Proteoform Analysis to Fulfill Unmet Clinical Needs and Reach Global Standardization of Protein Measurands in Clinical Chemistry Proteomics

Abstract

In clinical testing of protein markers, structure variants of the measurand are often not taken into account. This heterogeneous character of protein measurands in immunoassays often renders test standardization impossible. Consequently, test results from different methods can lead to underdiagnosis or overdiagnosis and, thus, undertreatment or overtreatment of patients. The systematic structural analysis of protein isoforms has been coined proteoform profiling and is performed through mass spectrometry-based proteomics strategies. Knowledge on proteoforms allows refining existing uni-marker tests and moreover has great potential to contribute to the urgent need for new tests to predict prognosis and severity of diseases.

Apolipoprotein profiling as a personalized approach to the diagnosis and treatment of dyslipidaemia

“…177 Because different proteoforms are likely to have different functionality, quantitation of proteoforms might be of importance, specifically in light of precision medicine and targeted therapeutics. 178 Proteoforms have been found for several apos. Specifically, differential O-glycosylation of apoCIII was initially observed using isoelectric focusing, 179 and, more recently, using intact protein MS analysis.…”

“…184 Currently the clinical relevance of the proteoforms of apos for CVD are unknown, and more research is warranted. MS is an excellent technique for the in-depth characterization of an individual’s proteoform-profile, 178,185 as mass-based differences either at the intact protein or at the peptide concentration can easily be identified. Specifically the introduction of high resolution MS-instruments, which can provide accurate mass information at resolutions >60,000, would be beneficial for the analysis of proteoforms.…”

Automated Plasma Glycomics with Linkage-Specific Sialic Acid Esterification and Ultrahigh Resolution MS

“… 4 , 5 This neglect of glycans, and proteoforms in general, can be rationalized from data complexity and technical challenges. 6 − 8 In MS-based proteomics, database searches methyl-, phosphate-, and acetyl-groups are accommodated by an exactly defined mass difference, whereas glycans have different sizes (monomeric to oligomeric) that moreover can be isobaric. With regard to technical considerations, the applied analytical methods require high-throughput (HT) platforms to study the clinical relevance of protein glycosylation in large-scale studies, as larger sample sizes have a higher risk of errors and inconsistencies in sample preparation, leading to poor repeatability.…”

Protein biomarker discovery is still relevant and has entered a new phase

“…In the second approach, data is combined from different -omics technologies, with proteogenomics as a growing example [4]. A third way to obtain a multimarker panel involves mapping proteoforms of one specific protein that is measured in a clinical immunoassay-based test [5]. For example, detailed MS analysis of prostate-specific antigen revealed that this widely used prostate cancer marker actually consist of many isoforms with varying N -glycan structures.…”