Optimized antiangiogenic reprogramming of the tumor microenvironment potentiates CD40 immunotherapy.

Cancer immunotherapies are increasingly combined with targeted therapies to improve therapeutic outcomes. We show that combination of agonistic anti-CD40 with antiangiogenic antibodies targeting 2 proangiogenic factors, vascular endothelial growth factor A (VEGFA) and angiopoietin 2 (Ang2/ANGPT2), induces pleiotropic immune mechanisms that facilitate tumor rejection in several tumor models. On the one hand, VEGFA/Ang2 blockade induced regression of the tumor microvasculature while decreasing the proportion of nonperfused vessels and reducing leakiness of the remaining vessels. On the other hand, both anti-VEGFA/Ang2 and anti-CD40 independently promoted proinflammatory macrophage skewing and increased dendritic cell activation in the tumor microenvironment, which were further amplified upon combination of the 2 treatments. Finally, combined therapy provoked brisk infiltration and intratumoral redistribution of cytotoxic CD8+ T cells in the tumors, which was mainly driven by Ang2 blockade. Overall, these nonredundant synergistic mechanisms endowed T cells with improved effector functions that were conducive to more efficient tumor control, underscoring the therapeutic potential of antiangiogenic immunotherapy in cancer.

Regulation of T cell survival through coronin-1-mediated generation of inositol-1,4,5-trisphosphate and calcium mobilization after T cell receptor triggering.

T cell homeostasis is essential for the functioning of the vertebrate immune system, but the intracellular signals required for T cell homeostasis are largely unknown. We here report that the WD-repeat protein family member coronin-1, encoded by the gene Coro1a, is essential in the mouse for T cell survival through its promotion of Ca2+ mobilization from intracellular stores. Upon T cell receptor triggering, coronin-1 was essential for the generation of inositol-1,4,5-trisphosphate from phosphatidylinositol-4,5-bisphosphate. The absence of coronin-1, although it did not affect T cell development, resulted in a profound defect in Ca2+ mobilization, interleukin-2 production, T cell proliferation and T cell survival. We conclude that coronin-1, through activation of Ca2+ release from intracellular stores, is an essential regulator of peripheral lymphocyte survival.

The novel TLR-9 agonist QbG10 shows clinical efficacy in persistent allergic asthma.

BACKGROUND: Allergen-specific TH2 responses contribute to the development of allergic asthma. Their increase may be due to a reduced early exposure to environmental pathogens, which induces a TH1 response, and thereby suppresses the allergic TH2 response. QbG10 (bacteriophage Qbeta-derived virus-like particle with CpG-motif G10 inside), a novel Toll-like receptor 9 agonist packaged into virus-like particles, was designed to stimulate the immune system toward a TH1-mediated protective response. OBJECTIVE: We examined clinical efficacy, safety, and tolerability of QbG10 with patient-reported and objective clinical outcome parameters in patients with mild-to-moderate persistent allergic asthma. METHODS: In this proof-of-concept parallel-group, double-blind, randomized trial, 63 asthmatic patients followed conversion to a standardized inhaled steroid and were treated with 7 injections of either QbG10 or placebo. Incorporating a controlled steroid withdrawal, the effects on patient-reported (day- and nighttime asthma symptoms, salbutamol usage, and 7-item-Asthma Control Questionnaire scores) and objective clinical outcome measures (FEV1, fraction of exhaled nitric oxide, and blood eosinophils) were assessed over 12 weeks (ClinicalTrials.gov number, NCT00890734). RESULTS: All patient-reported parameters improved overall between week 0 and 12 in QbG10-treated patients (n = 33) despite steroid withdrawal, compared with deteriorations observed under placebo (n = 30, P < .05). At week 12, two thirds of the QbG10-treated patients had their asthma "well controlled" (Asthma Control Questionnaire score ≤0.75) compared with one third under placebo. FEV1 had worsened to a clinically significant extent in patients on placebo, while it remained stable in QbG10 patients. Adverse events were mostly injection site reactions occurring after QbG10 administration. CONCLUSION: Treatment with QbG10 may contribute to continued asthma control during steroid reduction in patients on moderate or high-dose inhaled steroids.

