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

Abstract

Blood and serum N-glycans can be used as markers for cancer diagnosis, as alterations in protein glycosylation are associated with cancer pathogenesis and progression. We aimed to develop a platform for breast cancer (BrC) diagnosis based on serum N-glycan profiles using MALDI-TOF mass spectroscopy. Serum N-glycans from BrC patients and healthy volunteers were evaluated using NosQuest's software "NosIDsys." BrC-associated "NosID" N-glycan biomarkers were selected based on abundance and NosIDsys analysis, and their diagnostic potential was determined using NosIDsys and receiver operating characteristic curves. Results showed an efficient pattern recognition of invasive ductal carcinoma patients, with very high diagnostic performance [area under the curve (AUC): 0.93 and 95% confidence interval (CI): 0.917-0.947]. We achieved effective stage-specific differentiation of BrC patients from healthy controls with 82.3% specificity, 84.1% sensitivity, and 82.8% accuracy for stage 1 BrC and recognized hormone receptor-2 and lymph node invasion subtypes based on N-glycan profiles. Our novel technique supplements conventional diagnostic strategies for BrC detection and can be developed as an independent platform for BrC screening.

Mass spectrometry in clinical glycomics: The path from biomarker identification to clinical implementation

“…This is a challenge encountered also in other fields, such as metabolomics and proteomics [148] , [149] , [150] , and is partly caused by the shortage of large and well-defined prospective clinical cohorts (or their limited accessibility) and an insufficient statistical evaluation of the initial findings, resulting in poor validation of initial results. An example would be the reporting of 8 differentially expressed N -glycans in 82 breast cancer patients compared to 27 healthy controls in one study [151] , while a different study reported 25 different N -glycans to be differentially expressed in 256 breast cancer patients compared to 311 healthy controls [152] . While the analytical methods used were different, it is highly likely that these incongruent results are confounded by general descriptors of the population, such as age, sex and BMI [14] , [38] , [153] .…”

Potential of MALDI-TOF-based serum N-glycan analysis for the diagnosis and surveillance of breast cancer

“…solid phase extraction (SPE), was performed based on previously described methods 14 . Serum N-glycans were www.nature.com/scientificreports/ released by an enzymatic reaction using PNGase F, followed by a purification step using columns packed with porous graphite carbon (PGC).…”

“…Analytical methods. Sample and matrix preparation for MALDI-TOF MS analysis was conducted as described previously 14 . For preparing the matrix solution, 2,5-dihydroxybenzoic acid (DHB) was dissolved in a mixture of 40 mM sodium chloride solution and ACN (3:1, v/v) to a final concentration of 20 mg/mL.…”

“…The reference N-glycans are listed in Supplementary Table S2. Absolute peak intensity (API i ) of each N-glycan was normalized by the sum of all APIs, yielding the normalized absolute peak intensity (NAPI i ) using a previously described formula 14 .…”

“…breast cancer and healthy control samples 14 . In this study, we aimed to explore the possibility of expanding the application of MALDI-TOF-based N-glycan analysis to breast cancer diagnosis by obtaining recurrence samples and samples acquired during follow-up.…”