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In this study, we investigated the role of the Wnt/β-catenin signaling pathway in antitumor immune responses. We report that the concentration of secreted Wnt3a was significantly higher in conditioned medium from tumor or nontumor tissues obtained from all hepatocellular carcinoma or colorectal cancer patients tested, than in serum of healthy donors or patients. In addition, both Wnt3a and β-catenin were overexpressed by tumor-infiltrating and nontumor-infiltrating CD4+ or CD8+ T cells. The majority of these T cells expressed a dysfunctional effector memory Eomes+T-bet−phenotype that we defined as partially exhausted, because they performed effector functions (in terms of interferon-γ and tumor necrosis factor-α production, as well as CD107a mobilization) despite their PD-1 expression. Wnt3a/β-catenin signaling in T naïve cells in vitro recapitulated the T-cell setting in vivo. Indeed, the differentiation of cultured T naïve cells was arrested, producing cells that resembled the EomeshighT-betlowβ-cateninhigh T cells with moderate effector functions that infiltrated tumor and nontumor areas. Wnt3a blockade improved the capacity of T naïve cells to differentiate into effector cells in vitro. However, Wnt3a blockade did not affect the function and phenotype of differentiated, partially exhausted, tumor-infiltrating T cells ex vivo. Taken together, our data suggest that Wnt3a blockade halts the capacity of Wnt/β-catenin signaling to inhibit the differentiation of T naïve cells, but it does not restore the dysfunction of differentiated T cells, in the tumor setting. Cancer Immunol Res; 6(8); 941–52. ©2018 AACR.

Table S1. Characteristics of all enrolled patients (HCC=35, CRC=16), not previous treated with chemotherapy Table S2. List of all antibodies for FC analysis Figure S1: CD4 and CD8 T cells expressed high levels of PD1 in TUM district and showed mostly an effector memory phenotype in CRC Figure S2: Distribution of Wnt3a in serum, NTUM- or TUM-derived CM, and expression of Wnt3a/β-catenin axis in NTUM and TUM-infiltrating CD4 and CD8 T cells from CRC patients Figure S3: Analysis of Frizzled receptor 1 expression among CD4 and CD8 T cells in periphery, nontumor and tumor samples of HCC patients Figure S4: β-catenin+ tumor-infiltrating CD4 T cells in presence of anti-Wnt3a do not down-regulated TNF-α prduction

In our study, we investigated the role of CD39 on tumor-infiltrating CD8

In our study, we investigated the role of CD39 on tumor-infiltrating CD8+ T lymphocytes (CD8+ TILs) in colorectal, head and neck and pancreatic cancers. Partially confirming recent observations correlating the CD39 expression with T-cell exhaustion, we demonstrated a divergent functional activity in CD39+ CD8+ TILs. On the one hand, CD39+ CD8+ TILs (as compared to their CD39- counterparts) produced significantly lower IFN-γ and IL-2 amounts, expressed higher PD-1, and inversely correlated with perforin and granzyme B expression. On the other, they displayed a significantly higher proliferative capacity ex vivo that was inversely correlated with the PD-1 expression. Therefore, CD39+ CD8+ TILs, including those co-expressing the CD103 (a marker of T resident memory [TRM] cells), were defined as partially dysfunctional T cells that correlate with tumor patients with initial progression stages. Interestingly, our results identified for the first time a single nucleotide polymorphism (SNP rs10748643 A>G), as a genetic factor associated with CD39 expression in CD8+ TILs. Finally, we demonstrated that compounds inhibiting CD39-related ATPases improved CD39+ CD8+ T-cell effector function ex vivo, and that CD39+ CD8+ TILs displayed effective suppression function in vitro. Overall these data suggest that the SNP analysis may represent a suitable predictor of CD39+ CD8+ T-cell expression in cancer patients, and propose the modulation of CD39 as a new strategy to restore partially exhausted CD8+ TILs.

