Peptide Ligands Incorporated into the Threefold Spike Capsid Domain to Re-Direct Gene Transduction of AAV8 and AAV9 In Vivo

Efficiency and specificity of viral vectors are vital issues in gene therapy. Insertion of peptide ligands into the adeno-associated viral (AAV) capsid at receptor binding sites can re-target AAV2-derived vectors to alternative cell types. Also, the use of serotypes AAV8 and -9 is more efficient than AAV2 for gene transfer to certain tissues in vivo. Consequently, re-targeting of these serotypes by ligand insertion could be a promising approach but has not been explored so far. Here, we generated AAV8 and -9 vectors displaying peptides in the threefold spike capsid domain. These peptides had been selected from peptide libraries displayed on capsids of AAV serotype 2 to optimize systemic gene delivery to murine lung tissue and to breast cancer tissue in PymT transgenic mice (PymT). Such peptide insertions at position 590 of the AAV8 capsid and position 589 of the AAV9 capsid changed the transduction properties of both serotypes. However, both peptides inserted in AAV8 did not result in the same changes of tissue tropism as they did in AAV2. While the AAV2 peptides selected on murine lung tissue did not alter tropism of serotypes 8 and -9, insertion of the AAV2-derived peptide selected on breast cancer tissue augmented tumor gene delivery in both serotypes. Further, this peptide mediated a strong but unspecific in vivo gene transfer for AAV8 and abrogated transduction of various control tissues for AAV9. Our findings indicate that peptide insertion into defined sites of AAV8 and -9 capsids can change and improve their efficiency and specificity compared to their wild type variants and to AAV2, making these insertion sites attractive for the generation of novel targeted vectors in these serotypes

Phenotypic Detection of Clonotypic B Cells in Multiple Myeloma by Specific Immunoglobulin Ligands Reveals their Rarity in Multiple Myeloma

In multiple myeloma, circulating “clonotypic” B cells, that express the immunoglobulin rearrangement of the malignant plasma cell clone, can be indirectly detected by PCR. Their role as potential “feeder” cells for the malignant plasma cell pool remains controversial. Here we established for the first time an approach that allows direct tracking of such clonotypic cells by labeling with patient-specific immunoglobulin ligands in 15 patients with myeloma. Fifty percent of patients showed evidence of clonotypic B cells in blood or bone marrow by PCR. Epitope-mimicking peptides from random libraries were selected on each patient's individual immunoglobulin and used as ligands to trace cells expressing the idiotypic immunoglobulin on their surface. We established a flow cytometry and immunofluorescence protocol to track clonotypic B cells and validated it in two independent monoclonal B cell systems. Using this method, we found clonotypic B cells in only one out of 15 myeloma patients. In view of the assay's validated sensitivity level of 10−3, this surprising data suggests that the abundance of such cells has been vastly overestimated in the past and that they apparently represent a very rare population in myeloma. Our novel tracing approach may open perspectives to isolate and analyze clonotypic B cells and determine their role in myeloma pathobiology

Successful Expansion but Not Complete Restriction of Tropism of Adeno-Associated Virus by In Vivo Biopanning of Random Virus Display Peptide Libraries

Targeting viral vectors to certain tissues in vivo has been a major challenge in gene therapy. Cell type-directed vector capsids can be selected from random peptide libraries displayed on viral capsids in vitro but so far this system could not easily be translated to in vivo applications. Using a novel, PCR-based amplification protocol for peptide libraries displayed on adeno-associated virus (AAV), we selected vectors for optimized transduction of primary tumor cells in vitro. However, these vectors were not suitable for transduction of the same target cells under in vivo conditions. We therefore performed selections of AAV peptide libraries in vivo in living animals after intravenous administration using tumor and lung tissue as prototype targets. Analysis of peptide sequences of AAV clones after several rounds of selection yielded distinct sequence motifs for both tissues. The selected clones indeed conferred gene expression in the target tissue while gene expression was undetectable in animals injected with control vectors. However, all of the vectors selected for tumor transduction also transduced heart tissue and the vectors selected for lung transduction also transduced a number of other tissues, particularly and invariably the heart. This suggests that modification of the heparin binding motif by target-binding peptide insertion is necessary but not sufficient to achieve tissue-specific transgene expression. While the approach presented here does not yield vectors whose expression is confined to one target tissue, it is a useful tool for in vivo tissue transduction when expression in tissues other than the primary target is uncritical

