Systemically Administered TLR7/8 Agonist and Antigen-Conjugated Nanogels Govern Immune Responses against Tumors

[Image: see text] The generation of specific humoral and cellular immune responses plays a pivotal role in the development of effective vaccines against tumors. Especially the presence of antigen-specific, cytotoxic T cells influences the outcome of therapeutic cancer vaccinations. Different strategies, ranging from delivering antigen-encoding mRNAs to peptides or full antigens, are accessible but often suffer from insufficient immunogenicity and require immune-boosting adjuvants as well as carrier platforms to ensure stability and adequate retention. Here, we introduce a pH-responsive nanogel platform as a two-component antitumor vaccine that is safe for intravenous application and elicits robust immune responses in vitro and in vivo. The underlying chemical design allows for straightforward covalent attachment of a model antigen (ovalbumin) and an immune adjuvant (imidazoquinoline-type TLR7/8 agonist) onto the same nanocarrier system. In addition to eliciting antigen-specific T and B cell responses that outperform mixtures of individual components, our two-component nanovaccine leads in prophylactic and therapeutic studies to an antigen-specific growth reduction of different tumors expressing ovalbumin intracellularly or on their surface. Regarding the versatile opportunities for functionalization, our nanogels are promising for the development of highly customized and potent nanovaccines

Pointlike structure for super p-branes

We present an efficient method to understand the p-brane dynamics in a unified framework. For this purpose, we reformulate the action for super p-branes in the form appropriate to incorporate the pointlike (parton) structure of higher dimensional p-branes and intend to interpret the p-brane dynamics as the collective dynamics of superparticles. In order to examine such a parton picture of super p-branes, we consider various superparticle configurations that can be reduced from super p-branes, especially, a supermembrane, and study the partonic structure of classical p-brane solutions.Comment: 22 pages, corrected typos, to appear in Phys. Rev. D58, 085018 (1998

The Heat Shock Response of Mycobacterium Tuberculosis: Linking Gene Expression, Immunology and Pathogenesis

The regulation of heat shock protein (HSP) expression is critically important to pathogens such as Mycobacterium tuberculosis and dysregulation of the heat shock response results in increased immune recognition of the bacterium and reduced survival during chronic infection. In this study we use a whole genome spotted microarray to characterize the heat shock response of M. tuberculosis. We also begin a dissection of this important stress response by generating deletion mutants that lack specific transcriptional regulators and examining their transcriptional profiles under different stresses. Understanding the stimuli and mechanisms that govern heat shock in mycobacteria will allow us to relate observed in vivo expression patterns of HSPs to particular stresses and physiological conditions. The mechanisms controlling HSP expression also make attractive drug targets as part of a strategy designed to enhance immune recognition of the bacterium

Candida albicans Possesses Sap7 as a Pepstatin A-Insensitive Secreted Aspartic Protease

BACKGROUND: Candida albicans, a commensal organism, is a part of the normal flora of healthy individuals. However, once the host immunity is compromised, C. albicans opportunistically causes recurrent superficial or fatal systemic candidiasis. Secreted aspartic proteases (Sap), encoded by 10 types of SAP genes, have been suggested to contribute to various virulence processes. Thus, it is important to elucidate their biochemical properties for better understanding of the molecular mechanisms that how Sap isozymes damage host tissues. METHODOLOGY/PRINCIPAL FINDINGS: The SAP7 gene was cloned from C. albicans SC5314 and heterogeneously produced by Pichia pastoris. Measurement of Sap7 proteolytic activity using the FRETS-25Ala library showed that Sap7 was a pepstatin A-insensitive protease. To understand why Sap7 was insensitive to pepstatin A, alanine substitution mutants of Sap7 were constructed. We found that M242A and T467A mutants had normal proteolytic activity and sensitivity to pepstatin A. M242 and T467 were located in close proximity to the entrance to an active site, and alanine substitution at these positions widened the entrance. Our results suggest that this alteration might allow increased accessibility of pepstatin A to the active site. This inference was supported by the observation that the T467A mutant has stronger proteolytic activity than the wild type. CONCLUSIONS/SIGNIFICANCE: We found that Sap7 was a pepstatin A-insensitive protease, and that M242 and T467 restricted the accessibility of pepstatin A to the active site. This finding will lead to the development of a novel protease inhibitor beyond pepstatin A. Such a novel inhibitor will be an important research tool as well as pharmaceutical agent for patients suffering from candidiasis

