Commensal bacteria augment Staphylococcus aureus infection by inactivation of phagocyte-derived reactive oxygen species

Staphylococcus aureus is a human commensal organism and opportunist pathogen, causing potentially fatal disease. The presence of non-pathogenic microflora or their components, at the point of infection, dramatically increases S. aureus pathogenicity, a process termed augmentation. Augmentation is associated with macrophage interaction but by a hitherto unknown mechanism. Here, we demonstrate a breadth of cross-kingdom microorganisms can augment S. aureus disease and that pathogenesis of Enterococcus faecalis can also be augmented. Co-administration of augmenting material also forms an efficacious vaccine model for S. aureus. In vitro, augmenting material protects S. aureus directly from reactive oxygen species (ROS), which correlates with in vivo studies where augmentation restores full virulence to the ROS-susceptible, attenuated mutant katA ahpC. At the cellular level, augmentation increases bacterial survival within macrophages via amelioration of ROS, leading to proliferation and escape. We have defined the molecular basis for augmentation that represents an important aspect of the initiation of infection

Vaccination of stage III/IV melanoma patients with long NY-ESO-1 peptide and CpG-B elicits robust CD8(+) and CD4(+) T-cell responses with multiple specificities including a novel DR7-restricted epitope.

Long synthetic peptides and CpG-containing oligodeoxynucleotides are promising components for cancer vaccines. In this phase I trial, 19 patients received a mean of 8 (range 1-12) monthly vaccines s.c. composed of the long synthetic NY-ESO-179-108 peptide and CpG-B (PF-3512676), emulsified in Montanide ISA-51. In 18/18 evaluable patients, vaccination induced antigen-specific CD8(+) and CD4(+) T-cell and antibody responses, starting early after initiation of immunotherapy and lasting at least one year. The T-cells responded antigen-specifically, with strong secretion of IFNγ and TNFα, irrespective of patients' HLAs. The most immunogenic regions of the vaccine peptide were NY-ESO-189-102 for CD8(+) and NY-ESO-183-99 for CD4(+) T-cells. We discovered a novel and highly immunogenic epitope (HLA-DR7/NY-ESO-187-99); 7/7 HLA-DR7(+) patients generated strong CD4(+) T-cell responses, as detected directly ex vivo with fluorescent multimers. Thus, vaccination with the long synthetic NY-ESO-179-108 peptide combined with the strong immune adjuvant CpG-B induced integrated, robust and functional CD8(+) and CD4(+) T-cell responses in melanoma patients, supporting the further development of this immunotherapeutic approach

Immunoregulation of Dendritic Cell Subsets by Inhibitory Receptors in Urothelial Cancer.

Blockade of inhibitory receptors (IRs) overexpressed by T cells can activate antitumor immune responses, resulting in the most promising therapeutic approaches, particularly in bladder cancer, currently able to extend patient survival. Thanks to their ability to cross-present antigens to T cells, dendritic cells (DCs) are an immune cell population that plays a central role in the generation of effective antitumor T-cell responses. While IR function and expression have been investigated in T cells, very few data are available for DCs. Therefore, we analyzed whether DCs express IRs that can decrease their functions. To this end, we investigated several IRs (PD-1, CTLA-4, BTLA, TIM-3, and CD160) in circulating CD1c javax.xml.bind.JAXBElement@4f1331d4 DCs, CD141 javax.xml.bind.JAXBElement@68e4feef DCs, and plasmacytoid DCs from healthy donors and patients with urothelial cancer (UCa). Different DC subsets expressed BTLA and TIM-3 but not other IRs. More importantly, BTLA and TIM-3 were significantly upregulated in DCs from blood of UCa patients. Locally, bladder tumor-infiltrating DCs also overexpressed BTLA and TIM-3 compared to DCs from paired nontumoral tissue. Finally, in vitro functional experiments showed that ligand-mediated engagement of BTLA and TIM-3 receptors significantly reduced the secretion of effector cytokines by DC subpopulations. Our findings demonstrate that UCa induces local and systemic overexpression of BTLA and TIM-3 by DCs that may result in their functional inhibition, highlighting these receptors as potential targets for UCa treatment. We investigated the expression and function of a panel of inhibitory receptors in dendritic cells (DCs), an immune cell subpopulation critical in initiation of protective immune responses, among patients with urothelial carcinoma. We found high expression of BTLA and TIM-3 by blood and tumor DCs, which could potentially mediate decreased DC function. The results suggest that BTLA and TIM-3 might be new targets for urothelial carcinoma treatment

Adenosine mediates functional and metabolic suppression of peripheral and tumor-infiltrating CD8⁺ T cells.

Several mechanisms are present in the tumor microenvironment (TME) to impair cytotoxic T cell responses potentially able to control tumor growth. Among these, the accumulation of adenosine (Ado) contributes to tumor progression and represents a promising immunotherapeutic target. Ado has been shown to impair T cell effector function, but the role and mechanisms employed by Ado/Ado receptors (AdoRs) in modulating human peripheral and tumor-infiltrating lymphocyte (TIL) function are still puzzling. CD8 <sup>+</sup> T cell cytokine production following stimulation was quantified by intracellular staining and flow cytometry. The cytotoxic capacity of tumor infiltrating lymphocytes (TILs) was quantified by the chromium release assay following co-culture with autologous or anti-CD3-loaded tumor cell lines. The CD8 <sup>+</sup> T cell metabolic fitness was evaluated by the seahorse assay and by the quantification of 2-NBDG uptake and CD71/CD98 upregulation upon stimulation. The expression of AdoRs was assessed by RNA flow cytometry, a recently developed technology that we validated by semiquantitative RT-PCR (qRT-PCR), while the impact on T cell function was evaluated by the use of selective antagonists and agonists. The influence of Ado/AdoR on the PKA and mTOR pathways was evaluated by phosphoflow staining of p-CREB and p-S6, respectively, and validated by western blot. Here, we demonstrate that Ado signaling through the A2A receptor (A2AR) in human peripheral CD8 <sup>+</sup> T cells and TILs is responsible for the higher sensitivity to Ado-mediated suppression of T central memory cells. We confirmed that Ado is able to impair peripheral and tumor-expanded T cell effector functions, and we show for the first time its impact on metabolic fitness. The Ado-mediated immunosuppressive effects are mediated by increased PKA activation that results in impairment of the mTORC1 pathway. Our findings unveil A2AR/PKA/mTORC1 as the main Ado signaling pathway impairing the immune competence of peripheral T cells and TILs. Thus, p-CREB and p-S6 may represent useful pharmacodynamic and efficacy biomarkers of immunotherapies targeting Ado. The effect of Ado on T cell metabolic fitness reinforces the importance of the adenosinergic pathway as a target for next-generation immunotherapy

