Synthesis and characterization of nanometer-sized liposomes for encapsulation and microRNA transfer to breast cancer cells

Introduction: The use of liposomes as a drug delivery carrier (DDC) for the treatment of various diseases, especially cancer, is rapidly increasing, requiring more stringent synthesis, formulation, and preservation techniques to bolster safety and efficacy. Liposomes otherwise referred to as phospholipid vesicles are self-assembled colloidal particles. When formed in either the micrometer or nanometer size range, they are ideal candidates as DDC because of their biological availability, performance, activity, and compatibility. Defining and addressing the critical quality attributes (CQAs) along the pharmaceutical production scale will enable a higher level of quality control for reproducibility. More specifically, understanding the CQAs of nanoliposomes that dictate its homogeneity and stability has the potential to widen applications in biomedical science. Methods: To this end, we designed a study that aimed to define synthesis, characterization, formulation (encapsulation), preservation, and cargo delivery and trafficking as the major components within a target product profile for nanoliposomes. A series of synthetic schemes were employed to measure physicochemical properties relevant to nanomaterial drug product development, including concentration gradients, probe versus bath sonication, and storage temperature measured by microscopy (electron and light) and dynamic light scattering. Results: Concentration was found to be a vital CQA as reducing concentrations resulted in nanometer-sized liposomes of <350 nm. Liposomes were loaded with microRNA and fluorescence spectroscopy was used to determine loading efficacy and stability over time. Lyophilization was used to create a dry powder formulation that was then assessed for stability for 6 months. Lastly, breast cancer cell lines were used to ensure efficacy of microRNA delivery and localization. Conclusion: We conclude that microRNA can be loaded into nanometer-sized liposomes, preserved for months in a dried form, and maintain encapsulation after extended time periods in storage.