Study of the fusion mechanism of cationic fusogenic liposomes and their potential as carrier system

• Grant number: 17/09367-5
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• Start date: August 01, 2017
• End date: July 31, 2022
• Field of knowledge: Biological Sciences - Biophysics - Molecular Biophysics
• Principal Investigator: Karin Do Amaral Riske
• Grantee: Rafaela Ramos Mororó Cavalcanti
• Host Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil
• Associated research grant: 16/13368-4 - Nanostructured systems: from membrane biomimetic models to carriers of bioactives, AP.TEM
• Associated scholarship(s): 18/08014-4 - Study of the fusion mechanism of fusogenic cationic liposomes using advanced optical microscopy tools, BE.EP.DD

Abstract

Liposomes have been used as a drug delivery system in biological media. However, conventional liposomes typically enter the cell via endocytic routes, making it difficult to deliver the encapsulated material to the intracellular medium. To optimize direct intracellular delivery, a fusogenic liposomal system was recently developed. This system has presented high fusion efficiency with both biomimetic lipid vesicles and cells. These fusogenic liposomes are composed of equal fractions of DOTAP, a cationic lipid, and DOPE, a lipid with a curvature that is able to promote fusion, and a smaller fraction of a fluorescent lipid. In this project, we intend to extend studies with model systems to understand the role of lipid composition and membrane phase in the mechanism and fusion efficiency, and also to test the intracellular delivery of these systems in cancer cell culture. The DOTAP and DOPE lipids will be replaced by analogous lipids with other hydrocarbon tails (different length and degree of unsaturation) to test the role of the membrane phase in the fusion efficiency. On the other hand, the composition of the cellular mimetic system (small liposomes-LUVs and giant liposomes-GUVs) will also be varied to include cholesterol-containing systems and in biologically relevant phases, such as liquid-ordered and liquid-disordered phases. In particular, a fusogenic system which is triggered when the system reaches the physiological temperature will be tested. The thermal behavior of the lipid mixtures will be evaluated by Differential Scanning Calorimetry (DSC). The melt efficiency will be quantified by FRET efficiency measurements using confocal microscopy, Isothermal Titration Calorimetry (ITC) and zeta potential measurements and Dynamic Light Scattering (DLS). The fusion mechanism and efficiency will be discussed and compared with membrane properties that are relevant to the membrane fusion process, such as line tension, diffusion coefficient and bending modulus. Finally, the delivery capacity of materials encapsulated in the fusogenic liposomes will be evaluated both in biomimetic systems (GUVs) and in cancer cell culture. (AU)