Cosmic microwave background anisotropies in multi-connected flat spaces

This article investigates the signature of the seventeen multi-connected flat spaces in cosmic microwave background (CMB) maps. For each such space it recalls a fundamental domain and a set of generating matrices, and then goes on to find an orthonormal basis for the set of eigenmodes of the Laplace operator on that space. The basis eigenmodes are expressed as linear combinations of eigenmodes of the simply connected Euclidean space. A preceding work, which provides a general method for implementing multi-connected topologies in standard CMB codes, is then applied to simulate CMB maps and angular power spectra for each space. Unlike in the 3-torus, the results in most multi-connected flat spaces depend on the location of the observer. This effect is discussed in detail. In particular, it is shown that the correlated circles on a CMB map are generically not back-to-back, so that negative search of back-to-back circles in the WMAP data does not exclude a vast majority of flat or nearly flat topologies.Comment: 33 pages, 19 figures, 1 table. Submitted to PR

A Genome-Wide Association Study of Diabetic Kidney Disease in Subjects With Type 2 Diabetes

dentification of sequence variants robustly associated with predisposition to diabetic kidney disease (DKD) has the potential to provide insights into the pathophysiological mechanisms responsible. We conducted a genome-wide association study (GWAS) of DKD in type 2 diabetes (T2D) using eight complementary dichotomous and quantitative DKD phenotypes: the principal dichotomous analysis involved 5,717 T2D subjects, 3,345 with DKD. Promising association signals were evaluated in up to 26,827 subjects with T2D (12,710 with DKD). A combined T1D+T2D GWAS was performed using complementary data available for subjects with T1D, which, with replication samples, involved up to 40,340 subjects with diabetes (18,582 with DKD). Analysis of specific DKD phenotypes identified a novel signal near GABRR1 (rs9942471, P = 4.5 x 10(-8)) associated with microalbuminuria in European T2D case subjects. However, no replication of this signal was observed in Asian subjects with T2D or in the equivalent T1D analysis. There was only limited support, in this substantially enlarged analysis, for association at previously reported DKD signals, except for those at UMOD and PRKAG2, both associated with estimated glomerular filtration rate. We conclude that, despite challenges in addressing phenotypic heterogeneity, access to increased sample sizes will continue to provide more robust inference regarding risk variant discovery for DKD.Peer reviewe

GAMER-Ad: a novel and rapid method for generating recombinant adenoviruses

Oncolytic adenoviruses have become ideal agents in the path toward treating cancer. Such viruses have been engineered to conditionally replicate in malignant cells in which certain signaling pathways have been disrupted. Other than such oncolytic properties, the viruses need to activate the immune system in order to sustain a long-term response. Therefore, oncolytic adenoviruses have been genetically modified to express various immune-stimulatory agents to achieve this. However, genetically modifying adenoviruses is very time consuming and labor intensive with the current available methods. In this paper, we describe a novel method we have called GAMER-Ad to genetically modify adenovirus genomes within 2 days. Our method entails the replacement of the gp19k gene in the E3 region with any given gene of interest (GOI) using Gibson Assembly avoiding the homologous recombination between the shuttle and the parental plasmid. In this manuscript as proof of concept we constructed and characterized three oncolytic adenoviruses expressing CXCL9, CXCL10, and interleukin-15 (IL-15). We demonstrate that our novel method is fast, reliable, and simple compared to other methods. We anticipate that our method will be used in the future to genetically engineer oncolytic but also other adenoviruses used for gene therapy as well

Novel oncolytic adenovirus expressing enhanced cross-hybrid IgGA Fc PD-L1 inhibitor activates multiple immune effector populations leading to enhanced tumor killing in vitro, in vivo and with patient-derived tumor organoids

