Cell membrane disruption and recovery for intracellular delivery

PROJECT ABSTRACT Intracellular delivery plays an essential role in biological research and therapeutic applications, however, efficient intracellular delivery of exogenous compounds and macromolecular cargo remains a long-standing challenge. The complex mechanisms of established methods and their often unpredictable impact on cell behaviour have dramatically limited the scope of biological experiments and reduced efficacy of potentially promising cell therapy concepts. Membrane disruption-based approaches have emerged as key strategies for rapid, direct and universal intracellular delivery because they are less dependent on cargo properties and cell types, being able to deliver almost any submicron material dispersed in solution. The ability to rapidly switch membrane-perturbing effects on and off provides an additional level of control, enabling temporal manipulation and rapid, almost instantaneous delivery. However, key challenges of membrane disruption strategies have been: 1) inconsistent level of plasma membrane injury (which would lead to low viability and efficiency); 2) poor throughput or scalability (e.g. microinjection); and 3) inadequate understanding of plasma membrane disruption and recovery response. The PI's research group at University of Colorado Boulder centers on interdisciplinary research at the frontiers of Biology, Medicine, Physics, and Micro/Nano Engineering. The main research thrust in the PI's group is to develop new technologies to quantitatively understand cell membrane disruption and recovery, and explore its application for next generation precise intracellular drug delivery. The goals for the next five years are to i) develop a novel microfluidic platform, including NanoEngineered Surface Technology and Acoustofluidic devices, that can precisely generate uniform and homogenous disruptions at cell membrane with controllable number and size of the pores, to quantitatively understand cell membrane disruption and recovery dynamics at molecular, cellular, proteiomic and high throughput level; ii) demonstrate a precise intracellular drug delivery system with controllable dose, minimum toxicity, maximum efficiency, and high throughput, providing insight for or promoting the next generation intracellular drug delivery. So far, biologists have not applied the fundamental insights gleaned from membrane disruption and repair studies toward engineering cell permeability. The proposed research grogram will bridge the scientific gap between these two disparate fields: the engineering of intracellular delivery approaches; and the cellular mechanobiology of plasma membrane disruption and repair response. The biomedical research community would benefit greatly from a more mechanistic and transparent understanding of intracellular delivery, both to further the development of more robust techniques and to realize key medical and industrial applications. 1