AMYCNE: Confident copy number assessment using whole genome sequencing data.

Copy number variations (CNVs) within the human genome have been linked to a diversity of inherited diseases and phenotypic traits. The currently used methodology to measure copy numbers has limited resolution and/or precision, especially for regions with more than 4 copies. Whole genome sequencing (WGS) offers an alternative data source to allow for the detection and characterization of the copy number across different genomic regions in a single experiment. A plethora of tools have been developed to utilize WGS data for CNV detection. None of these tools are designed specifically to accurately estimate copy numbers of complex regions in a small cohort or clinical setting. Herein, we present AMYCNE (automatic modeling functionality for copy number estimation), a CNV analysis tool using WGS data. AMYCNE is multifunctional and performs copy number estimation of complex regions, annotation of VCF files, and CNV detection on individual samples. The performance of AMYCNE was evaluated using AMY1A ddPCR measurements from 86 unrelated individuals. In addition, we validated the accuracy of AMYCNE copy number predictions on two additional genes (FCGR3A and FCGR3B) using datasets available through the 1000 genomes consortium. Finally, we simulated levels of mosaic loss and gain of chromosome X and used this dataset for benchmarking AMYCNE. The results show a high concordance between AMYCNE and ddPCR, validating the use of AMYCNE to measure tandem AMY1 repeats with high accuracy. This opens up new possibilities for the use of WGS for accurate copy number determination of other complex regions in the genome in small cohorts or single individuals.

Comprehensive Assessment of the Association between FCGRs polymorphisms and the risk of systemic lupus erythematosus: Evidence from a Meta-Analysis.

We performed a meta analysis to assess the relationship of FCGRs polymorphisms with the risk of SLE. Thirty-five articles (including up to 5741 cases and 6530 controls) were recruited for meta-analysis. The strongest association was observed between FCGR2B rs1050501 and SLE under the recessive genotypic model of C allele in the overall population (CC vs CT/TT, OR = 1.754, 95%CI: 1.422-2.165, P = 1.61 × 10(-7)) and in Asian population (CC vs CT/TT, OR = 1.784, 95%CI; 1.408-2.261, P = 1.67 × 10(-6)). We also found that FCGR3A rs396991 were significant association with the susceptibility to SLE in overall population in recessive model of T allele (TT vs TG/GG, OR = 1.263, 95%CI: 1.123-1.421, P = 9.62 × 10(-5)). The results also showed that significant association between FCGR2A rs1801274 and SLE under the allelic model in the overall population (OR = 0.879 per A allele, 95%CI: 0.819-0.943, P = 3.31 × 10(-4)). The meta-analysis indicated that FCGR3B copy number polymorphism NA1·NA2 was modestly associated with SLE in overall population (OR = 0.851 per NA1, 95%CI: 0.772-0.938, P = 1.2 × 10(-3)). We concluded that FCGR2B rs1050501 C allele and FCGR3A rs396991 T allele might contribute to susceptibility and development of SLE, and were under recessive association model. While, FCGR2A rs1801274 A allele and FCGR3B NA1 were associated with SLE and reduced the risk of SLE.

Hardy-Weinberg equilibrium revisited for inferences on genotypes featuring allele and copy-number variations.

Copy number variations represent a substantial source of genetic variation and are associated with a plethora of physiological and pathophysiological conditions. Joint copy number and allelic variations (CNAVs) are difficult to analyze and require new strategies to unravel the properties of genotype distributions. We developed a Bayesian hidden Markov model (HMM) approach that allows dissecting intrinsic properties and metastructures of the distribution of CNAVs within populations, in particular haplotype phases of genes with varying copy numbers. As a key feature, this approach incorporates an extension of the Hardy-Weinberg equilibrium, allowing both a comprehensive and parsimonious model design. We demonstrate the quality of performance and applicability of the HMM approach with a real data set describing the Fcγ receptor (FcγR) gene region. Our concept, using a dynamic process to analyze a static distribution, establishes the basis for a novel understanding of complex genomic data sets.

Assessment of the Influence of Inflammation and FCGR3A Genotype on Infliximab Pharmacokinetics and Time to Relapse in Patients with Crohn's Disease.

