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| Author(s): |
Giovanne Delechiave
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | São Paulo. |
| Institution: | Universidade de São Paulo (USP). Conjunto das Químicas (IQ e FCF) (CQ/DBDCQ) |
| Defense date: | 2024-10-18 |
| Examining board members: |
Luiz Henrique Catalani;
Marisa Masumi Beppu;
Iolanda Midea Cuccovia
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| Advisor: | Luiz Henrique Catalani |
| Abstract | |
The efficacy of drug delivery systems is a major concern in medicinal therapies across diverse fields because drug potency is related to interactions with biological systems. The complexity of in vivo conditions creates barriers that impede the bioactive from reaching the site of action. A drug delivery system must efficiently: 1) load the bioactive; 2) provide stability and protection against degradation; 3) enable mobility and transport through the bloodstream and tissues; 4) facilitate cell uptake; 5) promote endosomal escape; 6) promote sustained delivery; 7) and minimize off-target effects and toxicity. The first part of the thesis explores the adsorption and desorption processes of model proteins Lysozyme (LYS) and Bovine Serum Albumin (BSA) encapsulated on Layer-by-Layer reservoirs assembled with Polyethyleneimine (PEI), heparin, and chitosan as polyelectrolytes. The multilayers with 10 layers-pair are supported on spin-coated flat surfaces of biodegradable polyesters, such as poly(-caprolactone) (PCL) and poly(L-lactic acid) (PLLA). The characterization of these protein reservoirs included analysis of ellipsometry, atomic force microscopy, fluorescence microscopy, and quartz-crystal microbalance. The results demonstrate control over the kinetics of protein sustained delivery by precisely positioning the protein within the assembly (bottom position vs. top position vs. evenly distributed) and suggest that size and total charge of the protein are key factors for controlling its overall contribution to the assembly. The second part of the thesis presents versatile multilayered polymeric nanoparticles as a vaccine approach against SARS-CoV-2. These nanoparticles, fabricated using a nanoemulsion/solvent evaporation method, are functionalized with heparin through Layer-by-Layer assembly and coupled with a recombinant form of the receptor binding domain (RBD), an antigen of the SARS-CoV-2 Spike proteins. The nanoparticles achieved a complexation efficiency of 95% for the RBD, resulting in a final hydrodynamic diameter of 140 nm with a zeta potential of -46 mV. No toxicity was observed with the nanoparticles when tested on cultured mammalian cells or when inoculated into mice. The immunization of mice with antigen-loaded nanoparticles induced strong adjuvant effects on the serum levels of RBD-specific IgG antibodies, which could neutralize the virus under in vitro conditions. Furthermore, vaccination with these nanoparticles enhances protective immunity against SARS-CoV-2 challenge in mice. This thesis investigates the Layer-by-Layer assembly as a technique for the controlled loading and release of proteins from polymeric surfaces. It also explores the development of multilayered polymeric nanoparticles as adjuvants in a vaccine platform. (AU) | |
| FAPESP's process: | 18/14532-8 - Development of transfection device from nanoparticles functionalized by LbL for specific delivery of CRISPR/Cas9 system |
| Grantee: | Giovanne Delechiave |
| Support Opportunities: | Scholarships in Brazil - Doctorate (Direct) |