Combined Prebiotic and Microbial Intervention Improves Oral Cholera Vaccination Responses in a Mouse Model of Childhood Undernutrition.

Undernourished children in low-income countries often exhibit poor responses to oral vaccination. Perturbed microbiota development is linked to undernutrition, but whether and how microbiota changes affect vaccine responsiveness remains unclear. Here, we show that gnotobiotic mice colonized with microbiota from undernourished Bangladeshi children and fed a Bangladeshi diet exhibited microbiota-dependent differences in mucosal IgA responses to oral vaccination with cholera toxin (CT). Supplementation with a nutraceutical consisting of spirulina, amaranth, flaxseed, and micronutrients augmented CT-IgA production. Mice initially colonized with a microbiota associated with poor CT responses exhibited improved immunogenicity upon invasion of bacterial taxa from cagemates colonized with a more "responsive" microbiota. Additionally, a consortium of five cultured bacterial invaders conferred augmented CT-IgA responses in mice fed the supplemented diet and colonized with the "hypo-responsive" community. These results provide preclinical proof-of-concept that diet and microbiota influence mucosal immune responses to CT vaccination and identify a candidate synbiotic formulation.

Mechanism of action of mRNA-based vaccines.

INTRODUCTION: The present review summarizes the growing body of work defining the mechanisms of action of this exciting new vaccine technology that should allow rational approaches in the design of next generation mRNA vaccines. Areas covered: Bio-distribution of mRNA, localization of antigen production, role of the innate immunity, priming of the adaptive immune response, route of administration and effects of mRNA delivery systems. Expert commentary: In the last few years, the development of RNA vaccines had a fast growth, the rising number of proof will enable rational approaches to improving the effectiveness and safety of this modern class of medicine.

Gene expression of MAGE-A3 and PRAME tumor antigens and EGFR mutational status in Taiwanese non-small cell lung cancer patients.

AIM: To determine the frequency of expression of the tumor-associated antigens (TAAs) melanoma-associated antigen A3 (MAGE-A3) and preferentially expressed antigen of melanoma (PRAME) and the rate of EGFR mutations in a Taiwanese non-small cell lung cancer (NSCLC) population including only adenocarcinomas and squamous cell carcinomas. Furthermore, to investigate associations between TAA expression and EGFR mutations and to evaluate these TAAs as prognostic markers for overall survival. The occurrence of single nucleotide polymorphisms in MAGEA3 and PRAME was also assessed. METHODS: Archival fresh-frozen tumor tissue specimens were tested by quantitative reverse transcription polymerase chain reaction assays to detect MAGE-A3 and PRAME expression. EGFR mutations were detected by mass spectroscopy and single nucleotide polymorphisms by gene sequencing. RESULTS: Of the 156 adenocarcinomas examined, 3.3% expressed MAGE-A3, 32.2% expressed PRAME and 62.8% had EGFR mutations. Of the 128 squamous cell carcinomas, 29.8% expressed MAGE-A3, 59.2% expressed PRAME and 20.5% harbored EGFR mutations. TAA expression was similar across subgroups determined by patient or tumor characteristics. There was no association between TAA expression and EGFR mutation status and TAA expression was found not to be a prognostic marker for survival. Single nucleotide polymorphisms were identified, one of which with a possible impact on MAGE-A3 expression. CONCLUSIONS: In this NSCLC population, expression of MAGE-A3 and PRAME was more frequent in squamous cell carcinomas than in adenocarcinomas tumors. EGFR mutations were not associated with TAA expression for either histology and were three times more frequent in adenocarcinomas than in squamous cell carcinomas tumors.

Clinical impact of the NKp30/B7-H6 axis in high-risk neuroblastoma patients.

