Characteristics of the examined proteins.

a<p>M±S.D: mean value±standard deviation of each protein is indicated in g/dl except for IgE, which is indicated as IU/ml.</p>b<p>Age and body mass index (BMI) are indicated as mean values±standard deviation.</p>*<p>Log-transformed values were applied in the analysis.</p><p>Abbreviations: GWAS: genome-wide association study, TP: total protein, ALB: albumin, NAP: non-albumin protein.</p

Manhattan plots for the GWAS of (A) TP, (B) NAP, and (C) ALB.

<p>SNPs were plotted based on their physical chromosomal positions (horizontal axis) together with their –log₁₀ (<i>P-</i>values) in the GWAS (vertical axis). The black horizontal line shows the genome-wide significance threshold of <i>P</i> = 5.0×10⁻⁸. The SNPs for which <i>P</i>-values were smaller than 1.0×10⁻¹⁵ are indicated at the upper limit of the plots.</p

Regional plots for the associations of the SNPs in the GWAS stage of TP, ALB and NAP.

<p>SNPs plotted with their –log₁₀ (<i>P</i>-values) in the GWAS based on their physical chromosomal positions. Genotyped SNPs are indicated as circles, while imputed SNPs are indicated as triangles. The color scheme indicated the linkage disequilibrium displayed as <i>r</i>² values between all SNPs and the top-ranked SNP in each plot. The tested trait, chromosomal locus, and the top-ranked SNPs (in purple color) in the GWAS and combined analyses together with their <i>P</i>-values are shown in each plot. The blue lines represent the recombination rates estimated based on HapMap Phase ΙΙ database. The plots were drawn using Locus Zoom software.</p

Summary results of the GWAS and the replication study of TP, ALB, and NAP.

a<p>A1/A2: major/minor alleles.</p>b<p>The effect of the minor allele on the normalized values based on an additive genetic model.</p>c<p>For the GWAS and replication analysis, <i>P</i>-values were obtained by linear regression test model, for the Meta analysis by inverse-variance method.</p>*<p>SNPs obtained by whole-genome imputation analysis.</p><p>Abbreviations: GWAS: genome-wide association study, MAF: minor allele frequency, TP: total protein, ALB: albumin, NAP: non-albumin protein, s.e: standard error.</p

Association of the SNPs in the GWAS of the NAP with immunoglobulin isotypes.

a<p>The effect of the minor alleles on the standardized values.</p>b<p><i>P</i>-values for the associations of SNPs with each normalized immunoglobulin isotype obtained by using a linear regression model.</p><p>Abbreviations: s.e: standard error, %EV: percentage of the explanatory variance.</p

Quantitative Structural Characterization of Local N‑Glycan Microheterogeneity in Therapeutic Antibodies by Energy-Resolved Oxonium Ion Monitoring

Site-specific characterization of glycoform heterogeneity currently requires glycan structure assignment and glycopeptide quantification in two independent experiments. We present here a new method combining multiple reaction monitoring mass spectrometry with energy-resolved structural analysis, which we termed “energy-resolved oxonium ion monitoring”. We demonstrated that monitoring the yields of oligosaccharide-derived fragment ions (oxonium ions) over a wide range of collision induced dissociation (CID) energy applied to a glycopeptide precursor exhibits a glycan structure-unique fragmentation pattern. In the analysis of purified immunoglobulin glycopeptides, the energy-resolved oxonium ion profile was shown to clearly distinguish between isomeric glycopeptides. Moreover, limit of detection (LOD) of glycopeptide detection was 30 attomole injection, and quantitative dynamic range spanned 4 orders magnitude. Therefore, both quantification of glycopeptides and assignment of their glycan structures were achieved by a simple analysis procedure. We assessed the utility of this method for characterizing site-specific N-glycan microheterogeneity on therapeutic antibodies, including validation of lot-to-lot glycoform variability. A significant change in the degree of terminal galactosylation was observed in different production lots of trastuzumab and bevacizumab. Cetuximab Fab glycosylation, previously known to cause anaphylaxis, was also analyzed, and several causative antigens including Lewis X motifs were quantitatively detected. The data suggests that energy-resolved oxonium ion monitoring could fulfill the regulatory requirement on the routine quality control analysis of forthcoming biosimilar therapeutics

Association of rs364663 with age at menarche.

*<p>Effect size and S.E. of allele1 on age at menarche (year per allele) and P-values were obtained by inverse-variance method.</p

Association results in Japanese woman of previously identified SNPs with age at menarche in Caucasian woman.

<p>Genotyping result of 15,495 Japanese subjects were anlayzed in this study. Imputed SNPs with R2 of less than 0.7 were excluded from this analysis. A1 frequency of JPT were those from release 24 Hapmap JPT. N.D.; no data. References: 1 Elks et al Nat Genet 2010, 2 He et al Nature Genet 2009, 3 Liu et al Plos Genet 2009. P-value of 0.0015 (0.05/33) was set at the significant threshold for this candidate analysis.</p>*<p>:Effect size and S.E. of allele1 on age at menarche (year per allele) and P-values were obtained by inverse-variance method.</p>**<p>Concordance of association direction between this study and the previous report.</p

Results from meta-analysis of four genome-wide association studies.

<p>A total of 15,495 female samples were analyzed in this study.</p

Characteristics of study population.

<p>Characteristics of study population.</p