Lectins 101: How Plant Lectins Can Accelerate Cancer Biomarker Discovery

Lectins 101: How Plant Lectins Can Accelerate Cancer Biomarker Discovery
Posted in: Tips and Tricks
Castor Bean Plant Seeds Growing in Guatemala
Castor Bean Plant Seeds Growing in Guatemala

The glycome of a cell is, quite frankly, a festival entry ticket for that cell; it determines how the cell functions, where it migrates, and what that implicates for its destination inside the festival area. We have constantly emphasized the importance of glycosylation in cell-to-cell interactions. For cells within the tumor microenvironment, this can lead to proliferation and metastatic behavior. So, how do these aberrantly glycosylated cells communicate with each other and the tumor microenvironment? Enter lectins, the festival buddies waiting for their besties at the entrance gate and accompanying them throughout the festival.

There is more to the story: the marriage between a glycan and a lectin is unique, and, because of this, lectins can be leveraged to help us achieve breakthroughs in biomedical research. Most importantly, they help us distinguish healthy from diseased cells.

How can science utilize lectins, or why is it worth the effort in the first place? Read on to find out.

Overview of Lectins and Their Functions

Lectins are part of a broader group called glycan-binding proteins that recognize glycan chains and mediate their functionality. These ubiquitous proteins serve a vast range of purposes through their smart recognition mechanisms in animals, plants, bacteria, and viruses.

There is an enormous diversity of lectins, depending on their structures and cellular locations, but they each have similar functions necessary for the survival of all species.

Take E. coli, for example. The lectins on their cell surfaces recognize glycan chains in our gastrointestinal tract, which is how they can adhere to our gut wall and survive our immune response [1].

The task list of lectins in eukaryotes is much longer. Some mediate the folding and transfer of glycoproteins throughout the cell for further glycosylation, or degradation (if the glycosylation was aberrant). They are also critical to the body’s immune response. Glycan binding by certain lectins, such as ficolin, mediates immune cell recruitment to the inflammation site and signals the cells to secrete anti-inflammatory cytokines [3].

How Do Lectins Bind Glycans?

The key to the attraction between a glycan and a lectin lies in its carbohydrate recognition domain (CRD). If you looked at a computer simulation of a protein surface, these domains would look like small cavities or pockets on the outside.

There is more to the interaction between two molecules than meets the eye, however. For starters, the lectins undergo protein folding, particularly beta-sheet formation, so their binding pockets become more exposed to the glycan. Inside these pockets, the lectin contains active site residues that make various non-bonded interactions with the glycan. This involves a combination of hydrogen bonds, van der Waals interactions, and ionic bonds. As a cherry on top, the presence of ions like calcium strengthens the binding even more [4].

It is also interesting to note that many lectins have more than one CRD, allowing them to bind multiple glycans simultaneously. This multifunctionality enhances their binding affinity to the cell surface.

Plant Lectins: From the 19th Century to Present Day

Galectin-3 (carbohydrate binding domain) protein, bound to N-acetyllactosamine
Galectin-3 (carbohydrate binding domain) protein, bound to N-acetyllactosamine

Now that we have covered the basics of lectins in their natural habitat, let’s now focus on how we can use lectins to ask research questions. In fact, lectin research dates back to the late 19th century. Because of their abundance in nature, plant lectins have been the protagonists of these discoveries.

The first highlight of lectin research was the ability of some plant lectins (extracted from castor bean seeds, to be specific) to agglutinate and precipitate red blood cells to the extent that some showed toxicity in mice models [5]. A few decades later, it was found that lectins agglutinated red blood cells by binding to the glycan chains on red blood cell surfaces. This marked their first use as cell biomarkers (to determine blood types) [6].

Lectins entered the cancer research marathon in the 1960s when researchers discovered that two plant lectins, phytohemagglutinin (PHA) [7] and wheat germ agglutinin (WGA) [8], specifically recognized and agglutinated tumor cells.

It was later established that not all lectins had agglutination features, but they all possessed at least one domain that specifically recognized carbohydrate structures and bound them reversibly [9]. The revelation of reversible binding was particularly game-changing because it meant that the lectin binding did not disrupt glycan structures. As the field of lectinology thrived, researchers discovered more and more properties that increased the feasibility of plant lectins, including stability at high temperatures and low pH [10] and resistance to digestive enzymes [11].

