Combater a doença de Alzheimer usando copolímeros multivalentes feitos sob medida

The main objective of the Copolym4AD project is to design and develop safe, sustainable and innovative materials for the production of multifunctional copolymers for the treatment of AD. To this end, the following Specific Objectives (SO) will be tackled: SO1. Development of safe, sustainable and innovative materials, in particular stimuli-responsive materials, following the principles of Safe and Sustainable by Design (SSbD). Advanced materials (alkyne-terminated PDMAEMA and amine-terminated methacrylate-based glycopolymers) will be synthesized using SARA-ATRP. Novel methacrylate-based glycopolymers are synthesized from glucose or lactobionic acid. After polymerization, the alkyne-terminated PDMAEMA are functionalized with acryloyl chloride to improve its mucoadhesive properties, resulting in quaternized polymers. The materials are characterized by RMN, SEC, FITR, DSC and TGA to determine the molecular weight, chemical composition, DPI and thermal stability. SO2. Optimization of the functional and biological properties of glycopolymers and their combination with PDMAEMA to generate “clickable” copolymers with two stimuli for spatio-temporally controlled release. The glycopolymers will be used as chemical drug models to study the effects of their structural properties on the inhibition or disaggregation of amyloid aggregates (structure-activity relationships) so that we can determine which of them have therapeutic potential. Possible structural changes in amyloid aggregates after incubation with glycopolymers are evaluated using different techniques. For the first time, the glycopolymers will be investigated as amyloid inhibitors in a neuronal environment. Novel “clickable” copolymers with two stimuli are generated by combining the functional polymer segments through the creation of an aminoacrylate bond (a pH/ROS-labile linker) and the physicochemical properties will be evaluated. To ensure the degradability of the aminoacrylate bond, different stimuli will be used to trigger a controlled degradation of the copolymer: a solution containing H2O2 and an acidic solution. SO3. Preparation and characterization of miR-29-loaded copolymers for intranasal delivery. The best dual-responsive copolymers will be conjugated to miR-29 by electrostatic interactions and will be monitored in vitro for key parameters: physicochemical properties, biodegradability, transfection efficiency, and live/dead kit assays. Nasal powders are obtained by spray drying or freeze drying and their storage stability will be evaluated. In turn, the potential mucoadhesive properties of the copolymers will be investigated by adhesion and permeation experiments on nasal cell lines and ex vivo porcine nasal mucosa. Histologic/immunohistochemical analysis of nasal tissue samples will evaluate tissue structure and inflammation. SO4. Provide evidence for a “Proof of Concept” of the applicability of the miR-29-loaded copolymers to the amyloid pathway. The optimal configuration of the new miR-29-loaded copolymers will be validated in in vitro studies with relevant disease models to confirm their safety, efficiency and efficacy in reducing BACE1 expression (BACE1 inhibitor) and inhibiting amyloid aggregation. Therefore, according to our design strategy, it is possible to produce a chemical drug with only two active molecules (glycopolymers and PDMAEMA) that can be self-administered by patients and represents an alternative to immunotherapies for the treatment of AD.