Migration and homeostasis of naive T cells depends on coronin 1-mediated prosurvival signals and not on coronin 1-dependent filamentous actin modulation.

Coronins are WD repeat-containing proteins highly conserved in the eukaryotic kingdom implicated in the regulation of F-actin. Mammalian coronin 1, one of the most conserved isoforms expressed in leukocytes, regulates survival of T cells, which has been suggested to be due to its role in preventing F-actin-induced apoptosis. In this study, we come to a different conclusion. We show that coronin 1 does not modulate F-actin and that induction of F-actin failed to induce apoptosis. Instead, coronin 1 was required for providing prosurvival signals, in the absence of which T cells rapidly underwent apoptosis. These results argue against a role for coronin 1 in F-actin-mediated T cell apoptosis and establish coronin 1 as an essential regulator of the balance between prosurvival and proapoptotic signals in naive T cells.

Coronin 1 is essential for IgM-mediated Ca2+ mobilization in B cells but dispensable for the generation of immune responses in vivo.

Coronin 1 is a leukocyte specific regulator of Ca(2+)-dependent signaling and is essential for the survival of peripheral T lymphocytes, but its role in B cells is unknown. In this study, we show that coronin 1 is essential for intracellular Ca(2+) mobilization and proliferation upon triggering of the BCR. However, the presence of costimulatory signals rendered coronin 1 dispensable for B cell signaling, consistent with the generation of normal immune responses against a variety of Ags in coronin 1-deficient mice. We conclude that coronin 1, while being essential for T cell function and survival, is dispensable for B cell function in vivo.

Analysis of Immunological Treatment Effects of Virotherapy in Tumor Tissue.

In addition of being directly tumoricidal, oncolytic viruses have emerged as potent partners for established and investigational immunotherapies due to their immune-stimulatory effects. The shifting focus on virus-mediated immune modulation calls for a comprehensive analysis of the tumor microenvironment (TME) and the factors orchestrating the antiviral and antitumor immune response. The oncolytic VSV-GP studied in our lab is a safe and potent antitumor agent with a fast replication cycle and killing of a broad range of different cancer types. It induces a robust local inflammatory conversion of the TME and drives a strong adaptive immune response toward the tumor. Here we present our multidisciplinary approach to study VSV-GP treatment effects in tumors by assessing both immune cells (tumor-infiltrating lymphocytes and tumor-associated macrophages) and immune-regulatory factors (cytokines) as well as characterizing immune signatures using an immune-targeted NanoString gene expression system.

Evaluating performance, degradation, and release behavior of a nanoform pigmented coating after natural and accelerated weathering.

Pigments with nanoscale dimensions are added to exterior coatings to achieve desirable color and gloss properties. The present study compared the performance, degradation, and release behavior of an acrylic coating that was pigmented by a nanoform of Cu-phthalocyanine after both natural (i.e., outdoor) and accelerated weathering. Samples were weathered outdoors in three geographically distinct locations across the United States (Arizona, Colorado, Maryland) continuously for 15 months. Identically prepared samples were also artificially weathered under accelerated conditions (increased ultraviolet (UV) light intensity and elevated temperatures) for three months, in one-month increments. After exposure, both sets of samples were characterized with color, gloss, and infrared spectroscopy measurements, and selectively with surface roughness measurements. Results indicated that UV-driven coating oxidation was the principal degradation pathway for both natural and accelerated weathering samples, with accelerated weathering leading to an increased rate of oxidation without altering the fundamental degradation pathway. The inclusion of the nanoform pigment reduced the rate of coating oxidation, via UV absorption by the pigment, leading to improved coating integrity compared to non-pigmented samples. Release measurements collected during natural weathering studies indicated there was never a period of weathering, in any location, that led to copper material release above background copper measurements. Lab-based release experiments performed on samples weathered naturally and under accelerated conditions found that the release of degraded coating material after each type of exposure was diminished by the inclusion of the nanoform pigment. Release measurements also indicated that the nanoform pigment remained embedded within the coating and did not release after weathering.