In this study, we investigated the role of the Wnt/β-catenin signaling pathway in antitumor immune responses. We report that the concentration of secreted Wnt3a was significantly higher in conditioned medium from tumor or nontumor tissues obtained from all hepatocellular carcinoma or colorectal cancer patients tested, than in serum of healthy donors or patients. In addition, both Wnt3a and β-catenin were overexpressed by tumor-infiltrating and nontumor-infiltrating CD4+ or CD8+ T cells. The majority of these T cells expressed a dysfunctional effector memory Eomes+T-bet−phenotype that we defined as partially exhausted, because they performed effector functions (in terms of interferon-γ and tumor necrosis factor-α production, as well as CD107a mobilization) despite their PD-1 expression. Wnt3a/β-catenin signaling in T naïve cells in vitro recapitulated the T-cell setting in vivo. Indeed, the differentiation of cultured T naïve cells was arrested, producing cells that resembled the EomeshighT-betlowβ-cateninhigh T cells with moderate effector functions that infiltrated tumor and nontumor areas. Wnt3a blockade improved the capacity of T naïve cells to differentiate into effector cells in vitro. However, Wnt3a blockade did not affect the function and phenotype of differentiated, partially exhausted, tumor-infiltrating T cells ex vivo. Taken together, our data suggest that Wnt3a blockade halts the capacity of Wnt/β-catenin signaling to inhibit the differentiation of T naïve cells, but it does not restore the dysfunction of differentiated T cells, in the tumor setting. Cancer Immunol Res; 6(8); 941–52. ©2018 AACR.

Background In Africa, peanuts are frequently consumed, but severe allergic reactions are rare. We investigated immunological patterns of clinical tolerance to peanut in peanut-sensitized but asymptomatic patients from central Africa compared to peanut-allergic and peanut-sensitized but asymptomatic patients from Sweden. Methods Sera from allergic patients (n = 54) from Zimbabwe sensitized to peanut but without allergic symptoms to peanut, and sera from peanut-allergic (n = 25) and peanut-sensitized but asymptomatic (n = 25) patients from Sweden were analyzed toward peanut allergen components (Ara h 1–3, 6, 8–9) and other allergen molecules from important allergen sources using microarray. IgE to Ara h 2 peptide epitopes was analyzed, and allergenic activity was assessed by basophil activation assay. Results Forty-six percent of the African and all peanut-allergic Swedish patients showed IgE toward one of the highly allergenic peanut allergens (Ara h 1–3, 6, 9). However, 48% of the African patients had IgE to cross-reactive carbohydrate determinants (CCDs) with low allergenic activity and 60% of the Swedish asymptomatic patients had IgE against the PR protein Ara h 8. IgG and IgG4 specificities and levels could not discriminate between the African asymptomatic and Swedish peanut-allergic patients. Asymptomatic patients almost lacked IgE to Ara h 2 peptides, which were recognized by peanut-allergic patients. Peanut IgE from peanut asymptomatic patients showed poor allergenic activity compared with IgE from peanut-allergic patients. Conclusions Natural clinical tolerance to peanut in the African patients can be caused by IgE to low allergenic peanut components and by poor allergenic activity of peanut-specific IgE.

Background: Rhinoviruses (RVs) are a major cause of common colds and induce exacerbations of asthma and chronic inflammatory lung diseases. Methods: We expressed and purified recombinant RV coat proteins VP1-4, non-structural proteins as well as N-terminal fragments of VP1 from four RV strains (RV14, 16, 89, C) covering the three known RV groups (RV-A, RV-B and RV-C) and measured specific IgG-subclass-, IgA- and IgM-responses by ELISA in subjects with different severities of asthma or without asthma before and after experimental infection with RV16. Findings: Before infection subjects showed IgG1 > IgA > IgM > IgG3 cross-reactivity with N-terminal fragments from the representative VP1 proteins of the three RV groups. Antibody levels were higher in the asthmatic group as compared to the non-asthmatic subjects. Six weeks after infection with RV16, IgG1 antibodies showed a group-specific increase towards the N-terminal VP1 fragment, but not towards other capsid and non-structural proteins, which was highest in subjects with severe upper and lower respiratory symptoms. Interpretation: Our results demonstrate that increases of antibodies towards the VP1 N-terminus are group-specific and associated with severity of respiratory symptoms and suggest that it may be possible to develop serological tests for identifying causative RV groups.