Background Despite the success of immune checkpoint inhibitors against PD-L1 in the clinic, only a fraction of patients benefit from such therapy. A theoretical strategy to increase efficacy would be to arm such antibodies with Fc-mediated effector mechanisms. However, these effector mechanisms are inhibited or reduced due to toxicity issues since PD-L1 is not confined to the tumor and also expressed on healthy cells. To increase efficacy while minimizing toxicity, we designed an oncolytic adenovirus that secretes a cross-hybrid Fc-fusion peptide against PD-L1 able to elicit effector mechanisms of an IgG1 and also IgA1 consequently activating neutrophils, a population neglected by IgG1, in order to combine multiple effector mechanisms. Methods The cross-hybrid Fc-fusion peptide comprises of an Fc with the constant domains of an IgA1 and IgG1 which is connected to a PD-1 ectodomain via a GGGS linker and was cloned into an oncolytic adenovirus. We demonstrated that the oncolytic adenovirus was able to secrete the cross-hybrid Fc-fusion peptide able to bind to PD-L1 and activate multiple immune components enhancing tumor cytotoxicity in various cancer cell lines, in vivo and ex vivo renal-cell carcinoma patient-derived organoids. Results Using various techniques to measure cytotoxicity, the cross-hybrid Fc-fusion peptide expressed by the oncolytic adenovirus was shown to activate Fc-effector mechanisms of an IgA1 (neutrophil activation) as well as of an IgG1 (natural killer and complement activation). The activation of multiple effector mechanism simultaneously led to significantly increased tumor killing compared with FDA-approved PD-L1 checkpoint inhibitor (Atezolizumab), IgG1-PDL1 and IgA-PDL1 in various in vitro cell lines, in vivo models and ex vivo renal cell carcinoma organoids. Moreover, in vivo data demonstrated that Ad-Cab did not require CD8+ T cells, unlike conventional checkpoint inhibitors, since it was able to activate other effector populations. Conclusion Arming PD-L1 checkpoint inhibitors with Fc-effector mechanisms of both an IgA1 and an IgG1 can increase efficacy while maintaining safety by limiting expression to the tumor using oncolytic adenovirus. The increase in tumor killing is mostly attributed to the activation of multiple effector populations rather than activating a single effector population leading to significantly higher tumor killing

PeptiCHIP: A Microfluidic Platform for Tumor Antigen Landscape Identification

Identification of HLA class I ligands from the tumor surface (ligandome or immunopeptidome) is essential for designing T-cell mediated cancer therapeutic approaches. However, the sensitivity of the process for isolating MHC-I restricted tumor-specific peptides has been the major limiting factor for reliable tumor antigen characterization, making clear the need for technical improvement. Here, we describe our work from the fabrication and development of a microfluidic-based chip (PeptiCHIP) and its use to identify and characterize tumor-specific ligands on clinically relevant human samples. Specifically, we assessed the potential of immobilizing a pan-HLA antibody on solid surfaces via well-characterized streptavidin-biotin chemistry, overcoming the limitations of the cross-linking chemistry used to prepare the affinity matrix with the desired antibodies in the immunopeptidomics workflow. Furthermore, to address the restrictions related to the handling and the limited availability of tumor samples, we further developed the concept toward the implementation of a microfluidic through-flow system. Thus, the biotinylated pan-HLA antibody was immobilized on streptavidin-functionalized surfaces, and immune-affinity purification (IP) was carried out on customized microfluidic pillar arrays made of thiol-ene polymer. Compared to the standard methods reported in the field, our methodology reduces the amount of antibody and the time required for peptide isolation. In this work, we carefully examined the specificity and robustness of our customized technology for immunopeptidomics workflows. We tested this platform by immunopurifying HLA-I complexes from 1 × 106 cells both in a widely studied B-cell line and in patients-derived ex vivo cell cultures, instead of 5 × 108 cells as required in the current technology. After the final elution in mild acid, HLA-I-presented peptides were identified by tandem mass spectrometry and further investigated by in vitro methods. These results highlight the potential to exploit microfluidics-based strategies in immunopeptidomics platforms and in personalized immunopeptidome analysis from cells isolated from individual tumor biopsies to design tailored cancer therapeutic vaccines. Moreover, the possibility to integrate multiple identical units on a single chip further improves the throughput and multiplexing of these assays with a view to clinical needs