Synthesis:PLUTONS: Investigating the relationship between pluton growth and volcanism in the Central Andes

The Central Andes is a key global location to study the enigmatic relation between volcanism and plutonism because it has been the site of large ignimbrite-forming eruptions during the past several million years and currently hosts the world’s largest zone of silicic partial melt in the form of the Altiplano-Puna Magma (or Mush) Body (APMB) and the Southern Puna Magma Body (SPMB). In this themed issue, results from the recently completed PLUTONS project are synthesized. This project focused an interdisciplinary study on two regions of large-scale surface uplift that have been found to represent ongoing movement of magmatic fluids in the middle to upper crust. The locations are Uturuncu in Bolivia near the center of the APMB and Lazufre on the Chile-Argentina border, on the edge of the SPMB. These studies use a suite of geological, geochemical, geophysical (seismology, gravity, surface deformation, and electromagnetic methods), petrological, and geomorphological techniques with numerical modeling to infer the subsurface distribution, quantity, and movements of magmatic fluids, as well as the past history of eruptions. Both Uturuncu and Lazufre show separate geophysical anomalies in the upper, middle, and lower crust (e.g., low seismic velocity, low resistivity, etc.) indicating multiple distinct reservoirs of magma and/or hydrothermal fluids with different physical properties. The characteristics of the geophysical anomalies differ somewhat depending on the technique used—reflecting the different sensitivity of each method to subsurface melt (or fluid) of different compositions, connectivity, and volatile content and highlight the need for integrated, multidisciplinary studies. While the PLUTONS project has led to significant progress, many unresolved issues remain and new questions have been raised

Food Sovereignty in the City: Challenging Historical Barriers to Food Justice

Local food initiatives are steadily becoming a part of contemporary cities around the world and can take on many forms. While some of these initiatives are concerned with providing consumers with farm-fresh produce, a growing portion are concerned with increasing the food sovereignty of marginalized urban communities. This chapter provides an analysis of urban contexts with the aim of identifying conceptual barriers that may act as roadblocks to achieving food sovereignty in cities. Specifically, this paper argues that taken for granted commitments created during the birth of the modern city could act as conceptual barriers for the implementation of food sovereignty programs and that urban food activists and programs that challenge these barriers are helping to achieve the goal of restoring food sovereignty to local communities, no matter their reasons for doing so. At the very least, understanding the complexities of these barriers and how they operate helps to strengthen ties between urban food projects, provides these initiatives with ways to undermine common arguments used to support restrictive ordinances and policies, and illustrates the transformative potential of food sovereignty movements

Non-affirmative Theory of Education as a Foundation for Curriculum Studies, Didaktik and Educational Leadership

This chapter presents non-affirmative theory of education as the foundation for a new research program in education, allowing us to bridge educational leadership, curriculum studies and Didaktik. We demonstrate the strengths of this framework by analyzing literature from educational leadership and curriculum theory/didaktik. In contrast to both socialization-oriented explanations locating curriculum and leadership within existing society, and transformation-oriented models viewing education as revolutionary or super-ordinate to society, non-affirmative theory explains the relation between education and politics, economy and culture, respectively, as non-hierarchical. Here critical deliberation and discursive practices mediate between politics, culture, economy and education, driven by individual agency in historically developed cultural and societal institutions. While transformative and socialization models typically result in instrumental notions of leadership and teaching, non-affirmative education theory, previously developed within German and Nordic education, instead views leadership and teaching as relational and hermeneutic, drawing on ontological core concepts of modern education: recognition; summoning to self-activity and Bildsamkeit. Understanding educational leadership, school development and teaching then requires a comparative multi-level approach informed by discursive institutionalism and organization theory, in addition to theorizing leadership and teaching as cultural-historical and critical-hermeneutic activity. Globalisation and contemporary challenges to deliberative democracy also call for rethinking modern nation-state based theorizing of education in a cosmopolitan light. Non-affirmative education theory allows us to understand and promote recognition based democratic citizenship (political, economical and cultural) that respects cultural, ethical and epistemological variations in a globopolitan era. We hope an American-European-Asian comparative dialogue is enhanced by theorizing education with a non-affirmative approach

