LC–MS/MS glycomic analyses of free and conjugated forms of the sialic acids, Neu5Ac, Neu5Gc and KDN in human throat cancers

Abstract

An elevated level of the free deaminated sialic acid, 2-keto-3-deoxy-D-glycero-D-galacto-nononic acid (KDN), was first discovered in human ovarian cancers (OCs), suggesting that KDN may be an oncodevelopmental antigen (Inoue S, Lin SL, Chang T, Wu SH, Yao CW, Chu TY, Troy FA II, Inoue Y. 1998. J Biol Chem. 273(42):27199-27204). To determine if this unexpected finding was unique to OC, we developed an LC-MS/MS glycomic approach to quantitatively determine the level of free and conjugated forms of KDN, Neu5Ac and Neu5Gc in head and neck cancers of the throat, and in a subpopulation of matched lymph nodes. These findings were correlated with tumor (T), nodal (N), metastatic (M) involvement and the differentiation status of the tumors. The following new findings are reported: (i) The level of free KDN in 49 throat cancers and a subpopulation of 10 regional lymph nodes accounted for 94.5 and 93.3%, respectively, of the total level of KDN (∼2 µg/g); (ii) in marked contrast, the level of free Neu5Ac in throat cancer and lymph nodes accounted for only 6.5 and 5.1% of the total level of Neu5Ac (85 µg/g); (3) The level of Neu5Gc (0.03 µg/g) in throat cancers was 0.30% of the level of Neu5Ac, two-thirds were conjugated and one-third was free. The central importance of these new findings is that the elevated level of free KDN relative to free Neu5Ac and Neu5GC in throat cancers showing no lymphatic metastasis, and which are poorly to moderately differentiated, suggests that free KDN may be useful as a biomarker for detecting some early-stage cancers at biopsy, and be of possible prognostic value in determining the potential degree of malignancy.

Human risk of diseases associated with red meat intake: Analysis of current theories and proposed role for metabolic incorporation of a non-human sialic acid

“…The beef-derived virus theory of Zur Hausen merits further research, as does the theory of Neu5Gc-induced xenosialitis. Based on the potential importance of Neu5Gc incorporation for the biology of human carcinomas, analytical methods for detection and quantification of Neu5Gc have been already been used (Samraj et al, 2014a; Wang et al, 2015). Although it is unquestionable that colon cancer is the type of cancer that has the strongest association with red meat consumption, quantitative analysis of the percentage of Neu5Gc in colorectal tumors still needs to be determined, to ascertain if this value correlates with cancer progression.…”

Recent advances in mass spectrometric analysis of glycoproteins

“…The levels of free and conjugated forms of deaminated sialic acid (KDN), Neu5Ac and Neu5Gc in head and neck cancers of the throat, and in a subpopulation of matched regional lymph nodes, using C 18 ‐LC–MS/MS technique were investigated by Wang et al. . The results revealed that higher levels of free KDN, relative to levels of free Neu5Ac and Neu5Gc, were expressed in throat cancers and in a subset of paired regional lymph nodes that would be useful as a biomarker for detecting some early‐stage cancers and the degree of malignancy.…”

Metabolic Glycoengineering with N‐Acyl Side Chain Modified Mannosamines

“…In mammalian species the level of KDN expression often exceeds that of Neu5Gc. [118] With respect to the amino substituents,both Neu5Ac and Neu5Gc can be expressed in mammalian tissues.H owever, because of am utation in the gene coding the synthesis of CMP-Neu5Ac-hydroxylase,h uman cells are not able to synthesize Neu5Gc or catalyze its covalent modifications. [119,120] Thep resence of Neu5Gc,o riginally thought to represent an oncofetal antigen, is now known to be present in human tissues as ar esult of ad iet rich in dairy products and red meat.…”

“…These findings suggest that the level of free KDN may be ap otential biomarker for detecting early stage tumors at biopsy,a nd be of potential prognostic value in determining the degree of malignancy. [118] Thes tructural role of Neu5Ac expression on cell-surface glycoconjugates has been widely reviewed, [2,3] particularly in regard to the aberrant glycosylation pattern of tumor cells. [136] An additional characteristic that is underreported is the turnover rates of membrane-bound N-glycans.W ith respect to individual membrane glycoproteins,ithas been shown that Neu5Ac and other peripheral sugar residues,for example,Gal and GlcNAc,s how af aster rate of turnover than the core sugars mannose and the chitobiose moiety (Scheme 5).…”

“…[344] PolySia not only fulfills critical functions during development in normal cells and tissues,b ut is also an important cancer-associated antigen involved in tumor metastasis. [66,[345][346][347][348][349][350] Since polySia is rarely found in healthy adult humans,other than in selected regions of the brain that show persistent expression, [118] the development of antibodies against this epitope is an attractive and relevant area for cancer research. In contrast to the physiological polySia glycan, N-propionylated and N-butanoylated polySia glycans are highly antigenic.…”