Leveraging membrane curvature to create phase-responsive lipid systems

Rueben, Jacob Kenneth

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https://hdl.handle.net/2142/122085 Copy

Description

Title

Leveraging membrane curvature to create phase-responsive lipid systems

Author(s)

Rueben, Jacob Kenneth

Issue Date

2023-05-26

Director of Research (if dissertation) or Advisor (if thesis)

Leal, Cecília

Doctoral Committee Chair(s)

Leal, Cecília

Committee Member(s)

• Murphy, Catherine J
• Statt, Antonia
• Espinosa-Marzal, Rosa
• Wang, Hua

Department of Study

Materials Science & Engineerng

Discipline

Materials Science & Engr

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

• Lipids
• Self-assembly
• Membranes
• Nonlamellar
• Saxs
• Gold Nanorods
• Red-shifted Dyes
• Biomolecules
• Nanotechnology
• Therapeutics
• Cosmetics

Language

eng

Abstract

A core component of life is the lipid, an unassuming amphiphilic class of molecules that spontaneously assemble in living organisms to create complex structures with myriad essential functions. Understanding the driving forces of lipid self-assembly, as well as harnessing their structural and functional diversity, is a goal that supports many intersecting research fields, including but not limited to those of therapeutics and cosmetics. It is therefore important from a fundamental and application standpoint to learn how these self-assembled systems function, in order to create novel beneficial formulations. Bilayer systems comprising lipid mixtures are the most well-studied model of biological membranes. While the plasma membrane of cells is a single bilayer, many intra- and extra-cellular biomembranes comprise stacks of bilayers. Most bilayer stacks in nature are periodic maintaining a precise water layer separation between bilayers. That equilibrium water separation is governed by multiple inter-bilayer forces and is highly responsive. Biomembranes re-configure inter-bilayer spacing in response to temperature, composition, or mass transport cues. In synthetic bilayer systems for applications in cosmetics or topical treatments, control of the hydration level is a critical design handle. Herein we investigated a binary lipid system that leverages key inter-bilayer forces leading to unprecedented levels of aqueous swelling. We found that combining cationic lipids with inverse cubic phase-forming lipids (lipids with positive Gaussian modulus), results in the stabilization of multilamellar phases against repulsive steric forces that typically lead to bilayer delamination at high degrees of swelling. Using Ultra-small-angle X-ray scattering alongside confocal laser scanning microscopy, we characterized various super-swelled states of 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and glycerol monooleate (GMO) lipids, as well as other analogous systems, at varied concentration and molar ratios. Through these experiments we established swelling profiles of various binary lipid systems that were near-linear with decreasing lipid volume fraction, showing maximum swelling distances above 200 nanometers. While traditional lamellar systems have been used in therapeutic applications, robust drug and gene delivery systems require innovative methods to control payload release and tune delivery efficiency. The most promising delivery materials are lipid-based and their efficiency often hinges on structural transformations activated by endogenous pH changes. Exogenously driving phase transitions in lipid assemblies is a tantalizing idea that could lead to better control of cargo release dynamics. Multiple reports have demonstrated phase transitions induced in lipid systems, achieved via plasmonic heating of entrained gold nanorods. However, undesirable non-localized heating is common because lipid assemblies often exhibit lattice dimensions of just a few nanometers that render gold particles challenging to integrate due to their incommensurate sizes, especially in lipid nanoparticle or colloidal forms. We investigate these processes using a judiciously chosen ternary lipid system with entrained small gold nanorods that undergoes transitions between bicontinuous cubic and inverse hexagonal phases on exposure to near-infrared light. Utilizing small-angle X-ray scattering alongside electron reconstruction, we show that gold nanorods integrate into the lipid assembly core lattice by co-localizing in the water nano-channels. We also found that plasmonically activated transformations occur in a couple of minutes and are reversible. In order to build on our ternary GOPEG-AuNR system and tune it for easier pre-clinical adoption, we set out to streamline formulations by finding a more scalable and readily available heating agent. We created a remotely-controlled activation system allowing for heightened control and targeting of genetic payload delivery using our well-documented bulk quaternary lipid-dye conjugate system. Herein we define a nonlamellar liquid crystalline lipid system