Interference of Mycobacterium tuberculosis with macrophage responses.

Tuberculosis, caused by Mycobacterium tuberculosis, has become an important health and economic burden, with more than four thousand people succumbing to the disease every day. Thus, there is an urgent need to understand the molecular basis of this pathogen's success in causing disease in humans, in order to develop new drugs superior to conventional drugs available at present. One reason why M. tuberculosis is such a dangerous microbe lies within its ability to survive within infected hosts, thereby efficiently circumventing host immune responses. Over the past few years, a number of mechanisms have been unravelled that are utilized by M. tuberculosis to survive within hosts and to avoid immune defence mechanisms. Several of these mechanisms have been described in this communication that may be useful for the development of novel compounds to treat tuberculosis.

Fraternal twins: distinction between PbPc and SnPc by their switching behaviour in a scanning tunnelling microscope.

In this contribution, we compare the optical absorbance behaviour and the structural properties of lead(II)-phthalocyanine (PbPc) and tin(II)-phthalocyanine (SnPc) thin films. To this end, we employ a Ag(1 1 1) substrate terminated with a monolayer of 3,4,9,10-perylene tetracarboxylic dianhydride constituting an internal interface whose main effect is an electronic decoupling of the phthalocyanine adlayer from the metal surface. As deduced from low-energy electron diffraction and scanning tunnelling microscopy (STM) measurements, the epitaxial relations and unit cell compositions of the prevailing PbPc monolayer and multilayer domains are confusingly similar to those of SnPc on PTCDA/Ag(1 1 1). However, SnPc and PbPc can be readily distinguished by their STM-induced switching behaviours: while the former is capable of reversible configurational changes, no effect on the latter could be achieved by us under comparable conditions. This corroborates earlier theoretical predictions and even renders the chemical identification of individual shuttlecock-shaped metal-phthalocyanines feasible.

Adsorption and Detection of Hazardous Trace Gases by Metal-Organic Frameworks.

The quest for advanced designer adsorbents for air filtration and monitoring hazardous trace gases has recently been more and more driven by the need to ensure clean air in indoor, outdoor, and industrial environments. How to increase safety with regard to personal protection in the event of hazardous gas exposure is a critical question for an ever-growing population spending most of their lifetime indoors, but is also crucial for the chemical industry in order to protect future generations of employees from potential hazards. Metal-organic frameworks (MOFs) are already quite advanced and promising in terms of capacity and specific affinity to overcome limitations of current adsorbent materials for trace and toxic gas adsorption. Due to their advantageous features (e.g., high specific surface area, catalytic activity, tailorable pore sizes, structural diversity, and range of chemical and physical properties), MOFs offer a high potential as adsorbents for air filtration and monitoring of hazardous trace gases. Three advanced topics are considered here, in applying MOFs for selective adsorption: (i) toxic gas adsorption toward filtration for respiratory protection as well as indoor and cabin air, (ii) enrichment of hazardous gases using MOFs, and (iii) MOFs as sensors for toxic trace gases and explosives.

Disruption of the cortical actin cytoskeleton does not affect store operated Ca2+ channels in human T-cells.

Lymphocyte signaling and activation leads to the influx of extracellular Ca(2+) via the activation of Ca(2+) release activated Ca(2+) (CRAC) channels in the plasma membrane. Activation of CRAC channels occurs following emptying of the endoplasmic reticulum intracellular Ca(2+) stores. One model to explain the coupling of store-emptying to CRAC activation is the secretion-like conformational coupling model. This model proposes that store depletion increases junctions between the endoplasmic reticulum and the plasma membrane in a manner that could be regulated by the cortical actin cytoskeleton. Here, we show that stabilization or depolymerization of the actin cytoskeleton failed to affect CRAC activation. We therefore conclude that rearrangement of the actin cytoskeleton is dispensable for store-operated Ca(2+) entry in T-cells.

