Selective glycan recognition using molecularly imprinted liposomes

Informations projet

GLYCANLIPO

N° de convention de subvention: 795415

DOI

10.3030/795415

Projet clôturé

Date de signature de la CE 28 Mars 2018

Date de début 3 Decembre 2018

Date de fin 15 Mars 2021

Financé au titre de

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Coût total

€ 195 454,80

Contribution de l’UE

€ 195 454,80

Coordonné par

THE UNIVERSITY OF BIRMINGHAM
United Kingdom

Periodic Reporting for period 1 - GLYCANLIPO (Selective glycan recognition using molecularly imprinted liposomes)

Période du rapport: 2018-12-03 au 2020-12-02

Résumé du contexte et des objectifs généraux du projet

Glycans, also referred to as carbohydrates, carry unique information in biological systems that make them an important source of biomarkers for wide range of diseases. While recognition of glycans with high affinity and exquisite specificity is at the heart of the early and accurate detection of such diseases, selective glycan recognition remains a daunting task due to their inherent diversity and complexity. In this framework, a key challenge is the design of a universal platform for highly specific recognition of a broad spectrum of glycan structures. Boronic acids (BAs), which react with diols, are the most commonly used recognition moieties for the synthesis of binders for glycans, which contain many hydroxyl groups. However, the challenge is to spatially arrange carbohydrate receptors, such as BAs, with the precision and stability required to differentiate a broad range of glycans. GLYCANLIPO addressed this challenge by using concepts and tools from lipid membrane biophysics and molecular imprinting to produce BA-functionalized liposomes with high affinity binding sites. The overall approach to create imprinted liposomes involves initial free rearrangement of polymerizable lipids functionalised with BAs to form binding sites towards template glycans followed by UV polymerisation of the lipid backbone of the liposome to fix the spatial arrangement of BAs.
The Specific Research Objectives (SRO) of GLYCANLIPO were: 1) to produce, characterize and optimize polymerized liposomes functionalised with sugar receptors and, 2) to perform molecular imprinting of glycans on polymerized liposomes.
Molecularly imprinted liposomes was successfully generated using the proposed idea and a 3 fold increase in the affinity was observed for a model oligosaccharide.Given that only one type of recognition moiety (i.e. BA) was used, the results indicate that the performance of imprinted liposomes can be enhanced and that the liposome surface imprinting strategy can be easily extended to more complex systems.

Travail effectué depuis le début du projet jusqu’à la fin de la période considérée dans le rapport et principaux résultats atteints jusqu’à présent

The overall workflow of this project was:
1) Design and synthesis of structurally diverse, polymerizable lipids to formulate liposomes for selective detection of glycans. The original plan was to synthesize a wider range of amphiphilic recognition motifs, such as PCDA functionalized with tryptophan and tyrosine, which are known to be involved in biological saccharide recognition via CH-π, hydrogen bonding and hydrophobic interactions. However, given time limitations due to the COVID-19 pandemic and lockdown measures in 2020, it was decided to focus on BA functionalized PCDA. A range of functional diacetylene monomers were designed and synthesized, including PCDA-BA and PCDA-PEG-BA, PCDA-PEG), in which a 4-hydroxyphenylboronic acid molecule that served as the carbohydrate receptor and Poly(ethylene glycol) (PEG) as a spacer were covalently attached to PCDA, respectively.
2) Preparation and characterization of polymerized PCDA liposomes doped with BA-functionalized PCDA. A secondment was planned in order to assess the behaviour of polymerized liposomes using Differential Scanning Calorimetry (DSC), Isothermal Titration Calorimetry (ITC) and thermal AFM. However, it was cancelled due to the COVID-19 pandemic and associated disruptions and lockdown measures in 2020. Instead the liposomes were mainly characterized by UV spectroscopy and dynamic light scattering (DLS).
3) To establish a polydiacetylene-based assay for label free detection of glycan binding to PCDA/BA-PCDA liposomal membrane. Assays based on the colorimetric/fluorescence response of the PCDA liposomes does not directly measure the binding, rather signal the perturbation of membrane and the original packing state of the PDA backbone caused by the binding. Therefore in addition to fluorescence spectroscopy, a surface plasmon resonance (SPR) assay was established to monitor glycan interactions with BA-functionalized surface-tethered liposomes.
4) Imprinting glycans on polymerizable liposomes. The original plan was to use cancer-related glycans such sa Thomsen–Friedenreich (TF) antigen, Sialyl-Tn Antigen (STn) and Sialyl Lewis x trisaccharide as template. However, given budgetary limitations, it was decided to focus on a tetrasaccharide stachyose as the template. In addition, the interaction of a glycoprotein (HRP) with imprinted liposomes was studied.

Progrès au-delà de l’état des connaissances et impact potentiel prévu (y compris l’impact socio-économique et les conséquences sociétales plus larges du projet jusqu’à présent)

The Researcher who was involved in GLYCANLIPO achieved scientific, intellectual and practical competence. The freedom and flexibility offered by the Fellowship also opened up many opportunities for the Researcher to enter into new research communities, create new collaborations, with leading experts on glycan-based cancer diagnostics and cancer research and therapy. The Researcher prepared a proposal and was awarded another fellowship to pursue his research in this area, based on the skills he gained during this fellowship.

Overview of GLYCANLIPO