title: Analysis and optimization of interactions between peptides mimicking the GD2 ganglioside and the monoclonal antibody 14G2a.
abstract: Overexpression of the GD2 ganglioside (GD2) is a hallmark of neuroblastoma. The antigen is used in neuroblastoma diagnosis and to target newly developed therapies to cancer cells. Peptide mimetics are novel approaches in the design of antigens for vaccine development. We previously reported the isolation of five GD2-mimicking peptides from the LX-8 phage display library with the monoclonal antibody (mAb) 14G2a. The goal of our current study was to analyze and optimize the binding of the peptide mimetics to the mAb 14G2a. Therefore, we performed further experiments and supported them with molecular modeling to investigate structure-activity relationships that are the basis for the observed mimicry of GD2 by our peptides. Here, we show that the peptides have overlapping binding sites on the mAb, 14G2a and restricted specificity, as they did not crossreact with other ganglioside-specific antibodies tested. In addition we demonstrate that the phage environment was involved in the process of selection of our peptides. The AAEGD sequence taken from the viral major coat protein, p8, and added to the C-termini of the peptides #65, #85 and #94 significantly improved their binding to the mAb, 14G2a. By application of analogs with amino acid substitutions and sequence truncations, we elucidated the structure-activity relationships necessary for the interactions between the 14G2a mAb and the peptide #94 (RCNPNMEPPRCF). We identified amino acids indispensable for the observed GD2-mimicry by #94 and confirmed a pivotal role of the disulphide bridge between the cysteine residues of #94 for binding to the mAb 14G2a. More importantly, we report five new peptides demonstrating a significant improvement of mAb 14G2a binding. The experimental data were supported and expanded with molecular modeling tools. Taken together, the experimental results and the in silico data allowed us to probe in detail the mechanism of the molecular mimicry of GD2 by the peptides. Additionally, we significantly optimized binding of the leading peptide sequence #94 to the mAb 14G2a. We can conclude that our findings add to the knowledge on factors governing selections of peptide mimetics from phage-display libraries.
authors: Laboratory of Molecular Genetics and Virology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, 7 Gronostajowa St, 30-387 Krakow, Poland. irena.horwacik@uj.edu.pl
authors: Irena
authors: I
authors: Horwacik
authors: Mateusz
authors: M
authors: Kurciński
authors: Małgorzata
authors: M
authors: Bzowska
authors: Aleksandra K
authors: AK
authors: Kowalczyk
authors: Dominik
authors: D
authors: Czaplicki
authors: Andrzej
authors: A
authors: Koliński
authors: Hanna
authors: H
authors: Rokita
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D001665:Binding Sites; D045744:Cell Line, Tumor; D005732:Gangliosides / Q000276:immunology*; D006801:Humans; D008958:Models, Molecular; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D009447:Neuroblastoma / Q000235:genetics / Q000276:immunology*; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D013329:Structure-Activity Relationship
chemical_list: D000911:Antibodies, Monoclonal; D005732:Gangliosides; D019151:Peptide Library; D010455:Peptides; C019403:ganglioside, GD2