The immunosurveillance mechanisms governing high-risk neuroblastoma (HR-NB), a major pediatric malignancy, have been elusive. We identify a potential role for natural killer (NK) cells, in particular the interaction between the NK receptor NKp30 and its ligand, B7-H6, in the metastatic progression and survival of HR-NB after myeloablative multimodal chemotherapy and stem cell transplantation. NB cells expressing the NKp30 ligand B7-H6 stimulated NK cells in an NKp30-dependent manner. Serum concentration of soluble B7-H6 correlated with the down-regulation of NKp30, bone marrow metastases, and chemoresistance, and soluble B7-H6 contained in the serum of HR-NB patients inhibited NK cell functions in vitro. The expression of distinct NKp30 isoforms affecting the polarization of NK cell functions correlated with 10-year event-free survival in three independent cohorts of HR-NB in remission from metastases after induction chemotherapy (n = 196, P < 0.001), adding prognostic value to known risk factors such as N-Myc amplification and age >18 months. We conclude that the interaction between NKp30 and B7-H6 may contribute to the fate of NB patients and that both the expression of NKp30 isoforms on circulating NK cells and the concentration of soluble B7-H6 in the serum may be clinically useful as biomarkers for risk stratification.

Alternatively spliced NKp30 isoforms affect the prognosis of gastrointestinal stromal tumors.

The natural killer (NK) cell receptor NKp30 is involved in the recognition of tumor and dendritic cells (DCs). Here we describe the influence of three NKp30 splice variants on the prognosis of gastrointestinal sarcoma (GIST), a malignancy that expresses NKp30 ligands and that is treated with NK-stimulatory KIT tyrosine kinase inhibitors. Healthy individuals and those with GIST show distinct patterns of transcription of functionally different NKp30 isoforms. In a retrospective analysis of 80 individuals with GIST, predominant expression of the immunosuppressive NKp30c isoform (over the immunostimulatory NKp30a and NKp30b isoforms) was associated with reduced survival of subjects, decreased NKp30-dependent tumor necrosis factor-α (TNF-α) and CD107a release, and defective interferon-γ (IFN-γ) and interleukin-12 (IL-12) secretion in the NK-DC cross-talk that could be restored by blocking of IL-10. Preferential NKp30c expression resulted partly from a single-nucleotide polymorphism at position 3790 in the 3' untranslated region of the gene encoding NKp30. The genetically determined NKp30 status predicts the clinical outcomes of individuals with GIST independently from KIT mutation.

Contribution of IL-17-producing gamma delta T cells to the efficacy of anticancer chemotherapy.

By triggering immunogenic cell death, some anticancer compounds, including anthracyclines and oxaliplatin, elicit tumor-specific, interferon-γ-producing CD8(+) αß T lymphocytes (Tc1 CTLs) that are pivotal for an optimal therapeutic outcome. Here, we demonstrate that chemotherapy induces a rapid and prominent invasion of interleukin (IL)-17-producing γδ (Vγ4(+) and Vγ6(+)) T lymphocytes (γδ T17 cells) that precedes the accumulation of Tc1 CTLs within the tumor bed. In T cell receptor δ(-/-) or Vγ4/6(-/-) mice, the therapeutic efficacy of chemotherapy was compromised, no IL-17 was produced by tumor-infiltrating T cells, and Tc1 CTLs failed to invade the tumor after treatment. Although γδ T17 cells could produce both IL-17A and IL-22, the absence of a functional IL-17A-IL-17R pathway significantly reduced tumor-specific T cell responses elicited by tumor cell death, and the efficacy of chemotherapy in four independent transplantable tumor models. Adoptive transfer of γδ T cells restored the efficacy of chemotherapy in IL-17A(-/-) hosts. The anticancer effect of infused γδ T cells was lost when they lacked either IL-1R1 or IL-17A. Conventional helper CD4(+) αß T cells failed to produce IL-17 after chemotherapy. We conclude that γδ T17 cells play a decisive role in chemotherapy-induced anticancer immune responses.

Integration of host-related signatures with cancer cell-derived predictors for the optimal management of anticancer chemotherapy.

