COVARIS, INC. — Department of Health and Human Services SBIR Phase I: 400

Description

The ex vivo production of cells engineered by material delivery through the cell wall is widely used in fundamental research and therapeutic developmentDNA and RNA enable the study of the functionexpressionand regulation of geneshigh throughput and patient specific screening of gene and conventional drug therapiestherapeutic tissue engineeringand the biomanufacture of proteins and other materialsTransfection methods such as viral transductionuse of cationic and liposomal materials or polymer nanoparticlesand physical methods such as electroporation nucleofection can have high efficiencies but poor cell viabilityInefficient cell transformation is emerging as a bottleneck for autologous and allogenic therapiesAs a resultthere is a significant need for an efficientscalableand more universal delivery methodCovaris will develop a platform for intracellular delivery based on precision sonoporation based on its Adaptive Focused AcousticsAFATMtechnologyIn sonoporationacoustic streaming generated by ultrasound induced oscillations of gas microbubbles disturbs the cell membranegenerating transient pores for material transferSonoporation has been explored with moderate successwith and without added microbubbles as cavitation nucleiMost previous experiments were performed with poor control of parameters such as pressure amplitudes and uniformity and the proximity and size of microbubblesExperiments which have attempted to control these parameters resulted in superior transfection rates but are not scalableThe proposed system will tightly control the acoustic fieldthe nature of shear generating microbubblesand the bubblesproximity to cellsA highly uniform acoustic field will be combined with a plastic microfluidic chipProximity to microbubbles will be controlled by either chemical means or through design of the microfluidic chipFeedback will control acoustic intensity within the chipBoth approaches lend themselves to high throughput transfection due to the short bubble excitation times required per transfectionPhase I will focus on exploring this concept in low throughput through different design approaches for three cell types inefficiently transfected by standard methodsThe project will have three componentsDevelopment of cell culture and analysis systems usinggold standardchemicallipofectionand physicalelectroporationmethodsthe development of an instrument and a series of embossed thermoplastic and cast elastomer microfluidic chips for precise sonoporationand the evaluation of cross membrane transport using fluorescent moleculeslow MW dextrans and high MW plasmidand transfection with a GFP coding plasmidThe goal of the Phase I project is to increasetransfection efficiencyXviabilitybywith afold increase in throughput relative to pate based methods This project seeks to develop a system comprised of an ultrasonic instrument and microfluidic chip for highthroughputhighly efficient delivery of payloads into cellsApplications include fundamental research into gene function and regulationgenetic engineering of cells for the production of therapeutic and other productsand for the engineering of immune and other cells for therapeutic infusion into patients