IL-18 is an inflammasome-related cytokine, member of the IL-1 family, produced by a wide range of cells in response to signals by several pathogen- or damage-associated molecular patterns. It can be highly represented in tumor patients, but its relevance in human cancer development is not clear. In this study, we provide evidence that IL-18 is principally expressed in tumor cells and, in concert with other conventional Th1 cell-driven cytokines, has a pivotal role in establishing a pro-inflammatory milieu in the tumor microenvironment of human non-small cell lung cancer (NSCLC). Interestingly, the analysis of tumor-infiltrating CD8+ T cell populations showed that (i) the relative IL-18 receptor (IL-18R) is significantly more expressed by the minority of cells with a functional phenotype (T-bet+Eomes+), than by the majority of those with the dysfunctional phenotype T-bet−Eomes+ generally resident within tumors; (ii) as a consequence, the former are significantly more responsive than the latter to IL-18 stimulus in terms of IFNγ production ex vivo; (iii) PD-1 expression does not discriminate these two populations. These data indicate that IL-18R may represent a biomarker of the minority of functional tumor-infiltrating CD8+ T cells in adenocarcinoma NSCLC patients. In addition, our results lead to envisage the possible therapeutic usage of IL-18 in NSCLC, even in combination with other checkpoint inhibitor approaches.

We used the microarray technology to develop chips containing a comprehensive set of proteins and peptides covering the proteome of HIV-1 clade C, which is the HIV-1 subtype that causes the majority of infections worldwide. We demonstrate that the HIV microarray allows simultaneous, sensitive and specific detection of antibody responses for the major immunoglobulin classes (IgG, IgA, IgM, IgE) and subclasses (IgG1-4) with minute amounts of serum samples towards a large number of HIV antigens and peptides. Furthermore, we show that the HIV chip can be used for the monitoring of antibody responses during the course of the disease and during treatment. The HIV microarray should be useful to study antibody responses to multiple HIV antigens and epitopes in HIV-infected patients to explore pathomechanisms of the disease, for diagnosis and for monitoring of treatment and of vaccine trials.

The infection of CD4+ cells by HIV leads to the progressive destruction of CD4+ T lymphocytes and, after a severe reduction of CD4+ cells, to AIDS. The aim of the study was to investigate whether HIV-infected patients with CD4 cell counts

Correction for 'HIV microarray for the mapping and characterization of HIV-specific antibody responses' by Daniela Gallerano et al., Lab Chip, 2015, 15, 1574-1589.

HIV-1 virus infectivity factor (Vif) is one of the four accessory proteins that are characteristic of primate lentiviruses and critically required for the infection of host cells. Vif plays a key role in replication and transmission of the virus in non-permissive cells, such as primary T cells and macrophages. Using co-precipitation and co-fractionation techniques, evidence has been provided that Vif interacts with a variety of host proteins, such as the cytidine deaminases APOBEC3G and 3F, the Cullin5/EloBC ubiquitin–ligase complex, Fyn and Hck tyrosine kinases, as well as with viral components, such as the immature Gag precursor and viral RNA. We report on the expression, purification and molecular characterization of a folded recombinant subtype C Vif. Vif was expressed in E. coli with a C-terminal hexahistidine tag and purified by nickel affinity chromatography. We obtained approximately 5 mg protein per liter of bacterial culture, with a purity >95%. The expected molecular mass of 23.7 kDa was confirmed by mass spectrometry. Although dynamic light scattering and small angle X-ray scattering measurements revealed the presence of high molecular weight aggregates in the protein preparation, circular dichroism analysis showed that the protein contains mainly folded β-sheet elements and exhibits remarkable thermal stability (Tm > 95°C). Recombinant Vif may be used as a tool to study its biological functions and tertiary structure, as well as for the development of diagnostic, therapeutic and preventive strategies for HIV-1 infections.