Applications of Plant Lectins

Coronavirus illustration
Coronavirus illustration.

Today, plant lectins are widely used in a range of applications. The toxicity of plant lectins towards insects makes them potential candidates for agricultural insecticide production [12]. Cell and animal studies have been generating results about the antimicrobial and antiviral activities of plant lectins, and you can find several studies revealing their potential for the treatment and prevention of HIV [13] and even SARS-CoV-2 [14].

From 1960 to the present day, their anti-cancer activity has been demonstrated in numerous studies involving leukemia, sarcoma, hepatoma, and breast cancer [15]. They have also been used in drug delivery to enhance the targeting abilities of their nanocarriers [16].

Even when they are not actively used to test anti-tumor activity, plant lectins still contribute a great deal. Their ability to bind surface glycans shined more light on the altered glycan

structures expressed in tumor cells [17]. Lectins do not merely differentiate between healthy cells and tumor cells. Their high-level specificity could even help us to distinguish between tumors with differing levels of aggression (e.g., in the detection of cells with metastatic behavior).

Plant Lectins in Glycan Analysis

The importance of glycosylation in cellular processes is undebatable. Research has also established how pathogenic changes in glycosylation wreak havoc in our bodies. Thus, the importance of precise biomarkers for these changes is more important than ever.

With plant lectins, the detection of glycan biomarkers has become a reality. Through many decades of lectinology studies, researchers have developed several techniques for repurposing plant lectins in glycan analysis.

Lectin affinity chromatography [18] and lectin arrays are ideal for identifying and isolating glycoproteins. They involve mounting and immobilizing lectins on a support platform. Then, through mass spectroscopy, you can determine the structures of carbohydrates that the immobilized lectins bind.

Mouse Tongue: endothelial cells stained with Dylight 594-labeled Griffonia simplififolia lectin, Isolectin B4, (red). Mounted with VECTASHIELD HardSet with DAPI (Blue)
Colon Carcinoma (FFPE) was antigen retrieved with Antigen Unmasking Solution stained with DyLight® 594 labeled UEA I. Mounted in VECTASHIELD® HardSet with DAPI.

Lectins can also be incorporated into enzyme-linked assays. The working principle of such assays is similar to ELISA, except you replace antibodies with lectins, hence the name enzyme-linked lectin assay (ELLA) [20]. This makes lectins suitable for high-throughput multi-well plates, through which you can quantitatively analyze specific glycan–lectin interactions with spectrophotometers. It is cost-effective, and you often need only small amounts, making ELLA quite popular in glycobiology.

Vectashield Plus antifade mounting medium, H-1900 from Vector Laboratories
Human Colon: Rabbit Anti-Cytokeratin (AE1/AE3) and Mouse Anti-Desmin detected simultaneously with VectaFluor duet Double Labeling Kit; Vasculature detected using DyLight 649 UEA I lectin purple. Mounted in VECTASHIELD PLUS.

Lectin histochemistry and cytochemistry can provide an even stronger perspective, allowing deeper insights into glycome analysis. Thanks to the development of lectin labels, you can monitor the pathogenic glycosylation in cells and tissues via fluorescent imaging and access a better overview of how cells with aberrant glycosylation proliferate, interact, and migrate inside your sample.

The combination of quantitative and qualitative plant lectin assays enables a better grasp of cancer biology, posing huge potential for precise characterization of tumor type and progression. Lectins can open doors to a more comprehensive understanding of cancer biology, with detailed glycome profile reports that can drive actionable insights.

Conclusion

From the early research of castor bean seeds and blood cell agglutination to the identification of biomarkers of cancer, lectins are providing accessible insights into the glycome. The broad spectrum of lectin assays allows us to approach cancer glycobiology from different yet complementary angles.

Determining the right lectin assays for your specific research goals can be overwhelming, but Vector Laboratories has your back! In fact, we will delve into the technical details of lectin assays in our next blog posts. In the meantime, you can download our lectin guide for a comprehensive understanding of various plant lectin assays and their working principles.