BACKGROUND AND OBJECTIVES: Infliximab is a monoclonal anti-tumor necrosis factor-α (anti-TNFα) antibody that profoundly modified the treatment of Crohn's disease (CD). The polymorphism of Fc fragment of IgG, low affinity IIIa, receptor (CD16a) [FCGR3A] influences the biological response to infliximab in patients with CD. Our aim was to study its influence on infliximab pharmacokinetics and risk of relapse after infliximab discontinuation. METHODS: In 111 CD patients in remission, infliximab was discontinued and its concentrations were measured for 30 months or until relapse. Infliximab pharmacokinetics were described using monocompartmental population modeling. RESULTS: The elimination rate of infliximab increased with C-reactive protein (CRP) [p = 0.00018] and was 16 % higher in FCGR3A-158V/V patients than in F carriers (p = 0.0028). Risk of relapse was higher in patients with baseline CRP ≥5 mg/L than in those with a lower value (p = 0.0000029). In addition, there was a first-order interaction between CRP and the FCGR3A genotype; in patients with high CRP, risk of relapse was higher for V/V patients than for F carriers (hazard ratio 4.80 and 2.84 for V/V and F carriers, respectively; p = 0.013). CONCLUSION: Both increased inflammation and FCGR3A-158V/V genotype are associated with increased infliximab elimination and risk of relapse after infliximab discontinuation in patients with CD.

The role of natural killer (NK) cells and NK cell receptor polymorphisms in the assessment of HIV-1 neutralization.

The importance of innate immune cells in HIV-1 pathogenesis and protection has been highlighted by the role of natural killer (NK) cells in the containment of viral replication. Use of peripheral blood mononuclear cells (PBMC) in immunologic studies provides both HIV-1 target cells (ie. CD4+ T cells), as well as anti-HIV-1 effector cells, such as NK cells. In this study, NK and other immune cell populations were analyzed in HIV-negative donor PBMC for an impact on the anti-HIV activity of polyclonal and monoclonal antibodies. NK cell percentages were significantly higher in donor PBMC that supported lower levels of viral replication. While the percentage of NK cells was not directly associated with neutralization titers, NK cell-depletion significantly diminished the antiviral antibody activity by up to three logs, and polymorphisms in NK killer immunoglobulin receptor (KIR) and FcγRIIIa alleles appear to be associated with this affect. These findings demonstrate that NK cells and NK cell receptor polymorphisms may influence assessment of traditional HIV-1 neutralization in a platform where antibody is continuously present. This format appears to simultaneously assess conventional entry inhibition (neutralization) and non-neutralizing antibody-dependent HIV inhibition, which may provide the opportunity to delineate the dominant antibody function(s) in polyclonal vaccine responses.

Association of FcgR2a, but not FcgR3a, with inflammatory bowel diseases across three Caucasian populations.

BACKGROUND: The Fc receptors II and III (FcgR2a, and FcgR3a) play a crucial role in the regulation of the immune response. The FcgR2a*519GG and FcgR3a*559CC genotypes have been associated with several autoimmune diseases including systemic lupus erythematosus, rheumatoid arthritis, nephritis, and possibly to type I diabetes, and celiac disease. In a large multicenter, two-stage study of 6570 people, we tested whether the FcgR2a and FcgR3a genes were also involved in inflammatory bowel disease (IBD), which includes Crohn's disease (CD) and ulcerative colitis (UC). METHODS: We genotyped the FcgR2a*A519G and FcgR3a*A559C functional variants in 4205 IBD patients in six well-phenotyped Caucasian IBD cohorts and 2365 ethnically matched controls recruited from the Netherlands, Spain, and New Zealand. RESULTS: In the initial Dutch study we found a significant association of FcgR2a genotypes with IBD (P-genotype = 0.02); while the FcgR2a*519GG was more common in controls (23%) than in IBD patients (18%; odds ratio [OR] = 0.75; 95% confidence interval [CI] 0.61-0.92; P = 0.004). This association was corroborated by a combined analysis across all the study populations (Mantel-Haenszel [MH] OR = 0.84; 0.74-0.95; P = 0.005) in the next stage. The Fcgr2a*GG genotype was associated with both UC (MH-OR = 0.84; 0.72-0.97; P = 0.01) and CD (MH-OR = 0.84; 0.73-0.97; P = 0.01), suggesting that this genotype confers a protective effect against IBD. There was no association of FcgR3a*A559C genotypes with IBD, CD, or UC in any of the three studied populations. CONCLUSIONS: The FcgR2a*519G functional variant was associated with IBD and reduced susceptibility to UC and to CD in Caucasians. There was no association between FcgR3a*5A559C and IBD, CD or UC.

