Developing artificial cells for delivering drug molecules to micro-engineered heart tissue
Creators: name: Hay, Callum
Publisher: name: Imperial College London
Year: 2022

description: Batch-based liposome production methods are widely applied within academia. Although batch-based methods offer poor mechanical control, limited size accessibility, and low batch size, therefore, their usage in the study of drug delivery liposomes can present challenges. Additionally, ever increasing specificity of drug delivery liposomes means more representative models of the human micro-environment are required to facilitate efficacy testing. This is especially pertinent in the field of cardiology, where current drug success rates are low. Herein, this work attempts to further understand these two challenges and first present a means for higher throughput, continuous liposome production that has scope for size exploration. Through the application of liposomes produced by this method, the efficacy of drug delivery in both conventional cardiac tissue culture and bioprinted cardiac tissue models can be studied. Initially addressed are the limitations of batch-based liposome synthesis by employing the previously reported method of microfluidic hydrodynamic flow focusing, a flow-based production method with well-defined mechanical formation. This is extended to another reported method of vertical flow focusing to achieve greater throughput, through the application of a larger channel aspect ratio. Increased throughput ensures the application to larger cell-based validation studies, without sacrificing sample quality. The vertical flow focusing device used in this work employed a differing approach of polymethylmethacrylate multilayer assembly method. This fabrication was designed to be achieved in non-specialist institutions, thus presenting a fully accessible method. Additionally, the multilayer assembly method is amenable to incorporating further functionality. This device was shown to produce liposomes in a therapeutically relevant size window (&lt; 200 nm diameter). In further exploration of the device both the impact of liposome formulation and residual ethanol were explored. Lipid formulation trends shown within the same device instruct how to achieve desired liposome size and distribution, empowering size control within the device application. The exploration into ethanol using molecular rotors is the first attempt at evaluating the impact of residual solvent on the lipid bilayer, this suggested the use of a high flow rate ratio (&gt; 25) for producing minimally perturbed lipid bilayers. Next, the development of tissue-on-chip models was explored, ranging from polydimethylsiloxane base devices to a bioprinted tissue model. The design and application of a bioprinted tissue model showed potential scalability and ease of analysis. Bioprinted models also enable the study of extracellular matrix, this is especially important for liposomal drug delivery systems as it presents another recognised barrier to liposomal delivery that is currently largely unaddressed. A study of liposomal drug delivery through model extracellular matrixes could further the development of liposome formulations for penetrating specific tissue compositions. Finally, the performance of targeting (I-1 peptide) and non-targeting control liposomes was explored using the encapsulation of the antagonist menadione. Within tissue culture, targeted liposomal menadione delivery was shown to inhibit cell viability (36.3 ± 12.4 %) whereas non-targeting exhibited minimal impact on viability (84.4 ± 11.7 %). This was the first exploration into menadione delivery and could potentially demonstrate a renewed application, given menadione has previously been reported as a chemotherapeutic agent. The study was replicated using a bioprinted tissue model, demonstrating the first application of liposomes to a bioprinted cardiac tissue model. Overall, this work achieved liposomal drug delivery within an academic setting using approaches that could be applied in scaled liposome validation studies.