RCT of an incentive intervention is feasible. The intervention may improve vaccination uptake but a fully powered RCT is needed.British Journal of Cancer advance online publication: 22 August 2017; doi:10.1038/bjc.2017.284 www.bjcancer.com

GDNF Selectively Induces Microglial Activation and Neuronal Survival in CA1/CA3 Hippocampal Regions Exposed to NMDA Insult through Ret/ERK Signalling

The glial cell line-derived neurotrophic factor (GDNF) is a potent survival factor for several neuronal populations in different brain regions, including the hippocampus. However, no information is available on the: (1) hippocampal subregions involved in the GDNF-neuroprotective actions upon excitotoxicity, (2) identity of GDNF-responsive hippocampal cells, (3) transduction pathways involved in the GDNF-mediated neuroprotection in the hippocampus. We addressed these questions in organotypic hippocampal slices exposed to GDNF in presence of N-methyl-D-aspartate (NMDA) by immunoblotting, immunohistochemistry, and confocal analysis. In hippocampal slices GDNF acts through the activation of the tyrosine kinase receptor, Ret, without involving the NCAM-mediated pathway. Both Ret and ERK phosphorylation mainly occurred in the CA3 region where the two activated proteins co-localized. GDNF protected in a greater extent CA3 rather than CA1 following NMDA exposure. This neuroprotective effect targeted preferentially neurons, as assessed by NeuN staining. GDNF neuroprotection was associated with a significant increase of Ret phosphorylation in both CA3 and CA1. Interestingly, confocal images revealed that upon NMDA exposure, Ret activation occurred in microglial cells in the CA3 and CA1 following GDNF exposure. Collectively, this study shows that CA3 and CA1 hippocampal regions are highly responsive to GDNF-induced Ret activation and neuroprotection, and suggest that, upon excitotoxicity, such neuroprotection involves a GDNF modulation of microglial cell activity

Envelope Deglycosylation Enhances Antigenicity of HIV-1 gp41 Epitopes for Both Broad Neutralizing Antibodies and Their Unmutated Ancestor Antibodies

The HIV-1 gp41 envelope (Env) membrane proximal external region (MPER) is an important vaccine target that in rare subjects can elicit neutralizing antibodies. One mechanism proposed for rarity of MPER neutralizing antibody generation is lack of reverted unmutated ancestor (putative naive B cell receptor) antibody reactivity with HIV-1 envelope. We have studied the effect of partial deglycosylation under non-denaturing (native) conditions on gp140 Env antigenicity for MPER neutralizing antibodies and their reverted unmutated ancestor antibodies. We found that native deglycosylation of clade B JRFL gp140 as well as group M consensus gp140 Env CON-S selectively increased the reactivity of Env with the broad neutralizing human mAbs, 2F5 and 4E10. Whereas fully glycosylated gp140 Env either did not bind (JRFL), or weakly bound (CON-S), 2F5 and 4E10 reverted unmutated ancestors, natively deglycosylated JRFL and CON-S gp140 Envs did bind well to these putative mimics of naive B cell receptors. These data predict that partially deglycoslated Env would bind better than fully glycosylated Env to gp41-specific naïve B cells with improved immunogenicity. In this regard, immunization of rhesus macaques demonstrated enhanced immunogenicity of the 2F5 MPER epitope on deglyosylated JRFL gp140 compared to glycosylated JRFL gp140. Thus, the lack of 2F5 and 4E10 reverted unmutated ancestor binding to gp140 Env may not always be due to lack of unmutated ancestor antibody reactivity with gp41 peptide epitopes, but rather, may be due to glycan interference of binding of unmutated ancestor antibodies of broad neutralizing mAb to Env gp41

Genetic Loci Involved in Antibody Response to Mycobacterium avium ssp. paratuberculosis in Cattle

Background: Mycobacterium avium subsp. paratuberculosis (MAP) causes chronic enteritis in a wide range of animal species. In cattle, MAP causes a chronic disease called Johne's disease, or paratuberculosis, that is not treatable and the efficacy of vaccine control is controversial. The clinical phase of the disease is characterised by diarrhoea, weight loss, drop in milk production and eventually death. Susceptibility to MAP infection is heritable with heritability estimates ranging from 0.06 to 0.10. There have been several studies over the last few years that have identified genetic loci putatively associated with MAP susceptibility, however, with the availability of genome-wide high density SNP maker panels it is now possible to carry out association studies that have higher precision. Methodology/Principal Findings: The objective of the current study was to localize genes having an impact on Johne's disease susceptibility using the latest bovine genome information and a high density SNP panel (Illumina BovineSNP50 BeadChip) to perform a case/control, genome-wide association analysis. Samples from MAP case and negative controls were selected from field samples collected in 2007 and 2008 in the province of Lombardy, Italy. Cases were defined as animals serologically positive for MAP by ELISA. In total 966 samples were genotyped: 483 MAP ELISA positive and 483 ELISA negative. Samples were selected randomly among those collected from 119 farms which had at least one positive animal. Conclusion/Significance: The analysis of the genotype data identified several chromosomal regions associated with disease status: a region on chromosome 12 with high significance (P<5 710-6), while regions on chromosome 9, 11, and 12 had moderate significance (P<5 710-5). These results provide evidence for genetic loci involved in the humoral response to MAP. Knowledge of genetic variations related to susceptibility will facilitate the incorporation of this information into breeding programmes for the improvement of health status