Current cancer management aims to integrate molecular signatures into the design of personalized therapies. Recent advances in "omics" done on tumor specimens have led to the identification of factors that either recognize cancers of dismal prognosis or pinpoint "druggable" signaling pathways, which can be interrupted by targeted therapies. However, accumulating evidence underscores the biological and clinical significance of immune predictors in several compartments (blood, serum, tumor) in a variety of malignancies. An additional aspect that has been overlooked is the bidirectional, tumor-host interaction during therapeutic intervention, suggesting that dynamic molecular, biochemical, and metabolic signatures should be developed in the future. We review immune parameters of prognostic or predictive value during cancer therapy, and highlight existing "descriptive-prognostic" and "functional-therapeutic" molecular signatures, with the hindsight of designing appropriate compensatory therapies.

Natural killer cell IFN-gamma levels predict long-term survival with imatinib mesylate therapy in gastrointestinal stromal tumor-bearing patients.

Clinical outcomes of gastrointestinal stromal tumor (GIST)-bearing patients treated with imatinib mesylate (IM) are variable. Other than the site of mutation within the c-kit gene, prognostic features of GIST remain undefined. IM can exhibit off-target effects such as triggering natural killer (NK) cell activity. We addressed whether NK cell functions could predict long term survival with IM. NK cell functions were followed up in 77 GIST patients enrolled onto two phase III trials. "Immunologic responders" were defined as patients whose NK cell IFN-gamma values after 2 months of IM were higher than or equal to the baseline value at entry into the trial. The prognostic effect of IFN-gamma on progression-free survival was assessed by a Wald test in a Cox regression analysis using the landmark method and stratified by trial and on the c-kit mutational status. Fifty-six patients were evaluable for the NK cell IFN-gamma responses at baseline and 2 months. Their median follow-up for progression-free survival was 3.7 years. Thirty-four of 56 patients were immunologic responders to IM. In the Cox regression analysis, immunologic responders possessed a hazard ratio of progression or death equal to 0.29 (95% confidence interval, 0.12-0.70; P = 0.006) compared with nonresponders. Kaplan-Meier 2-year survival estimates were 85% for immunologic responders and 50% for nonresponders. Moreover, the immunologic response added prognostic value to the c-kit mutation. The NK cell IFN-gamma production after 2 months of treatment could be considered an independent predictor of long term survival in advanced GISTs treated with IM.

Family-based association of a low producing lymphotoxin-alpha allele with reduced Plasmodium falciparum parasitemia.

Tumor necrosis factor (TNF)-related genes are thought to play a role in human malaria. TNF polymorphisms have been associated with severe malaria, mild malaria, and parasitemia. Lymphotoxin-alpha gene (LTA) that belongs to the TNF family is one such candidate gene. Here we report the family-based association analysis of a cis-regulatory lymphotoxin-alpha polymorphism with parasitemia in two independent populations living in Burkina Faso. Analysis of 199 subjects (34 families) living in an urban endemic area revealed the association of the low producing LTA+80A allele with reduced parasitemia. Furthermore, there was evidence of significant LTA+80-by-age and LTA+80-by-gender interactions. In another set of 318 residents (55 families) in a rural endemic area, we found both the association of the low producing LTA+80A allele with reduced parasitemia and LTA+80-by-age and LTA+80-by-gender interactions. This study suggests that LTA+80 polymorphism influences parasitemia and acts in an age- and gender-dependent manner.

Natural killer cell-directed therapies: moving from unexpected results to successful strategies.

Natural killer (NK) cells influence innate and adaptive immune host defenses. Existing data indicate that manipulating the balance between inhibitory and activating NK receptor signals, the sensitivity of target cells to NK cell-mediated apoptosis, and NK cell cross-talk with dendritic cells might hold therapeutic promise. Efforts to modulate NK cell trafficking into inflamed tissues and/or lymph nodes, and to counteract NK cell suppressors, might also prove fruitful in the clinic. However, deeper investigation into the benefits of combination therapy, greater understanding of the functional distinctions between NK cell subsets, and design of new tools to monitor NK cell activity are needed to strengthen our ability to harness the power of NK cells for therapeutic aims.

Gene expression analysis reveals early changes in several molecular pathways in cerebral malaria-susceptible mice versus cerebral malaria-resistant mice.