The Volcanic Evolution of Cerro Uturuncu: A High-K, Composite Volcano in the Back-Arc of the Central Andes of SW Bolivia

Cerro Uturuncu, southwest Bolivia, is a high-K, calc-alkaline, composite volcano constructed upon extremely thick continental crust approximately 125 km behind the arc-front of the Andean Central Volcanic Zone (CVZ). Eruptive activity occurred between 890 - 271 ka in a single phase of volcanism lasting ~620,000 years. The edifice consists of a central cone and several flank vents where dacitic and andesitic lava flows and domes erupted. Volumes of individual eruptive units range from 0.1 to ~10 km3; the composite volume of Uturuncu is ~89 km3. In this paper, we present new field, petrographic, and geochemical data in an effort to understand the volcanic and magmatic evolution of Uturuncu. Lava flows and domes have a restricted range in whole rock compositions ranging from 61 wt% - 67 wt% SiO2; magmatic inclusions contained within these units have a larger range from 53 wt% - 64 wt% SiO2. Typical phenocryst assemblages are plagioclase \u3e orthopyroxene \u3e biotite \u3e\u3e quartz and Fe-Ti oxides. Pb isotope ratios are characteristic of the southern CVZ by containing high 207Pb/204Pb and 206Pb/204Pb and moderate to high 208Pb/204Pb. Sr and Nd isotope ratios indicate that Uturuncu magmas were modified by high 87Sr/86Sr and low 143Nd/144Nd felsic basement lithology during magma migration and differentiation. In all eruptive units, there is petrographic and geochemical evidence for magma mixing and mingling. In this regard, magma mixing and mingling is considered to be responsible for the small range in lava flow and dome compositions throughout the eruptive history of the center

Interstitial lung disease increases mortality in systemic sclerosis patients with pulmonary arterial hypertension without affecting hemodynamics and exercise capacity

Published data suggest that coexisting interstitial lung disease (ILD) has an impact on mortality in patients with systemic sclerosis (SSc) and pulmonary arterial hypertension (PAH), but there is scarce knowledge if this is reflected by hemodynamics, exercise capacity, autoantibody profile, or pulmonary function. In this partially retrospective study, 27 SSc-PAH patients were compared to 24 SSc-PAH patients with coexisting ILD respecting to survival, pulmonary function, hemodynamics, exercise capacity, and laboratory parameters. Survival was significantly worse in SSc-PAH-ILD patients than in SSc patients with isolated PAH (1, 5, and 10-year survival rates 86, 54, and 54% versus 96, 92, and 82%, p = 0.013). Compared to isolated SSc-PAH patients, patients with SSc-PAH-ILD revealed lower forced expiratory volume after 1 s (FEV1) values at the time of PAH diagnosis as well as 1 and 2 years later (p = 0.002) without significant decrease in the PAH course in both groups. At PAH diagnosis, diffusion capacity for carbon monoxide (DLCO) values were lower in the ILD-PAH group. Coexisting ILD was not associated with lower exercise capacity, different FEV1/forced vital capacity (FVC) ratio, higher WHO functional class, or reduced hemodynamics. Higher levels of antibodies against angiotensin and endothelin receptors predict mortality in all SSc-PAH patients but could not differentiate between PAH patients with and without ILD. Our study confirmed an impact of ILD on mortality in SSc-PAH patients. Pulmonary function parameters can be used to distinguish PAH from PAH-ILD. The higher mortality rate cannot be explained by differences in hemodynamics, exercise capacity, or autoantibody levels. Mechanisms of mortality remain to be studied