Plasmonic nanobubbles for target cell-specific gene and drug delivery and multifunctional processing of heterogeneous cell systems

Abstract

Cell and gene cancer therapies require ex vivo cell processing of human grafts. Such processing requires at least three steps -cell enrichment, cell separation (destruction), and gene transfer -each of which requires the use of a separate technology. While these technologies may be satisfactory for research use, they are of limited usefulness in the clinical treatment setting because they have a low processing rate, as well as a low transfection and separation efficacy and specificity in heterogeneous human grafts. Most problematic, because current technologies are administered in multiple steps -rather than in a single, multifunctional, and simultaneous procedure -they lengthen treatment process and introduce an unnecessary level of complexity, labor, and resources into clinical treatment; all these limitations result in high losses of valuable cells. We report a universal, high-throughput, and multifunctional technology that simultaneously (1) inject free external cargo in target cells, (2) destroys unwanted cells, and (3) preserve valuable non-target cells in heterogeneous grafts. Each of these functions has single target cell specificity in heterogeneous cell system, processing rate > 45 mln cell/min, injection efficacy 90% under 96% viability of the injected cells, target cell destruction efficacy > 99%, viability of not-target cells >99% The developed technology employs novel cellular agents, called plasmonic nanobubbles (PNBs). PNBs are not particles, but transient, intracellular events, a vapor nanobubbles that expand and collapse in mere nanoseconds under optical excitation of gold nanoparticles with short picosecond laser pulses. PNBs of different, cell-specific, size (1) inject free external cargo with small PNBs, (2) Destroy other target cells mechanically with large PNBs and (3) Preserve non-target cells. The multi-functionality, precision, and high throughput of all-in-one PNB technology will tremendously impact cell and gene therapies and other clinical applications that depend on ex vivo processing of heterogeneous cell systems.

Role of cytoskeletal mechanics and cell membrane fluidity in the intracellular delivery of molecules mediated by laser‐activated carbon nanoparticles

“…Efficient delivery of molecules into cells is needed for applications in the laboratory and the clinic that seek to alter cell behavior using small molecules, proteins, nucleic acids, and other bioactive compounds (Fu et al, 2014;Yang and Hinner, 2015). In addition to methods that rely on active transport mechanisms to enter the cell (e.g., endocytosis), there are a number of biophysical approaches that transiently permeabilize the cell membrane to deliver molecules intracellularly, such as ultrasound (Castle et al, 2013;Lentacker et al, 2014;Liu et al, 2012), electroporation (Kim and Lee, 2017;Yarmush et al, 2014), and photoporation methods (Chakravarty et al, 2010;Clark et al, 2014;Davis et al, 2013;Delcea et al, 2012;Kalies et al, 2014;Kodama et al, 2000;Lukianova-Hleb et al, 2014;Sengupta et al, 2014;Terakawa et al, 2013;Waleed et al, 2013;Wu et al, 2015;Xiong et al, 2016). When using these methods, there is a delicate balance between delivery of molecules and retaining high cell viability, wherein excessive physical stresses can damage the cell irreversibly, resulting in cell death.…”