References

  1. Lindhorst, Thisbe K. 2015. Small molecule ligands for bacterial lectins: letters of an antiadhesive glycopolymer code. Glycopolymer Code: Synthesis of Glycopolymers and their Applications.
  2. Słomińska-Wojewódzka, Monika, and Kirsten Sandvig. 2015. The role of lectin-carbohydrate interactions in the regulation of ER-associated protein degradation. Molecules
  3. Mason, Christopher P., and Alexander W. Tarr. 1992. Human lectins and their roles in viral infections. Molecules.
  4. Imberty A, and James H. Prestegard. 2017. Chapter 30 – Structural Biology of Glycan Recognition. Essentials of Glycobiology [Internet]. 3rd edition.
  5. Stillmark, H. 1888. Über Ricin ein giftiges Ferment aus den Samen von Ricinus communis L. und einige anderen Euphorbiaceen. MD Thesis, University of Dorpat, Dorpat, Estonia
  6. Renkonen, K. O. 1948. Studies on hemagglutinins present in seeds of some representatives of the family of Leguminoseae. Annales medicinae experimentalis et biologiae fenniae. Vol. 26. No. 1. KALEVANKATU 11 A, 00100 HELSINKI, FINLAND: FINNISH MEDICAL SOC DUODECIM.
  7. Nowell, Peter C. 1960. Phytohemagglutinin: an initiator of mitosis in cultures of normal human leukocytes. Cancer research.
  8. Aub, Joseph C., Carol Tieslau, and Ann Lankester. 1963. Reactions of normal and tumor cell surfaces to enzymes, I. Wheat-germ lipase and associated mucopolysaccharides. Proc Natl Acad Sci USA.
  9. Peumans, Willy J., and E. J. Van Damme. 1995. Lectins as plant defense proteins. Plant Physiology.
  10. Pérez-Giménez, Julieta, et al. 2009. Soybean lectin enhances biofilm formation by Bradyrhizobium japonicum in the absence of plants. Int J Microbiol.
  11. Zhu-Salzman, Keyan, et al. 1998. Carbohydrate binding and resistance to proteolysis control insecticidal activity of Griffonia simplicifolia lectin II. Proc Natl Acad Sci USA.
  12. Reyes-Montaño, Edgar Antonio, and Nohora Angélica Vega-Castro. 2018. Plant lectins with insecticidal and insectistatic activities. Insecticides - Agriculture and Toxicology.
  13. Akkouh, Ouafae, et al. 2015. Lectins with anti-HIV activity: a review. Molecules.
  14. Sohrab, Sayed S., et al. 2020. The emergence of human pathogenic coronaviruses: Lectins as antivirals for SARS-CoV-2. Current Pharmaceutical Design.
  15. Yau, Tammy, et al. 2015. Lectins with potential for anti-cancer therapy. Molecules.
  16. Neutsch, L, et al. 2013. Synergistic targeting/prodrug strategies for intravesical drug delivery—Lectin-modified PLGA microparticles enhance cytotoxicity of stearoyl gemcitabine by contact-dependent transfer. J Control Release.
  17. Cummings RD, and M.E. Etzler. 2009. Chapter 45 – Antibodies and Lectins in Glycan Analysis. Essentials of Glycobiology. 2nd edition.
  18. Merkle, Roberta K., and Richard D. Cummings. 1987. Lectin affinity chromatography of glycopeptides. Methods Enzymol.
  19. Hu, Shen, and David T. Wong. 2009. Lectin microarray. Proteomics Clin Appl.
  20. McCoy Jr, J. Philip, James Varani, and Irwin J. Goldstein. 1983. Enzyme-linked lectin assay (ELLA): use of alkaline phosphatase-conjugated Griffonia simplicifolia B4 isolectin for the detection of α-D-galactopyranosyl end groups. Anal Biochem .
  21. Hashim, Onn Haji, Jaime Jacqueline Jayapalan, and Cheng-Siang Lee. 2017. Lectins: an effective tool for screening of potential cancer biomarkers. PeerJ.
November 10, 2021
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