Candida albicans Scavenges Host Zinc via Pra1 during Endothelial Invasion

The ability of pathogenic microorganisms to assimilate essential nutrients from their hosts is critical for pathogenesis. Here we report endothelial zinc sequestration by the major human fungal pathogen, Candida albicans. We hypothesised that, analogous to siderophore-mediated iron acquisition, C. albicans utilises an extracellular zinc scavenger for acquiring this essential metal. We postulated that such a “zincophore” system would consist of a secreted factor with zinc-binding properties, which can specifically reassociate with the fungal cell surface. In silico analysis of the C. albicans secretome for proteins with zinc binding motifs identified the pH-regulated antigen 1 (Pra1). Three-dimensional modelling of Pra1 indicated the presence of at least two zinc coordination sites. Indeed, recombinantly expressed Pra1 exhibited zinc binding properties in vitro. Deletion of PRA1 in C. albicans prevented fungal sequestration and utilisation of host zinc, and specifically blocked host cell damage in the absence of exogenous zinc. Phylogenetic analysis revealed that PRA1 arose in an ancient fungal lineage and developed synteny with ZRT1 (encoding a zinc transporter) before divergence of the Ascomycota and Basidiomycota. Structural modelling indicated physical interaction between Pra1 and Zrt1 and we confirmed this experimentally by demonstrating that Zrt1 was essential for binding of soluble Pra1 to the cell surface of C. albicans. Therefore, we have identified a novel metal acquisition system consisting of a secreted zinc scavenger (“zincophore”), which reassociates with the fungal cell. Furthermore, functional similarities with phylogenetically unrelated prokaryotic systems indicate that syntenic zinc acquisition loci have been independently selected during evolution

A case of mistaken identity: HSPs are no DAMPs but DAMPERs

Until recently, the immune system was seen solely as a defense system with its primary task being the elimination of unwanted microbial invaders. Currently, however, the functional significance of the immune system has obtained a much wider perspective, to include among others the maintenance and restoration of homeostasis following tissue damage. In this latter aspect, there is a growing interest in the identification of molecules involved, such as the so-called danger or damage-associated molecular patterns (DAMPs), also called alarmins. Since heat shock proteins are archetypical molecules produced under stressful conditions, such as tissue damage or inflammation, they are frequently mentioned as prime examples of DAMPs (Bianchi, J Leukoc Biol 81:1–5, 2007; Kono and Rock, Nat Rev Immunol 8:279–289, 2008; Martin-Murphy et al., Toxicol Lett 192:387–394, 2010). See for instance also a recent review (Chen and Nunez, Science 298:1395–1401, 2010). Contrary to this description, we recently presented some of the arguments against a role of heat shock protein as DAMPs (Broere et al., Nat Rev Immunol 11:565-c1, 2011). With this perspective and reflection article, we hope to elaborate on this debate and provide additional thoughts to further ignite this discussion on this critical and evolving issue

Inclusive e+^+e−^- production in collisions of pions with protons and nuclei in the second resonance region of baryons

Inclusive e$^+$e$^-$ production has been studied with HADES in $\pi^-$ + p, $\pi^-$ + C and $\pi^- + \mathrm{CH}_2$ reactions, using the GSI pion beam at $\sqrt{s_{\pi p}}$ = 1.49 GeV. Invariant mass and transverse momentum distributions have been measured and reveal contributions from Dalitz decays of $\pi^0$, $\eta$ mesons and baryon resonances. The transverse momentum distributions are very sensitive to the underlying kinematics of the various processes. The baryon contribution exhibits a deviation up to a factor seven from the QED reference expected for the dielectron decay of a hypothetical point-like baryon with the production cross section constrained from the inverse $\gamma$ n$\rightarrow \pi^-$ p reaction. The enhancement is attributed to a strong four-momentum squared dependence of the time-like electromagnetic transition form factors as suggested by Vector Meson Dominance (VMD). Two versions of the VMD, that differ in the photon-baryon coupling, have been applied in simulations and compared to data. VMD1 (or two-component VMD) assumes a coupling via the $\rho$ meson and a direct coupling of the photon, while in VMD2 (or strict VMD) the coupling is only mediated via the $\rho$ meson. The VMD2 model, frequently used in transport calculations for dilepton decays, is found to overestimate the measured dielectron yields, while a good description of the data can be obtained with the VMD1 model assuming no phase difference between the two amplitudes. Similar descriptions have also been obtained using a time-like baryon transition form factor model where the pion cloud plays the major role.Comment: (HADES collaboration

Impact of the Coulomb field on charged-pion spectra in few-GeV heavy-ion collisions

In nuclear collisions the incident protons generate a Coulomb field which acts on produced charged particles. The impact of these interactions on charged-pion transverse-mass and rapidity spectra, as well as on pion–pion momentum correlations is investigated in Au + Au collisions at $\sqrt{^{S}NN}$ = 2.4 GeV. We show that the low-m$_{t}$ region (m$_{t}$ < 0.2 GeV / c$^{2}$) can be well described with a Coulomb-modified Boltzmann distribution that also takes changes of the Coulomb field during the expansion of the fireball into account. The observed centrality dependence of the fitted mean Coulomb potential energy deviates strongly from a $A_{part}^{2/3}$ scaling, indicating that, next to the fireball, the non-interacting charged spectators have to be taken into account. For the most central collisions, the Coulomb modifications of the HBT source radii are found to be consistent with the potential extracted from the single-pion transverse-mass distributions. This finding suggests that the region of homogeneity obtained from two-pion correlations coincides with the region in which the pions freeze-out. Using the inferred mean-square radius of the charge distribution at freeze-out, we have deduced a baryon density, in fair agreement with values obtained from statistical hadronization model fits to the particle yields