Preparation of (TA)MUC1 CTB-Glycoconjugates for Detecting Antibody-Like Proteins and as Potential Vaccine Candidates

Session: 
S2.3 Synthetic glycoproteins
Code: 
FL2.3.4
Location (hall): 
Galactose
Start/end time: 
Monday, July 1, 2019 - 15:30 to 15:35
Jonathan
Dolan

Jonathan Dolan1,2, Bruce Turnbull1, Michael Webb1, Stephan Goetz2, Robert Field2

1University of Leeds, Leeds, United Kingdom, 2Iceni Diagnosics, Norwich, United Kingdom

Healthy human MUC1 is a highly glycosylated extracellular peptide, containing a highly conserved tandem repeat sequence of 20 amino acids (GVTSAPDTRPAPGSTAPPAH)). Tumour associated MUC1 ((TA)MUC1) shows enhanced expression in 90% of breast cancers [1]. (TA)MUC1 is characterised by the incomplete glycosylation of the peptide backbone, resulting in a change of conformation and exposure of the peptide backbone to the immune system [1]. Autoantibodies from the sera of patients bound specifically to the incompletely glycosylated Ser¹⁴ and Thr¹⁵ of the GSTA domain of the tandem repeat sequence [1]. Seven glycopeptides were produced by solid phase peptide synthesis based off two regions of the MUC1 peptide sequence, one region containing Thr⁸ of the DTR region and the other containing Ser¹⁴ and Thr¹⁵ of the GSTA region. Each of the seven glycopeptides exhibits a different level of Tn antigen (α-GalNAc) expression.

Cholera toxin B-subunit has previously been investigated for the display of peptide antigens, but not for carbohydrate antigens [2]. CTB-glycoconjugates of (TA)MUC1 were prepared using C-terminal sortase-mediated transpeptidase ligation methodology. These glycoconjugates were produced on a preparative scale and will go on to be used for the identification of antibody-like proteins via phag

C-terminal labelling of CTB-LPXTG using sortase-mediated transpeptidase ligation methodology. CTB-glycoconjugates will be used in phage display experiments. 

References: 
  1. Stergiou, N.; Gaidzik, N.; Heimes, A.-S.; Dietzen, S.; Besenius, P.; Jäkel, J.; Brenner, W.; Schmidt, M.; Kunz, H.; Schmitt, E. Reduced Breast Tumor Growth after Immunization with a Tumor-Restricted MUC1 Glycopeptide Conjugated to Tetanus Toxoid. Cancer Immunol. Res. 2019, 7 (1), 113–122. https://doi.org/10.1158/2326-6066.CIR-18-0256.
  2. Langridge, W.; Dénes, B.; Fodor, I. Cholera Toxin B Subunit Modulation of Mucosal Vaccines for Infectious and Autoimmune Diseases. Curr. Opin. Investig. drugs 2010, 11 (8), 919–928.
  3. Tiede, C.; Bedford, R.; Heseltine, S. J.; Smith, G.; Wijetunga, I.; Ross, R.; AlQallaf, D.; Roberts, A. P.; Balls, A.; Curd, A.; et al. Affimer Proteins Are Versatile and Renewable Affinity Reagents. Elife 2017, 6. https://doi.org/10.7554/eLife.24903.

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