Identification of mycobacterial GarA as a substrate of protein kinase G from M. tuberculosis using a KESTREL-based proteome wide approach.

Signal transduction in bacteria is generally mediated via two-component systems. These systems depend on the transfer of a phosphate molecule from a donor to an acceptor by histidine kinases, thereby activating the acceptor to allow downstream signaling/activation. Several bacterial genomes, including the genome of M. tuberculosis, were shown to encode eukaryotic-like kinases. To better understand the function of these kinases and the regulatory networks within which they operate, identification of downstream targets is essential. We here present a straightforward approach for the identification of bacterial Ser/Thr-kinase substrates. This approach is based on the KESTREL (Kinase Tracking and Substrate Elucidation) procedure combined with reversed-phase chromatography and two-dimensional gel electrophoresis. Using this method, GarA was identified as one potential substrate for the mycobacterial Ser/Thr-protein kinase G (PknG). These results show that the modified KESTREL approach can be successfully employed for the identification of substrates for bacterial Ser/Thr-kinases.

Modulation of macrophage antimicrobial mechanisms by pathogenic mycobacteria.

Tuberculosis remained a mysterious disease until Koch was able to demonstrate in the late 1800s that it was caused by a bacterium spread by aerosols, Mycobacterium tuberculosis. Today, tuberculosis still is a major health problem causing approximately 2 million deaths annually with about one third of the world's population being latently infected with M. tuberculosis. The secret of success for M. tuberculosis lies in its ability to persist inside host cells, the macrophages. Whereas macrophages are designed to destroy any incoming microbe, pathogenic mycobacteria have evolved strategies to survive within macrophages by preventing phagosome-lysosome fusion, thereby creating a niche that allows them to persist within an otherwise hostile environment. In this contribution, we discuss recent advances in our understanding of the interplay between the host and this pathogen that lead to survival of mycobacteria within macrophages.

Does avascular necrosis of the jaws in cancer patients only occur following treatment with bisphosphonates?

INTRODUCTION: In the last decade, bisphosphonates were regularly used to treat osteoporosis and bone pain from diseases such as metastatic breast cancer, multiple myeloma and Paget's disease. Currently, the influence of bisphosphonates in development of avascular osteonecrosis of the jaws has been recognized by various authors. In many cancer patients chemotherapy and medications like steroids have also to be applied. Agreement exists that these drugs can initiate vascular endothelial cell damage and accelerate disturbances in the microcirculation of the jaws possibly resulting in thrombosis of nutrient end arteries. The role of bisphosphonates in cancer patients with previously treated jaws has yet to be elucidated. PATIENTS: Four case reports of 'cancer' patients are described in whom osteonecrosis of the jaws was found. In two patients, the nitrogen-containing bisphosphonate zoledronic acid was prescribed for additional therapy of malignancy for a period of 45 up to 70 months. In another case, supportive treatment of breast cancer was offered using ibandronate. The fourth patient suffered avascular necrosis of the mandible without ever having taken bisphosphonates. In any case, revisional, as well as extended surgery has to be performed for osteonecrosis because neither conservative debridement nor antibiotic therapy have shown long term success, with or without bisphosphonates. No withdrawal of bisphosphonates was performed in view of the information on the direct correlation of total dosage and duration of drug intake to systemic incorporation and the long time for drug release. CONCLUSION: According to our observations, withdrawal of bisphosphonates is not recommended when necrosis of the jaws has occurred.

Immunotherapy of allergic rhinitis: new therapeutic opportunities with virus-like particles filled with CpG motifs.

