Disease prevention

The prevalence of protein-carbohydrate interactions in pathogen mechanisms makes them an important target for disease prevention in the form of carbohydrate vaccine targets, new bacterial species-specific antibiotic targets and anti-adhesion therapy. These will now be addressed.

1.2.8.1 Carbohydrate vaccine targets

Despite many pathogens mimicking the host glycome, many still have pathogen specific oligosaccharide presentations, which could provide targets suitable for carbohydrate-based vaccines. For example, the antigenic response to arabinogalatan from M. tuberculosis is a potential vaccine candidate as the component galactofuranose is not present in mammalian cells; this minimising the chance of generating an autoimmune response.20 Monoclonal antibodies have been generated that are directed at the cluster presented mannose-rich oligosaccharides present on human immunodeficiency virus (HIV) glycoprotein 120 and new broadly neutralizing carbohydrate based antibodies have also been extracted from patients.89, 90 A carbohydrate specific immune response to the Streptococcus pneumonia bacteria has also been generated in mice in response to gold nanoparticles coated with a T-helper

peptide and the tetrasaccharide repeating unit of the S. pneumoniae capsular polysaccharide.91

Protein conjugated carbohydrate vaccines have also been developed for H. influenza

type b, where meningitis caused by this bacteria has been eradicated in areas with a vaccine program.92 A species specific tetrasaccharide has also been identified for

Bacillus anthracis93 and administration of this allowed the generation of antibodies specific to this bacteria although more work needs to be done to develop this into a usable vaccine against anthrax.94 Proteins conjugated to the P. falciparum specific glycophosphatidylinositol (GPI) hexasaccharide have also been developed as a potential conjugate vaccine. This vaccine conferred reduced mortality to malaria in mice without any cross-reactivity with human GPI although more work to develop this is still on going.95

Carbohydrate vaccines are also being pursued as an anti-cancer treatment by utilising antibodies produced by the aberrant glycosylation of mucins on tumour cells. This involved the coupling of the sialylated Tn antigen (where the Tn Antigen is just N- Acetyl-D-Galactosamine coupled to a serine or a threonine residue) that is produced on aberrant mucins. This vaccine was able to generate a humoral and a cellular immune response to the cancer cells.96

1.2.8.2 Pathogen specific glycosyltransferase targets

As glycosylation is a process involving the expression of many glycosyltransferases and the existence of non-mammalian monosaccharides there will be pathogen specific glycosyltransferases. Drugs that target these enzymes may make the pathogen more

25

susceptible to the host’s immune system by removing its ability to mimic host glycan presentation on its surface or by reducing its pathogenicity through other mechanisms.

Neuraminidases play a crucial role in influenza infection. Influenza particles adhere to the cell surface through adhesion to sialic acid residues. Upon creation of a new virus particle, the viral neuraminidase clips the terminal sialic acid residue from the cellular glycan thus allowing release of the viral particle from the cell surface where it can then go on and infect other cells.97 This process is the target of neuraminidase inhibitors such as oseltamivir and zanamivir although resistance to such drugs is becoming more widespread.

V. cholerae can also produce a bacterial neuraminidase and this capability is found amongst all strains that cause the most severe cholera infection.98 The role of this neuraminidase is to clip sialic acid residues from glycans on the cell surface in order to truncate the glycans to form GM1 (the natural binding ligand of the cholera toxin).99, 100 This increases the cellular presentation of the cholera toxin binding ligand thereby increasing the amount of toxin binding and causing increased disease severity.101 Inhibition of this neuraminidase could reduce disease severity in severe cholera infections.

As previously mentioned, the presence of sialic acid terminated glycans play a crucial role in the protection of H. influenza and Neisseria spp. from the complement immune system. As such, inhibition of the enzymes responsible for addition of the sialic acid cap to the glycans on the surface of these bacteria will make them more susceptible to killing by the immune system and offers a potential drug target (vide infra).

1.2.8.3 Anti-adhesion therapy

For those pathogens that have an essential carbohydrate mediated adhesion phase (Figure 1.15A), anti-adhesion therapy has potential for prevention of infection and treatment of disease. Many examples of carbohydrate based anti-adhesion compounds exist, but some examples include the use of glycopolymers for the inhibition of cholera toxin binding102, 103 and for the specific inhibition of fimbriated (and thus pathogenic) strains of E. coli.104105 _{This mechanism is also used in nature to protect}

cells from bacterial infection, mucins which are found on the surface of epithelial cells are also secreted thus trapping pathogens before they can invade the underlying tissue (Figure 1.15B).21, 106

Figure 1.15 Schematic depicting anti-adhesion therapy. (A) Normally a bacteria will

bind to the underlying cell glycans resulting in infection (B) whereas a drug or a mucin that mimcs these glycans can prevent binding and thus prevent infection.

A B Bacterial adhesion Infection Prevention of adhesion Infection

27

A form of anti-adhesion therapy has also been proposed for the prevention of cancer metastases. Many cancer cells contain regions of aberrant glycosylation, one common modification is the presence of many selectin ligands. This would mask the cancer cells from the immune system as selectins are used to suppress the immune system and prevent it from attacking host cells. Several compounds have been designed to mimic the glycan-binding domain of selectins in order to prevent binding of selectins to tumour cells.107-109

1.2.8.4 Carbohydrate based drugs

Carbohydrates also form the basis of a number of other drug compounds including a number of antibiotics such as kanamycin, vancomycin and teicoplannin.110 _Heparin

and related compounds such as dermatan sulphate form the basis of many anti- thrombotic agents.111, 112 Glycoproteins have been used as enzyme replacement therapy in patients with Hurler and Hurler-Scheie forms of mucopolysaccharidosis I.113