Anti-phospholipid human monoclonal antibodies inhibit CCR5-tropic HIV-1 and induce β-chemokines

Traditional antibody-mediated neutralization of HIV-1 infection is thought to result from the binding of antibodies to virions, thus preventing virus entry. However, antibodies that broadly neutralize HIV-1 are rare and are not induced by current vaccines. We report that four human anti-phospholipid monoclonal antibodies (mAbs) (PGN632, P1, IS4, and CL1) inhibit HIV-1 CCR5-tropic (R5) primary isolate infection of peripheral blood mononuclear cells (PBMCs) with 80% inhibitory concentrations of <0.02 to ∼10 µg/ml. Anti-phospholipid mAbs inhibited PBMC HIV-1 infection in vitro by mechanisms involving binding to monocytes and triggering the release of MIP-1α and MIP-1β. The release of these β-chemokines explains both the specificity for R5 HIV-1 and the activity of these mAbs in PBMC cultures containing both primary lymphocytes and monocytes

A Limited Number of Antibody Specificities Mediate Broad and Potent Serum Neutralization in Selected HIV-1 Infected Individuals

A protective vaccine against HIV-1 will likely require the elicitation of a broadly neutralizing antibody (bNAb) response. Although the development of an immunogen that elicits such antibodies remains elusive, a proportion of HIV-1 infected individuals evolve broadly neutralizing serum responses over time, demonstrating that the human immune system can recognize and generate NAbs to conserved epitopes on the virus. Understanding the specificities that mediate broad neutralization will provide insight into which epitopes should be targeted for immunogen design and aid in the isolation of broadly neutralizing monoclonal antibodies from these donors. Here, we have used a number of new and established technologies to map the bNAb specificities in the sera of 19 donors who exhibit among the most potent cross-clade serum neutralizing activities observed to date. The results suggest that broad and potent serum neutralization arises in most donors through a limited number of specificities (1–2 per donor). The major targets recognized are an epitope defined by the bNAbs PG9 and PG16 that is associated with conserved regions of the V1, V2 and V3 loops, an epitope overlapping the CD4 binding site and possibly the coreceptor binding site, an epitope sensitive to a loss of the glycan at N332 and distinct from that recognized by the bNAb 2G12 and an epitope sensitive to an I165A substitution. In approximately half of the donors, key N-linked glycans were critical for expression of the epitopes recognized by the bNAb specificities in the sera

Complete sequence of the 22q11.2 allele in 1,053 subjects with 22q11.2 deletion syndrome reveals modifiers of conotruncal heart defects

The 22q11.2 deletion syndrome (22q11.2DS) results from non-allelic homologous recombination between low-copy repeats termed LCR22. About 60%-70% of individuals with the typical 3 megabase (Mb) deletion from LCR22A-D have congenital heart disease, mostly of the conotruncal type (CTD), whereas others have normal cardiac anatomy. In this study, we tested whether variants in the hemizygous LCR22A-D region are associated with risk for CTDs on the basis of the sequence of the 22q11.2 region from 1,053 22q11.2DS individuals. We found a significant association (FDR p &lt; 0.05) of the CTD subset with 62 common variants in a single linkage disequilibrium (LD) block in a 350 kb interval harboring CRKL. A total of 45 of the 62 variants were associated with increased risk for CTDs (odds ratio [OR) ranges: 1.64-4.75). Associations of four variants were replicated in a meta-analysis of three genome-wide association studies of CTDs in affected individuals without 22q11.2DS. One of the replicated variants, rs178252, is located in an open chromatin region and resides in the double-elite enhancer, GH22J020947, that is predicted to regulate CRKL (CRK-like proto-oncogene, cytoplasmic adaptor) expression. Approximately 23% of patients with nested LCR22C-D deletions have CTDs, and inactivation of Crkl in mice causes CTDs, thus implicating this gene as a modifier. Rs178252 and rs6004160 are expression quantitative trait loci (eQTLs) of CRKL. Furthermore, set-based tests identified an enhancer that is predicted to target CRKL and is significantly associated with CTD risk (GH22J020946, sequence kernal association test (SKAT) p = 7.21&nbsp;× 10-5) in the 22q11.2DS cohort. These findings suggest that variance in CTD penetrance in the 22q11.2DS population can be explained in part by variants affecting CRKL expression

Antibody light-chain-restricted recognition of the site of immune pressure in the RV144 HIV-1 vaccine trial is phylogenetically conserved.

CAPRISA, 2014.Abstract available in pdf