Altered expression of glycan genes in cancers induced by epigenetic silencing and tumor hypoxia: clues in the ongoing search for new tumor markers.

The glycan molecules that preferentially appear in cancers are clinically utilized as serum tumor markers. The exact reason, however, why glycans are useful as tumor markers remain elusive. Here, we will summarize lessons learned from well-established cancer-associated glycans, and propose strategies to develop new cancer markers. Our recent results on cancer-associated glycans, sialyl Lewis A and sialyl Lewis X, indicated that the repressed transcription of some glycan genes by epigenetic silencing during early carcinogenesis, and the transcriptional induction of some other glycan genes by tumor hypoxia accompanying cancer progression at locally advanced stages, are two major factors determining cancer-associated glycan expression. Multiple genes are involved in glycan synthesis, and epigenetic silencing of a part of such genes leads to accumulation of glycans having truncated incomplete structures, which are readily detected by specific antibodies. Glycans are very unique and advantageous as marker molecules because they are capable of reflecting epigenetic silencing in their structures. Transcriptional induction of some glycan genes by tumor hypoxia at the later stages produces further glycan modifications, such as an unusual increase of the N-glycolyl sialic acid residues in the glycan molecules. The entire process of malignant transformation thus creates abnormal glycans, whose structures reveal the effects of both epigenetic silencing and tumor hypoxia. The second advantage of a glycan marker over a proteinous marker is that they can reflect the plurality of genetic anomalies in a singular molecule, as it is synthesized by the cooperative action of multiple genes. Glycans are sometimes covalently bound to well-known cancer-associated proteins, such as CD44v, and this eventually contributes to a high cancer specificity and functional relevancy in cancer progression.

Clinical application of functional glycoproteomics - dissection of glycotopes carried by soluble CD44 variants in sera of patients with cancers.

We provide here an example of clinical application of functional glycoproteomics for cancer diagnosis. Sialyl Lewis a and sialyl Lewis x glycotopes, which are the specific ligands for selectins, and variant forms of CD44, which are the adhesion molecules recognizing hyaluronate, are both implicated in cancer metastasis. The CD44 variants modified by the sialyl Lewis a and sialyl Lewis x glycotopes are expected to have dual functions, serving as ligands for vascular selectins, and simultaneously having binding activity to vascular bed hyaluronate, and are expected to figure heavily in cancer metastasis. We developed a heterogeneous sandwich assay system to detect soluble CD44v specifically modified by the cancer-associated sialyl Lewis a/x glycotopes, using the extracellular domain of CD44v cleaved by the metalloproteinase ADAM10 as standard molecules. We also developed the assay system for CD44v modified by normal epithelial glycotopes including disialyl Lewis a and sialyl 6-sulfo Lewis x. The results indicated that serum levels of soluble CD44v modified by cancer-associated glycotopes were frequently increased in patients with cancers, while those of CD44v modified by the nonmalignant glycotopes tended to be elevated in patients with benign disorders.