with incorporated FDA-approved amphiphilic red-shifted dye indocyanine green (ICG) that allows for remotely triggered nonlamellar phase transitions to the fusogenic inverse hexagonal phase via near-infrared (NIR) irradiation. We characterize this system with synchrotron small-angle X-ray scattering (SAXS) during both external and laser-induced heating experiments, and establish ideal compositions for future colloidal drug delivery applications. Robust drug and gene delivery systems require innovative methods to control payload release and tune delivery efficiency. The most promising delivery materials are lipid-based and their efficiency often hinges on structural transformations activated by endogenous pH changes. Exogenously driving phase transitions in lipid assemblies is a tantalizing idea that could lead to better control of cargo release dynamics. Multiple reports have demonstrated phase transitions induced in lipid systems, achieved via plasmonic heating of entrained gold nanorods. However, undesirable non-localized heating is common because lipid assemblies often exhibit lattice dimensions of just a few nanometers that render gold particles challenging to integrate due to their incommensurate sizes, especially in lipid nanoparticle or colloidal forms. We investigate these processes using a judiciously chosen ternary lipid system with entrained small gold nanorods that undergoes transitions between bicontinuous cubic and inverse hexagonal phases on exposure to near-infrared light. Utilizing small-angle X-ray scattering alongside electron reconstruction, we show that gold nanorods integrate into the lipid assembly core lattice by co-localizing in the water nanochannels. We also found that plasmonically activated transformations occur in a couple of minutes and are reversible. Bilayer systems comprising lipid mixtures are the most well-studied model of biological membranes. While the plasma membrane of cells is a single bilayer, many intra- and extra-cellular biomembranes comprise stacks of bilayers. Most bilayer stacks in nature are periodic maintaining a precise water layer separation between bilayers. That equilibrium water separation is governed by multiple inter-bilayer forces and is highly responsive. Biomembranes re-configure inter-bilayer spacing in response to temperature, composition, or mass transport cues. In synthetic bilayer systems for applications in cosmetics or topical treatments, control of the hydration level is a critical design handle. Herein we investigate a binary lipid system that leverages key inter-bilayer forces leading to unprecedented levels of aqueous swelling. We found that combining cationic lipids with inverse cubic phase-forming lipids (lipids with positive Gaussian modulus), results in the stabilization of multilamellar phases against repulsive steric forces that typically lead to bilayer delamination at high degrees of swelling. Using Ultra-small-angle X-ray scattering alongside confocal laser scanning microscopy, we characterized various super-swelled states of 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and glycerol monooleate (GMO) lipids, as well as other analogous systems, at varied concentration and molar ratios. Through these experiments we established swelling profiles of various binary lipid systems that were near-linear with decreasing lipid volume fraction, showing maximum swelling distances above 200 nanometers. With the recent advent of lipid nanoparticle (LNP) mRNA vaccines, the formulation of next-generation lipidic nanovectors is already underway. An ever-significant barrier to the cellular delivery of RNA payloads is endosomal escape, a process we have previously shown nonlamellar phases such as bicontinuous cubic to be superior at when compared to traditional lamellar lipid formulations. However, increased delivery rates are still desirable, and while many modern formulations rely on endogenous environmental triggers such as late-endosomal pH to induce conformational changes, these compositions rely on systemic processes and are uncontrollable post-injection. We propose a remotely-controlled activation system allowing for heightened control and targeting of genetic payload delivery using our well-documented bulk quaternary lipid-dye conjugate system. Herein we define a nonlamellar liquid crystalline lipid system with incorporated amphiphilic red-shifted dye indocyanine green (ICG) that allows for remotely triggered nonlamellar phase transitions to the fusogenic inverse hexagonal phase via near-infrared (NIR) irradiation. We characterize this system with synchrotron small-angle X-ray scattering (SAXS) during both external and laser-induced heating experiments, and establish ideal compositions for future colloidal drug delivery applications.

Graduation Semester

2023-12

Type of Resource

Text

Handle URL

https://hdl.handle.net/2142/122085

Copyright and License Information

Copyright 2023 Jacob Rueben

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