Decreased Levels of Bisecting GlcNAc Glycoforms of IgG Are Associated with Human Longevity

BACKGROUND: Markers for longevity that reflect the health condition and predict healthy aging are extremely scarce. Such markers are, however, valuable in aging research. It has been shown previously that the N-glycosylation pattern of human immunoglobulin G (IgG) is age-dependent. Here we investigate whether N-linked glycans reflect early features of human longevity. METHODOLOGY/PRINCIPAL FINDINGS: The Leiden Longevity Study (LLS) consists of nonagenarian sibling pairs, their offspring, and partners of the offspring serving as control. IgG subclass specific glycosylation patterns were obtained from 1967 participants in the LLS by MALDI-TOF-MS analysis of tryptic IgG Fc glycopeptides. Several regression strategies were applied to evaluate the association of IgG glycosylation with age, sex, and longevity. The degree of galactosylation of IgG decreased with increasing age. For the galactosylated glycoforms the incidence of bisecting GlcNAc increased as a function of age. Sex-related differences were observed at ages below 60 years. Compared to males, younger females had higher galactosylation, which decreased stronger with increasing age, resulting in similar galactosylation for both sexes from 60 onwards. In younger participants (<60 years of age), but not in the older age group (>60 years), decreased levels of non-galactosylated glycoforms containing a bisecting GlcNAc reflected early features of longevity. CONCLUSIONS/SIGNIFICANCE: We here describe IgG glycoforms associated with calendar age at all ages and the propensity for longevity before middle age. As modulation of IgG effector functions has been described for various IgG glycosylation features, a modulatory effect may be expected for the longevity marker described in this study

Systematic Evaluation of Normalization Methods for Glycomics Data Based on Performance of Network Inference

Glycomics measurements, like all other high-throughput technologies, are subject to technical variation due to fluctuations in the experimental conditions. The removal of this non-biological signal from the data is referred to as normalization. Contrary to other omics data types, a systematic evaluation of normalization options for glycomics data has not been published so far. In this paper, we assess the quality of different normalization strategies for glycomics data with an innovative approach. It has been shown previously that Gaussian Graphical Models (GGMs) inferred from glycomics data are able to identify enzymatic steps in the glycan synthesis pathways in a data-driven fashion. Based on this finding, here, we quantify the quality of a given normalization method according to how well a GGM inferred from the respective normalized data reconstructs known synthesis reactions in the glycosylation pathway. The method therefore exploits a biological measure of goodness. We analyzed 23 different normalization combinations applied to six large-scale glycomics cohorts across three experimental platforms: Liquid Chromatography – ElectroSpray Ionization-Mass Spectrometry (LC-ESI-MS), Ultra High Performance Liquid Chromatography with Fluorescence Detection (UHPLC-FLD), and Matrix Assisted Laser Desorption Ionization – Furier Transform Ion Cyclotron Resonance – Mass Spectrometry (MALDI-FTICR-MS). Based on our results, we recommend normalizing glycan data using the ‘Probabilistic Quotient’ method followed by log-transformation, irrespective of the measurement platform. This recommendation is further supported by an additional analysis, where we ranked normalization methods based on their statistical associations with age, a factor known to associate with glycomics measurements

The Antioxidant Protein Peroxiredoxin 4 Is Epigenetically Down Regulated in Acute Promyelocytic Leukemia

The antioxidant peroxiredoxin (PRDX) protein family comprises 6 members, which are implicated in a variety of cellular responses, including growth factor signal transduction. PRDX4 resides in the endoplasmic reticulum (ER), where it locally controls oxidative stress by reducing H2O2levels. We recently provided evidence for a regulatory function of PRDX4 in signal transduction from a myeloid growth factor receptor, the granulocyte colony-stimulating factor receptor (G-CSFR). Upon activation, the ligand-induced G-CSFR undergoes endocytosis and routes via the early endosomes where it physically interacts with ER-resident PRDX4. PRDX4 negatively regulates G-CSFR mediated signaling. Here, we investigated whether PRDX4 is affected in acute myeloid leukemia (AML); genomic alterations and expression levels of PRDX4 were investigated. We show that genomic abnormalities involving PRDX4 are rare in AML. However, we find a strong reduction in PRDX4 expression levels in acute promyelocytic leukemia (APL) compared to normal promyelocytes and different molecular subtypes of AML. Subsequently, the possible role of DNA methylation and histone modifications in silencing of PRDX4 in APLs was investigated. We show that the reduced expression is not due to methylation of the CpG island in the promoter region of PRDX4 but correlates with increased trimethylation of histone 3 lysine residue 27 (H3K27me3) and lysine residue 4 (H3K4me3) at the transcriptional start site (TSS) of PRDX4, indicative of a bivalent histone code involved in transcriptional silencing. These findings suggest that the control of G-CSF responses by the antioxidant protein PRDX4 may be perturbed in APL