First Observation and Measurement of the Decay K+- -> pi+- e+ e- gamma

Using the full data set of the NA48/2 experiment, the decay K+- -> pi+- e+ e- gamma is observed for the first time, selecting 120 candidates with 7.3 +- 1.7 estimated background events. With K+- -> pi+- pi0D as normalisation channel, the branching ratio is determined in a model-independent way to be Br(K+- -> pi+- e+ e- gamma, m_eegamma > 260 MeV/c^2) = (1.19 +- 0.12_stat +- 0.04_syst) x 10^-8. This measured value and the spectrum of the e+ e- gamma invariant mass allow a comparison with predictions of Chiral Perturbation Theory.Comment: 13 pages, 3 figures. Accepted for publication in Phys.Lett.

Empirical parameterization of the K+- -> pi+- pi0 pi0 decay Dalitz plot

As first observed by the NA48/2 experiment at the CERN SPS, the \p0p0 invariant mass (M00) distribution from \kcnn decay shows a cusp-like anomaly at M00=2m+, where m+ is the charged pion mass. An analysis to extract the pi pi scattering lengths in the isospin I=0 and I=2 states, a0 and a2, respectively, has been recently reported. In the present work the Dalitz plot of this decay is fitted to a new empirical parameterization suitable for practical purposes, such as Monte Carlo simulations of K+- -> pi+- pi0 pi0 decays.Comment: 9 pages, 3 figures

Search for direct CP-violation in K+- --> pi+-pi0pi0 decays

A search for direct CP-violation in K+- --> pi+-pi0pi0 decays based on 47.14 million events has been performed by the NA48/2 experiment at the CERN SPS. The asymmetry in the Dalitz plot linear slopes A_g=(g^+ - g^-)/(g^+ + g^-) is measured to be A_g=(1.8 +- 2.6).10^{-4}. The design of the experiment and the method of analysis provide good control of instrumental charge asymmetries in this measurement. The precision of the result is limited by statistics and is almost one order of magnitude better than that of previous measurements by other experiments.Comment: 14 page

ChPT tests at the NA48 and NA62 experiments at CERN

The NA48/2 Collaboration at CERN has accumulated unprecedented statistics of rare kaon decays in the Ke4 modes: Ke4(+-) ($K^\pm \to \pi^+ \pi^- e^\pm \nu$) and Ke4(00) ($K^\pm \to \pi^0 \pi^0 e^\pm \nu$) with nearly one percent background contamination. The detailed study of form factors and branching rates, based on these data, has been completed recently. The results brings new inputs to low energy strong interactions description and tests of Chiral Perturbation Theory (ChPT) and lattice QCD calculations. In particular, new data support the ChPT prediction for a cusp in the $\pi^0\pi^0$ invariant mass spectrum at the two charged pions threshold for Ke4(00) decay. New final results from an analysis of about 400 $K^\pm \to \pi^\pm \gamma \gamma$ rare decay candidates collected by the NA48/2 and NA62 experiments at CERN during low intensity runs with minimum bias trigger configurations are presented. The results include a model-independent decay rate measurement and fits to ChPT description.Comment: XIIth International Conference on Heavy Quarks and Leptons 2014, Mainz, German

Tumour-derived PGD2 and NKp30-B7H6 engagement drives an immunosuppressive ILC2-MDSC axis.

Group 2 innate lymphoid cells (ILC2s) are involved in human diseases, such as allergy, atopic dermatitis and nasal polyposis, but their function in human cancer remains unclear. Here we show that, in acute promyelocytic leukaemia (APL), ILC2s are increased and hyper-activated through the interaction of CRTH2 and NKp30 with elevated tumour-derived PGD2 and B7H6, respectively. ILC2s, in turn, activate monocytic myeloid-derived suppressor cells (M-MDSCs) via IL-13 secretion. Upon treating APL with all-trans retinoic acid and achieving complete remission, the levels of PGD2, NKp30, ILC2s, IL-13 and M-MDSCs are restored. Similarly, disruption of this tumour immunosuppressive axis by specifically blocking PGD2, IL-13 and NKp30 partially restores ILC2 and M-MDSC levels and results in increased survival. Thus, using APL as a model, we uncover a tolerogenic pathway that may represent a relevant immunosuppressive, therapeutic targetable, mechanism operating in various human tumour types, as supported by our observations in prostate cancer.Group 2 innate lymphoid cells (ILC2s) modulate inflammatory and allergic responses, but their function in cancer immunity is still unclear. Here the authors show that, in acute promyelocytic leukaemia, tumour-activated ILC2s secrete IL-13 to induce myeloid-derived suppressor cells and support tumour growth