By Nicole A. Cowan
Carbohydrates, also known as glycans, are one of the four foundational biological macromolecules, along with nucleic acids, proteins, and lipids. All of these macromolecules have distinct, essential roles in biological systems. Indeed, an improved understanding of the molecular components of biological systems has informed disease research. It has also yielded increasingly individualized biological insights, which have, in turn, informed patient care, culminating in the rise of precision medicine. However, to date precision medicine has benefited little from glycomics—the comprehensive study of glycans in biological systems. Until recently, glycomics applications were decades behind proteomics and genomics applications.
In part, this lag was due to the limitations of research tools that could address the complexity of glycan synthesis, structure, and function. Unlike proteins and nucleic acids, which are made from a DNA template, glycans are products of interconnected biosynthetic pathways affected simultaneously by the environment and heredity. In addition, glycans have diverse functions as unbound macromolecules and can also modify the function and identity of other biological molecules when covalently linked as glycoconjugates. For example, the glycans bound to a protein during glycosylation may influence how the protein—now the glycoprotein—migrates to the cell surface. The glycans may also influence how the glycoprotein interacts with surrounding molecules and cells once it arrives.
Increasing evidence of the glycome’s role in human health has stressed the necessity of integrating glycomics into precision medicine. Cell surface expression of glycoconjugates determines intra-organismal recognition and extra-organismal interactions. Microbes that contribute key functions to their host—such as food digestion, vitamin anabolism, and absorption—rely on glycoconjugates to communicate to immune cells that they are not harmful. Pathogens that cause deadly diseases, such as tuberculosis and malaria, use a similar strategy to evade immune attack. In this article, we’ll explore the technologies propelling glycomics research, and the existing applications of glycan biomarkers and therapeutics for treating disease.
New tools propel glycomics research
Glycomics has the potential to pioneer novel therapeutic approaches, and it can significantly complement genomics and proteomics tools by improving the characterization of molecules critical to disease progression. Comparative glycomics will be critical for the identification of glycans that contribute to disease, and further research will unlock insights into how glycomic abnormalities are related to clinical outcomes.
Key technologies used to advance glycomics include immunohistochemistry (IHC) and liquid chromatography-mass spectrometry (LC-MS), as well as high-throughput fluorescence flow cytometry, which has long been used to differentiate cell populations using glycoproteins.
LC-MS provides structural information about glycoconjugates by deciphering multiple glycoprotein isoforms from single protein species. High-throughput MS-based proteomics now provides unbiased surveys of cellular proteomes, and the approach is quickly being adapted to glycomics. One barrier to full realization of the technology is the diversity and complexity in glycan structures. However, structural reference databases are rapidly emerging to fill the gap.
Disease-related glycoconjugates that are identified by LC-MS can be further evaluated or verified with IHC, which utilizes antibody-based capture to visualize glycoconjugate expression with cell-staining or fluorescence. Immunocytochemistry (ICC) uncovers glycoprotein expression patterns at the cellular level using the same visualization strategies as IHC by targeting intact cells instead of tissue fragments.
Adapting IHC and ICC to focus on additional glycoconjugates, such as glycolipids, will open up new visualization capabilities in glycomics, including the broader characterization of metabolic pathways and junctions. Tandem or multitechnology workflows, such as MS-IHC, will offer laboratories added analytical power to tackle glycoconjugate complexity and optimize glycoconjugate diagnostics.
Therapeutic applications of glycomics
Glycomics research is now at the forefront of exciting opportunities in the diagnosis, monitoring, and treatment of diseases, including diseases that were clinically baffling when investigated from a genomics or proteomics perspective. Therapeutic areas where glycomics has been especially impactful include oncology, along with cardiovascular, gastrointestinal, liver, and infectious diseases.
Glycan biomarkers in diagnosis
Glycoconjugate abnormalities that are characteristic of disease can be turned into novel biomarkers, with a wide range of clinical applications. For example, Helicobacter pylori an infectious bacteria, linked to the development of gastritis and gastric cancer, expresses Lex and Ley antigens, which mimic the cell-surface glycoconjugates of gastric endothelial cells.
These glycans act as camouflage for H. pylori bacteria, allowing it to infect the epithelium of the stomach. Identifying monoclonal antibodies that bind to Lex and Ley enabled researchers to assess the presence of Lex and Ley in patients using IHC, thereby improving the diagnosis and prediction of gastric cancer due to H. pylori infection.
When abnormal glycosylation is a driver of disease, the resulting glycoconjugates can serve as disease-specific biomarkers. For example, high levels of fucosylation—the addition of the sugar fucose—has been reported in several cancer lines including hepatocellular carcinoma. The traditional tumor biomarker for this carcinoma is alpha-fetoprotein (AFP), a glycoprotein abundant in human fetuses, but with an unknown function in adults.
When AFP-L3—the fucosylated form of AFP—is used as a tumor marker instead of AFP, the detection sensitivity of primary hepatocellular carcinoma is enhanced to nearly 50%. The U.S. Food and Drug Administration has recently approved AFP-L3 as a tumor marker for primary hepatocellular carcinoma. Further characterization of the glycoconjugates linked to cancers and other diseases will help researchers develop additional biomarkers, alongside novel therapies.
Understanding how abnormal glycans are implicated in disease progression has facilitated the identification of novel drugs. One area already demonstrating strong therapeutic potential is chronic inflammatory gastrointestinal diseases.
In the gastrointestinal tract, glycoconjugates help to regulate the microbiome and microbial interactions with immune cells. Abnormalities in the intestinal glycome have been associated with a disproportionate number of harmful gut bacteria and with chronic inflammation.
Researchers at Kaleido Biosciences are developing therapeutics for immune-mediated and inflammatory diseases with synthetic glycans that are selectively metabolized in the gut microbiome to suppress the growth of bacteria associated with inflammation. One synthetic glycan candidate, KB295, is intended to treat ulcerative colitis, an inflammatory bowel disorder (IBD).
Preclinical studies of the orally administered KB295 have shown promising results, and clinical data are expected later in 2021. The administration of microbiome-mediating glycans could radically improve treatment for the 16% of IBD sufferers who do not respond to the current standard of care, and other patients with hard-to-treat autoimmune and inflammatory conditions.
A recent study in patients with congenital disorders of glycosylation (CDG) confirmed the importance of glycosylation in the regulation of low-density lipoprotein (LDL) metabolism. Elevated levels of LDL have long been associated with a higher risk of heart disease and stroke. In CDG patients, mutations in the glycoprotein ALG6, as well as PMM2, an enzyme in the synthesis of oligosaccharides, were shown to correlate with elevated levels of LDL.
CDG has debilitating neuromuscular and psychomotor symptoms, and these research results suggest ALG6 and PMM2 have potential as CDG therapeutic targets by reducing LDL accumulation. Investigating lipoprotein glycosylation pathways glycomically could identify further therapeutic targets for diverse cardiovascular diseases.
With the development of new high-throughput tools to characterize the structure and expression of glycans, along with continued innovation of glycan biomarkers and therapeutics, the clinical applications of glycomics are catching up to those of genomics and proteomics. Integrating glycomics into a comprehensive omics approach will provide insights into the interplay of genes, proteins, and glycans, which will revolutionize precision medicine. Clinical trials also stand to benefit from the integration of digital health platforms and omics insights, a development that promises to help researchers develop a more nuanced understanding of disease, and to help clinicians address the healthcare gaps of high-need patients.
Nicole A. Cowan is senior director, Project Management and IVD Operations and Strategy, Medical Device and Diagnostic Research, ICON.