103 research outputs found
In vivo interrogation of regulatory genomes reveals extensive quasi-insufficiency in cancer evolution
No full textAn integrated cellular and molecular model of gastric neuroendocrine cancer evolution highlights therapeutic targets
No full textIL-24 intrinsically regulates Th17 cell pathogenicity in mice
No full textIn certain instances, Th17 responses are associated with severe immunopathology. T cell–intrinsic mechanisms that restrict pathogenic effector functions have been described for type 1 and 2 responses but are less well studied for Th17 cells. Here, we report a cell-intrinsic feedback mechanism that controls the pathogenicity of Th17 cells. Th17 cells produce IL-24, which prompts them to secrete IL-10. The IL-10–inducing function of IL-24 is independent of the cell surface receptor of IL-24 on Th17 cells. Rather, IL-24 is recruited to the inner mitochondrial membrane, where it interacts with the NADH dehydrogenase (ubiquinone) 1 α subcomplex subunit 13 (also known as Grim19), a constituent of complex I of the respiratory chain. Together, Grim19 and IL-24 promote the accumulation of STAT3 in the mitochondrial compartment. We propose that IL-24–guided mitochondrial STAT3 constitutes a rheostat to blunt extensive STAT3 deflections in the nucleus, which might then contribute to a robust IL-10 response in Th17 cells and a restriction of immunopathology in experimental autoimmune encephalomyelitis
The glioblastoma multiforme tumor site promotes the commitment of tumor-infiltrating lymphocytes to the T(H)17 lineage in humans
No full textAlthough glioblastoma multiforme (GBM) is not an invariably cold tumor, checkpoint inhibition has largely failed in GBM. In order to investigate T cell–intrinsic properties that contribute to the resistance of GBM to endogenous or therapeutically enhanced adaptive immune responses, we sorted CD4(+) and CD8(+) T cells from the peripheral blood, normal-appearing brain tissue, and tumor bed of nine treatment-naive patients with GBM. Bulk RNA sequencing of highly pure T cell populations from these different compartments was used to obtain deep transcriptomes of tumor-infiltrating T cells (TILs). While the transcriptome of CD8(+) TILs suggested that they were partly locked in a dysfunctional state, CD4(+) TILs showed a robust commitment to the type 17 T helper cell (T(H)17) lineage, which was corroborated by flow cytometry in four additional GBM cases. Therefore, our study illustrates that the brain tumor environment in GBM might instruct T(H)17 commitment of infiltrating T helper cells. Whether these properties of CD4(+) TILs facilitate a tumor-promoting milieu and thus could be a target for adjuvant anti-T(H)17 cell interventions needs to be further investigated
Limit of blank and limit of detection of Plasmodium falciparum thick blood smear microscopy in a routine setting in Central Africa
No full textA flow cytometry-based workflow for detection and quantification of anti-plasmodial antibodies in vaccinated and naturally exposed individuals
No full textABSTRACT: BACKGROUND: Antibodies play a central role in naturally acquired immunity against Plasmodium falciparum. Current assays to detect anti-plasmodial antibodies against native antigens within their cellular context are prone to bias and cannot be automated, although they provide important information about natural exposure and vaccine immunogenicity. A novel, cytometry-based workflow for the quantitative detection of anti-plasmodial antibodies in human serum is presented. METHODS: Fixed red blood cells (RBCs), infected with late stages of P. falciparum were utilized to detect malaria-specific antibodies by flow cytometry with subsequent automated data analysis. Available methods for data-driven analysis of cytometry data were assessed and a new overlap subtraction algorithm (OSA) based on open source software was developed. The complete workflow was evaluated using sera from two GMZ2 malaria vaccine trials in semiimmune adults and pre-school children residing in a malaria endemic area. RESULTS: Fixation, permeabilization, and staining of infected RBCs were adapted for best operation in flow cytometry. As asexual vaccine candidates are designed to induce antibody patterns similar to semi-immune adults, serial dilutions of sera from heavily exposed individuals were compared to naive controls to determine optimal antibody dilutions. To eliminate investigator effects introduced by manual gating, a non-biased algorithm (OSA) for data-driven gating was developed. OSA derived results correlated well with those obtained by manual gating (r between 0.79 and 0.99) and outperformed other model-driven gating methods. Bland-Altman plots confirmed the agreement of manual gating and OSA derived results. A-1.33 fold increase (p=0.003) in the number of positive cells after vaccination in a subgroup of preschool children vaccinated with 100 mug GMZ2 was present and in vaccinated adults from the same region we measured a baseline-corrected 1.23-fold, vaccine-induced increase in mean fluorescence intensity of positive cells (p=0.03). CONCLUSIONS: The current workflow advances detection and quantification of anti-plasmodial antibodies through improvement of a bias-prone, low-throughput to an unbiased, semi-automated, scalable method. In conclusion, this work presents a novel method for immunofluorescence assays in malaria research