BACKGROUND: Microarray analyses allow the identification and assessment of molecular signatures in whole tissues undergoing pathological processes. To better understand cerebral malaria pathogenesis, we investigated intra-cerebral gene-expression profiles in well-defined genetically cerebral malaria-resistant (CM-R) and CM-susceptible (CM-S) mice, upon infection by Plasmodium berghei ANKA (PbA). We investigated mouse transcriptional responses at early and late stages of infection by use of cDNA microarrays. RESULTS: Through a rigorous statistical approach with multiple testing corrections, we showed that PbA significantly altered brain gene expression in CM-R (BALB/c), and in CM-S (CBA/J and C57BL/6) mice, and that 327 genes discriminated between early and late infection stages, between mouse strains, and between CM-R and CM-S mice. We further identified 104, 56, 84 genes with significant differential expression between CM-R and CM-S mice on days 2, 5, and 7 respectively. The analysis of their functional annotation indicates that genes involved in metabolic energy pathways, the inflammatory response, and the neuroprotection/neurotoxicity balance play a major role in cerebral malaria pathogenesis. In addition, our data suggest that cerebral malaria and Alzheimer's disease may share some common mechanisms of pathogenesis, as illustrated by the accumulation of beta-amyloid proteins in brains of CM-S mice, but not of CM-R mice. CONCLUSION: Our microarray analysis highlighted marked changes in several molecular pathways in CM-S compared to CM-R mice, particularly at early stages of infection. This study revealed some promising areas for exploration that may both provide new insight into the knowledge of CM pathogenesis and the development of novel therapeutic strategies.

Association analyses of NCR3 polymorphisms with P. falciparum mild malaria.

Plasmodium falciparum malaria is a major cause of morbidity and mortality in many developing countries especially in sub-Saharan Africa. A susceptibility locus for mild malaria has been mapped to the MHC region, and TNF polymorphisms have been associated with mild malaria. The Natural Cytotoxicity-triggering Receptor 3 (NCR3) gene is located in the peak region of linkage, and is 15kb distal to TNF. In this study, we considered NCR3 as a candidate gene, and we genotyped ten NCR3 single nucleotide polymorphisms (SNPs). Here, we report evidence of an association between mild malaria and NCR3 -412G>C polymorphism located within the promoter. Population-based association analysis showed that NCR3 -412C carriers had more frequent mild malaria attacks than NCR3 -412GG individuals (P=0.001). Using the family-based association test (FBAT) program and its phenotype (PBAT) option, we further found that NCR3 -412C (P=0.0009) and a haplotype containing NCR3 -412C (P=0.008) were significantly associated with increased risk of mild malaria, and that the association was not due to the association of TNF with mild malaria. These observations suggest that there are at least two genes located on the central region of MHC involved in genetic control of human malaria. The association of NCR3 with malaria should provide new insights into the role of Natural Killer cells in this common disease.

Gene-expression profiling discriminates between cerebral malaria (CM)-susceptible mice and CM-resistant mice.

The development of cerebral malaria (CM) in mice with Plasmodium berghei ANKA infection is under genetic control. Brain gene-expression patterns were investigated in well-defined genetically CM-resistant (CM-R; BALB/c and DBA/2) and CM-susceptible (CM-S; C57BL/6 and CBA/J) mice by use of cDNA microarrays. By combining transcriptional profiling with rigorous statistical methods and cluster analysis, we identified a set of 69 genes that perfectly discriminated between mouse strains and between CM-R and CM-S mice. The analysis of gene ontological terms revealed that the genes that clustered and were related to susceptibility to CM preferentially belonged to some biological process classes, such as those pertaining to immune responses. Using a false discovery rate of 5% and the Welch t test, we identified 31 genes with consistent differential expression between CM-R and CM-S mice. These data indicate that microarray analysis may be useful for identification of candidate genes that are potentially responsible for resistance or susceptibility to mouse CM and suggest that candidate genes identified in mice could be specifically tested in humans for an association with disease severity.