Gliadin-Blocking Peptides In vitro assessment of their potential to alleviate celiac disease development

Doctoral thesis, 2010

This thesis work aimed to investigate the potential of synthetic peptides as agents to block wheat prolamins (gliadins) that trigger the development of symptoms in celiac disease. The first step of this investigation included selection and synthesis of peptides with high affinity to gliadins and assessment of the potential of these so-called blocking peptides to limit gliadin reactivity in vitro. Wheat proteins targeted by blocking peptides were characterized, and peptides ability to reduce gliadin recognition was evaluated in in vitro assays. It was tested whether blocking peptides could reduce gliadin processing by tissue transglutaminase, and its recognition by anti-gliadin antibodies. The digestive stability of complexes formed by gliadin and blocking peptides was also studied. Finally, their potential to reduce a negative non-immunological effect of gliadin on intestinal mucosal cells was assessed in vitro in a Caco-2 cell line model. A large pool of 12-mer peptides with a high affinity to gliadins was selected with the phage display technology, and two peptides denoted P61, and P64, were chosen for experiments. Blocking peptides expressed an affinity to a broad spectrum of gliadin proteins and to α-amylase/trypsin inhibitors, wheat allergenic proteins that are involved in eliciting an immune response in baker’s asthma. Both blocking peptides significantly reduced the tissue transglutaminase processing of intact gliadin and partially reduced its recognition by anti-gliadin antibodies. The blocking peptides also partially reduced the negative non-immunological effect that gliadin had on a Caco-2 cell line. However, complexes of blocking peptides with gliadin were only partially stable after the pancreatic phase of in vitro digestion, with P64 complex with gliadin being more stable than that of P61. The two chosen blocking peptides, P61 and P64, have proven the potential to bind to gliadin and to partially prevent its toxicity and recognition in in vitro assays. In order to be used in celiac disease therapy, however, more efficient gliadin blocking peptide complexes need to be explored. The large pool of 12-mer peptides obtained during selection with the phage display will further be screened in a search for the most effective peptides.