Synergistic targeting and resistance to PARP inhibition in DNA damage repair-deficient pancreatic cancer
No full text© Author(s) (or their employer(s)) 2020.[Objective]: ATM serine/threonine kinase (ATM) is the most frequently mutated DNA damage response gene, involved in homologous recombination (HR), in pancreatic ductal adenocarcinoma (PDAC).[Design]: Combinational synergy screening was performed to endeavour a genotype-tailored targeted therapy.[Results]: Synergy was found on inhibition of PARP, ATR and DNA-PKcs (PAD) leading to synthetic lethality in ATM-deficient murine and human PDAC. Mechanistically, PAD-induced PARP trapping, replication fork stalling and mitosis defects leading to P53-mediated apoptosis. Most importantly, chemical inhibition of ATM sensitises human PDAC cells toward PAD with long-term tumour control in vivo. Finally, we anticipated and elucidated PARP inhibitor resistance within the ATM-null background via whole exome sequencing. Arising cells were aneuploid, underwent epithelial-mesenchymal-transition and acquired multidrug resistance (MDR) due to upregulation of drug transporters and a bypass within the DNA repair machinery. These functional observations were mirrored in copy number variations affecting a region on chromosome 5 comprising several of the upregulated MDR genes. Using these findings, we ultimately propose alternative strategies to overcome the resistance.[Conclusion]: Analysis of the molecular susceptibilities triggered by ATM deficiency in PDAC allow elaboration of an efficient mutation-specific combinational therapeutic approach that can be also implemented in a genotype-independent manner by ATM inhibition.Main funding is provided by the German Cancer Aid grant to AK (111879). Additional funding came from the Deutsche Forschungsgemeinschaft (DFG) ’Sachbeihilfe’ (KL 2544/1–1, 1–2, 5–1, 7-1) and ’Heisenberg-Programm’ (KL 2544/6–1), the Baden-Württemberg-Foundation ExPoChip and the INDIMEDVerbund PancChip. AK, FA, MI, SB, LW and TS are either Principal Investigators or students of HEIST RTG funded by the DFG GRK 2254/1. AK is an Else-KrönerFresenius Excellence fellow. LP received funds by the Bausteinprogramm of Ulm University. PCH is supported by a Max Eder Fellowship of the German Cancer Aid (111746), a German Cancer Aid Priority Program ’Translational Oncology’ 70112505 and by a Collaborative Research Centre grant (316249678 – SFB 1279) of the German Research Foundation. EH received funding from the German Cancer Aid (PiPAC, 70112505) and the Volkswagenstiftung/Ministry for Science and Culture in Lower Saxony (ZN3222). This work was also supported by the Deutsche Forschungsgemeinschaft (AZ.96/1–3) to NA, by the Deutsche Krebshilfe (111264) to AL and by the German Cancer Aid Priority Program Translational Oncology (70112504) to LW
Lactate as a Novel Quantitative Measure of Viability in Schistosoma mansoni Drug Sensitivity Assays
No full textmRNA 3’UTR lengthening by alternative polyadenylation attenuates inflammatory responses and correlates with virulence of Influenza A virus
No full textAbstract Changes of mRNA 3’UTRs by alternative polyadenylation (APA) have been associated to numerous pathologies, but the mechanisms and consequences often remain enigmatic. By combining transcriptomics, proteomics and recombinant viruses we show that all tested strains of IAV, including A/PR/8/34(H1N1) (PR8) and A/Cal/07/2009 (H1N1) (Cal09), cause APA. We mapped the effect to the highly conserved glycine residue at position 184 (G184) of the viral non-structural protein 1 (NS1). Unbiased mass spectrometry-based analyses indicate that NS1 causes APA by perturbing the function of CPSF4 and that this function is unrelated to virus-induced transcriptional shutoff. Accordingly, IAV strain PR8, expressing an NS1 variant with weak CPSF binding, does not induce host shutoff but only APA. However, recombinant IAV (PR8) expressing NS1(G184R) lacks binding to CPSF4 and thereby also the ability to cause APA. Functionally, the impaired ability to induce APA leads to an increased inflammatory cytokine production and an attenuated phenotype in a mouse infection model. Investigating diverse viral infection models showed that APA induction is a frequent ability of many pathogens. Collectively, we propose that targeting of the CPSF complex, leading to widespread alternative polyadenylation of host transcripts, constitutes a general immunevasion mechanism employed by a variety of pathogenic viruses
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