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theses

A major challenge for DNA vaccination platforms is the need for a secondary delivery mechanism to promote cellular transfection. In this work, we develop and investigate intradermal drug delivery approaches for the purpose of enhancing transfection of DNA-based vaccines and therapeutics. We focus our efforts so that these approaches are cost-effective, pain-free, require minimal training, and are highly efficient.
The first part of this dissertation examines the molecular distribution of materials post intradermal injection. The boundary of the dome-like bleb formed during injection is assumed to represent the lateral extent of the injected material. We systematically characterize cargo molecule distribution (puddle) as a function of injection volume and molecular/particle size post injection. In general, results indicate that the puddle forms a subdomain laterally contained within the bleb, with an area inversely correlating to the molecular size of the injected material. The lateral distribution appears to have no time-dependency up to 10 min post injection. The trend in the depth of cargo penetration is also similar, with smaller particles extending deeper into the dermis and subcutaneous fat layers. Establishing a baseline characterization for the puddle is essential to successfully targeting the injected drug and focusing secondary applications onto it.
The second part of the dissertation reports on three approaches to deliver DNA-based vaccines and therapeutics. The suction-based approach consists of applying a moderate negative pressure (65-90 kPa for 5-300 seconds) atop the injection site; a technique similar to Chinese báguàn and Middle Eastern hijama cupping therapies. Strong GFP expression was demonstrated with pEGFP-N1 plasmids where fluorescence was observed as early as 1 hour after dosing. Modeling indicates a strong correlation between focal strain/stress and expression patterns. The absence of visible and/or histological tissue injury contrasts with current in vivo transfection systems such as electroporation. Specific utility was demonstrated with a synthetic SARS-CoV-2 DNA vaccine, which generated host humoral immune response in rats with notable antibody production. The microneedle insertion approach is a minimally invasive approach. The insertion of a 1-2 mm long, 8x8 microneedle array for 5 seconds atop of the injection site can significantly enhance GFP plasmid uptake over injection alone. In fact, GFP signal quantification shows that the transfection enhancement due to the microneedle insertion is comparable to the suction approach. In addition to having a very short application time, this approach is non-battery powered, which allows for mass-vaccinations in remote areas where electricity is scarce. Finally, we present our ongoing work on the electroporation delivery approach. Firstly, we have developed a hand-held, battery powered, simple-to-use electroporation device with an intradermal microneedle-electrode array attachment for in vivo electroporation. Electrical testing of this prototype shows comparable capabilities against the conventional function generator/amplifier system. Our extensive experimental sets revealed two major limitations to the electroporation approach: (i) the electric current limitation of the instrumentation set and (ii) the colocalization between the micro-electrodes and the puddle. These two limitations will be addressed in future works.

http://dissertations.umi.com/gsnb.rutgers:12387

Ph.D.

Includes bibliographical references

doi:10.7282/t3-ncwj-jx87

The author owns the copyright to this work.