[Heparin induced autoimmune thrombocytopenia: assessment of a rapid functional assay]. / La thrombopénie immunoallergique induite par l'héparine: evaluation d'un test fonctionnel rapide.

Heparin-induced thrombocytopenia (HIT) diagnosis is often difficult. Depending on the patients, the thrombocytopenia could be due to various causes. Despite their poor sensitivity and specificity, biological tests are necessary to clarify the diagnosis. In 1985, a new heparin-induced platelet aggregation assay was described that consists in determining the percentage of aggregated platelets by using an automated cell counter. Plasma samples from 18 patients with a definite HIT were tested with this quick easy-to-perform HIT diagnosis test. Positive results were obtained with 11 plasma (61%) when the test was performed with platelets from 3 different healthy volunteers (control platelets). As for the other functional tests, results are depending on control platelets and sensitivity seems to be increased when control platelets FcgammaRIIa-131 polymorphism was of His/His (but difference is not significant). In emergency situation, it is difficult to perform a functional test with control platelets from several healthy donors, and it is even more difficult to select volunteers on their FcgammaRIIa-131 polymorphism. In conclusion, in spite of its practicability, the test is not reliable to help in the rapid diagnosis of HIT. Indeed, 7 patients of 18 (39%) with definite HIT have been found negative with this test.

Crucial role of FcgammaRIIa (CD32) in assessment of functional anti-Streptococcus pneumoniae antibody activity in human sera.

Immunoglobulin G (IgG)-mediated phagocytosis by polymorphonuclear leukocytes (PMNL) constitutes the main defense against Streptococcus pneumoniae. Two leukocyte IgG receptors, FcgammaRIIa and FcgammaRIIIb, are constitutively expressed on PMNL. Blocking experiments showed FcgammaRIIa is crucial for opsonophagocytosis of serum-opsonized S. pneumoniae. The biallelic, genetically determined FcgammaRIIa polymorphism (FcgammaRIIa-R131 vs. IIa-H131) determines the capacity of IgG2-mediated phagocytosis via this receptor. Comparative studies with PMNL from donors either homozygous for FcgammaRIIa-R131 or IIa-H131 showed the latter had higher phagocytic capacity. These results were confirmed in FcgammaRIIa-R131- and FcgammaRIIa-H131-transfected IIA1.6 cells. The performance of FcgammaRIIa-transfected cells in S. pneumoniae phagocytosis was validated using sera from adults and children. Serum-induced phagocytic activity depended mainly on anti-pneumococcal IgG2 antibodies. Results obtained with PMNL and IIA1.6 cells showed high correlation (r=0.94; P<.001), and support that FcgammaRIIa transfectants are a good alternative to PMNL as effector cells in opsonophagocytosis assays.

Systemic anaphylaxis in the mouse can be mediated largely through IgG1 and Fc gammaRIII. Assessment of the cardiopulmonary changes, mast cell degranulation, and death associated with active or IgE- or IgG1-dependent passive anaphylaxis.

We attempted to elicit active anaphylaxis to ovalbumin, or passive IgE- or IgG1-dependent anaphylaxis, in mice lacking either the Fc epsilonRI alpha chain or the FcR gamma chain common to Fc epsilonRI and Fc gammaRI/III, or in mice lacking mast cells (KitW/ KitW-v mice), and compared the responses to those in the corresponding wild-type mice. We found that the FcR gamma chain is required for the death, as well as for most of the pathophysiological changes, associated with active anaphylaxis or IgE- or IgG1-dependent passive anaphylaxis. Moreover, some of the physiological changes associated with either active, or IgG1-dependent passive, anaphylactic responses were significantly greater in Fc epsilonRI alpha chain -/- mice than in the corresponding normal mice. Finally, while both KitW/KitW-v and congenic +/+ mice exhibited fatal active anaphylaxis, mast cell-deficient mice exhibited weaker physiological responses than the corresponding wild-type mice in both active and IgG1-dependent passive systemic anaphylaxis. Our findings strongly suggest that while IgE antibodies and Fc epsilonRI may influence the intensity and/or kinetics of some of the pathophysiological changes associated with active anaphylaxis in the mouse, the mortality associated with this response can be mediated largely by IgG1 antibodies and Fc gammaRIII.