G-CSF and its receptor in myeloid malignancy

Granulocyte colony-stimulating factor (G-CSF) has been used in the clinic for more than 2 decades to treat congenital and acquired neutropenias and to reduce febrile neutropenia before or during courses of intensive cytoreductive therapy. In addition, healthy stem cell donors receive short-term treatment with G-CSF for mobilization of hematopoietic stem cells. G-CSF has also been applied in priming strategies designed to enhance the sensitivity of leukemia stem cells to cytotoxic agents, in protocols aimed to induce their differentiation and accompanying growth arrest and cell death, and in severe aplastic anemia and myelodysplastic syndrome (MDS) to alleviate anemia. The potential adverse effects of G-CSF administration, particularly the risk of malignant transformation, have fueled ongoing debates, some of which can only be settled in follow-up studies extending over several decades. This specifically applies to children with severe congenital neutropenia who receive lifelong treatment with G-CSF and in which the high susceptibility to develop MDS and acute myeloid leukemia (AML) has now become a major clinical concern. Here, we will highlight some of the controversies and challenges regarding the clinical application of G-CSF and discuss a possible role of G-CSF in malignant transformation, particularly in patients with neutropenia harboring mutations in the gene encoding the G-CSF receptor. (Blood. 2010; 115(25):5131-5136

Classification for Longevity Potential: The Use of Novel Biomarkers

Background: In older people, chronological age may not be the best predictor of residual lifespan and mortality, because with age the heterogeneity in health is increasing. Biomarkers for biological age and residual lifespan are being developed to predict disease and mortality better at an individual level than chronological age. In the current paper, we aim to classify a group of older people into those with longevity potential or controls. Methods: In the Leiden Longevity Study participated 1671 offspring of nonagenarian siblings, as the group with longevity potential, and 744 similarly aged controls. Using known risk factors for cardiovascular disease, previously reported markers for human longevity and other physiological measures as predictors, classification models for longevity potential were constructed with multiple logistic regression of the offspring-control status. Results: The Framingham Risk Score (FRS) is predictive for longevity potential [area under the receiver operating characteristic curve (AUC) = 64.7]. Physiological parameters involved in immune responses and glucose, lipid and energy metabolism further improve the prediction performance for longevity potential (AUCmale = 71.4, AUCfemale = 68.7). Conclusion: Using the FRS, the classification of older people in groups with longevity potential and controls is moderate, but can be improved to a reasonably good classification in combination with markers of immune response, glucose, lipid, and energy metabolism. We show that individual classification of older people for longevity potential may be feasible using biomarkers from a wide variety of different biological processes

Sequential gain of mutations in severe congenital neutropenia progressing to acute myeloid leukemia

Severe congenital neutropenia (SCN) is a BM failure syndrome with a high risk of progression to acute myeloid leukemia (AML). The underlying genetic changes involved in SCN evolution to AML are largely unknown. We obtained serial hematopoietic samples from an SCN patient who developed AML 17 years after the initiation of G-CSF treatment. Next-generation sequencing was performed to identify mutations during disease progression. In the AML phase, we found 12 acquired nonsynonymous mutations. Three of these, in CSF3R, LLGL2, and ZC3H18, co-occurred in a subpopulation of progenitor cells already in the early SCN phase. This population expanded over time, whereas clones harboring only CSF3R mutations disappeared from the BM. The other 9 mutations were only apparent in the AML cells and affected known AML-associated genes (RUNX1 and ASXL1) and chromatin remodelers (SUZ12 and EP300). In addition, a novel CSF3R mutation that conferred autonomous proliferation to myeloid progenitors was found. We conclude that progression from SCN to AML is a multistep process, with distinct mutations arising early during the SCN phase and others later in AML development. The sequential gain of 2 CSF3R mutations implicates abnormal G-CSF signaling as a driver of leukemic transformation in this case of SCN. (Blood. 2012; 119(22): 5071-5077

Reduced PRDX4 protein and transcript levels in APL.

<p><b>A</b>. Lysates of APL and AML samples were analyzed for PRDX4 protein levels. Because human <i>PRDX4</i> gene is located on the X-chromosome, male samples and female samples were compared with male and female AML control samples respectively to rule out the possibility of X-inactivation involvement. Patient number followed by F indicates female sample while patient number followed by M indicates male sample. <b>B. Upper panel</b>: Western blot analysis of PRDX4 expression in different normal bone marrow fractions (myeloblasts, promyelocytes, neutrophils). <b>Lower panel</b>: Micrographs of May Grunwald Giemsa stained fractions of cells used for Western blotting.</p