Exploring serum and immunoglobulin G N-glycome as diagnostic biomarkers for early detection of breast cancer in Ethiopian women

Abstract

Background Alterations in protein glycosylation patterns have potentially been targeted for biomarker discovery in a wide range of diseases including cancer. Although there have been improvements in patient diagnosis and survival for breast cancer (BC), there is no clinically validated serum biomarker for its early diagnosis. Here, we profiled whole serum and purified Immunoglobulin G (IgG) fraction N -glycome towards identification of non-invasive glycan markers of BC. Methods We employed a comprehensive glycomics approach by integrating glycoblotting-based glycan purification with MALDI-TOF/MS based quantitative analysis. Sera of BC patients belonging to stages I-IV and normal controls (NC) were collected from Ethiopian women during 2015–2016. IgG was purified by affinity chromatography using protein G spin plate and further subjected to glycoblotting for glycan release. Mass spectral data were further processed and evaluated rigorously, using various bioinformatics and statistical tools. Results Out of 35 N -glycans that were significantly up-regulated in the sera of all BC patients compared to the NC, 17 complex type N -glycans showed profound expression abundance and diagnostic potential (AUC = 0.8–1) for the early stage (I and II) BC patients. Most of these glycans were core-fucosylated, multiply branched and sialylated structures, whose abundance has been strongly associated with greater invasive and metastatic potential of cancer. N -glycans quantified form IgG confirmed their abundance in BC patients, of which two core-fucosylated and agalactosylated glycans ( m/z 1591, 1794) could specifically distinguish (AUC = 0.944 and 0.921, p ≤ 0.001) stage II patients from NC. Abundance of such structural features in IgG is associated with a decrease in its immunosuppressive potential towards tumor cells, which in part may correlate with the aggressive nature of BC commonly noticed in black population. Conclusions Our comprehensive study has addressed for the first time both whole serum and IgG N -glycosylation signatures of native black women suffering from BC and revealed novel glyco-biomarkers with marked overexpression and distinguishing ability at early stage patients. Further studies on direct identification of the intact glycoproteins using a glycoprteomics approach will provide a deeper understanding of specific biomarkers towards their clinical utility. Graphical abstract Electronic supplementary material The online version of this article (10.1186/s12885-019-5817-8) contains supplementary material, which is available to authorized users.

Glycan-specific antibodies as potential cancer biomarkers: a focus on microarray applications

“…With the use of suspension arrays (beads), glycan levels may be analyzed. Hydrazide-coated silica particles are used for the isolation of glycans with further analysis with MALDI-TOF MS [93,94]. At the same time, analysis of anti-glycan antibodies can be held using micro-and suspension arrays.…”

Antibody Panel Based N‐Glycan Imaging for N‐Glycoprotein Biomarker Discovery

“…The majority of serum N ‐glycan methods focus on the analysis of a pool of N ‐glycans released from all proteins in the serum (Gornik et al., ; Kirmiz et al., ; Novokmet et al., ; Reiding et al., ; Ruhaak et al., ; Ruhaak, Xu, Li, Goonatilleke, & Lebrilla, ), or the analysis of one target protein's N ‐glycan profile (Comunale et al., ; Pompach et al., ; Ruhaak et al., , ; Shubhakar et al., ; Simunovic et al., ; Šimurina et al., ; Theodoratou et al., ; Zhang et al., ). Pooled serum analyses have shown trends in overall N ‐glycan changes in the presence of cancer, such as increased fucosylation, branching, and bisects (Gebrehiwot et al., ; Hecht et al., ; Snyder et al., ; Vučković et al., ).…”

“…Current Protocols in Protein Science 2014; Reiding et al, 2019;Ruhaak et al, 2008;Ruhaak, Xu, Li, Goonatilleke, & Lebrilla, 2018), or the analysis of one target protein's N-glycan profile (Comunale et al, 2010;Pompach et al, 2016;Ruhaak et al, 2013Ruhaak et al, , 2018Shubhakar et al, 2016;Simunovic et al, 2019;Šimurina et al, 2018;Theodoratou et al, 2016;Zhang et al, 2016). Pooled serum analyses have shown trends in overall N-glycan changes in the presence of cancer, such as increased fucosylation, branching, and bisects (Gebrehiwot et al, 2019;Hecht et al, 2015;Snyder et al, 2016;Vučković et al, 2016).…”

Breast cancer diagnosis by analysis of serum N-glycans using MALDI-TOF mass spectroscopy

“…In this study, we analyzed serum N-glycomic patterns based on N stage, which is directly related to biomolecular signatures within bodily fluid. Because we acquired N-glycans from patient serum samples, the N stage was primarily considered rather than the overall cancer stage [9].…”