BACKGROUND: The incidence of allergic rhinitis (AR) has increased constantly over the last decades. The disease can significantly lower quality of life and subsequently might progress to allergic asthma. Allergen-specific immunotherapy is mostly used to cope with the cause of the disease. However, incidence of systemic reactions or limited compliance hampers the widespread use of this therapeutic approach. Therefore, new candidates are examined to improve immunotherapy of allergies. Recently, a new technology was developed with the aim to positively influence the immune system of allergic patients. Virus-like particles (VLPs) represent a potent vaccine platform that has been proven to be immunogenic and clinically effective. To enhance immune cell activation, addition of Toll-like receptor ligands and/or depot-forming adjuvants seems to be helpful. In this context, CpG motifs represent intensive investigated and potent stimulators of T cells. This article focuses on the function of VLPs and CpG motifs and their clinical experience for treatment of AR. METHODS: A literature review was performed. RESULTS: Several published studies showed a beneficial impact of the treatment on allergic symptoms. They tested VLPs filled with or without CpG motifs in combination with or without allergen. CONCLUSION: Results encourage further investigations of VLPs and CpG motifs as adjuncts to or even alternative candidates for immunotherapy of allergic disorders.

Phototriggered growth of crystalline Au structures in the presence of a DNA-surfactant complex.

A method for the preparation of complex anisotropic gold structures with potential uses in catalysis and photonics is presented based on the use of salmon sperm DNA and a photosensitizer to enable light triggered gold salt reduction.

Structural basis for the specific inhibition of protein kinase G, a virulence factor of Mycobacterium tuberculosis.

The pathogenicity of mycobacteria such as Mycobacterium tuberculosis is closely associated with their capacity to survive within host macrophages. A crucial virulence factor for intracellular mycobacterial survival is protein kinase G (PknG), a eukaryotic-like serine/threonine protein kinase expressed by pathogenic mycobacteria that blocks the intracellular degradation of mycobacteria in lysosomes. Inhibition of PknG with the highly selective low-molecular-weight inhibitor AX20017 results in mycobacterial transfer to lysosomes and killing of the mycobacteria. Here, we report the 2.4 A x-ray crystal structure of PknG in complex with AX20017. The unique multidomain topology of PknG reveals a central kinase domain that is flanked by N- and C-terminal rubredoxin and tetratrico-peptide repeat domains, respectively. Directed mutagenesis suggests that the rubredoxin domain functions as a regulator of PknG kinase activity. The structure of PknG-AX20017 further reveals that the inhibitor is buried deep within the adenosine-binding site, targeting an active conformation of the kinase domain. Remarkably, although the topology of the kinase domain is reminiscent of eukaryotic kinases, the AX20017-binding pocket is shaped by a unique set of amino acid side chains that are not found in any human kinase. Directed mutagenesis of the unique set of residues resulted in a drastic loss of the compound's inhibitory potency. Our results explain the specific mode of action of AX20017 and demonstrate that virulence factors highly homologous to host molecules can be successfully targeted to block the proliferation of M. tuberculosis.

Survival of mycobacteria in macrophages is mediated by coronin 1-dependent activation of calcineurin.

Pathogenic mycobacteria survive within macrophages by avoiding lysosomal delivery, instead residing in mycobacterial phagosomes. Upon infection, the leukocyte-specific protein coronin 1 is actively recruited to mycobacterial phagosomes, where it blocks lysosomal delivery by an unknown mechanism. Analysis of macrophages from coronin 1-deficient mice showed that coronin 1 is dispensable for F-actin-dependent processes such as phagocytosis, motility, and membrane ruffling. However, upon mycobacterial infection, coronin 1 was required for activation of the Ca(2+)-dependent phosphatase calcineurin, thereby blocking lysosomal delivery of mycobacteria. In the absence of coronin 1, calcineurin activity did not occur, resulting in lysosomal delivery and killing of mycobacteria. Furthermore, blocking calcineurin activation with cyclosporin A or FK506 led to lysosomal delivery and intracellular mycobacterial killing. These results demonstrate a role for coronin 1 in activating Ca(2+) dependent signaling processes in macrophages and reveal a function for calcineurin in the regulation of phagosome-lysosome fusion upon mycobacterial infection.