Rachel, J Kulchar
Smruti Nair
Francis Mante
Henry Daniell
| Rachel, J Kulchar | Smruti Nair | Francis Mante | Henry Daniell |
Rachel, J Kulchar, Smruti Nair, Francis Mante, Henry Daniell
Department of Basic and Translational Sciences, University of Pennsylvania, School of Dental Medicine
Introduction
Our laboratory has shown that enzymes engineered in plant cells incorporated into chewing gums debulk viruses in the saliva of COVID-19 patients or disrupt the dental biofilm and kill pathogens that cause dental caries, to which there are no comparable options. Advantages of this approach include affordability, stability of enzymes in chewing gums for several years at ambient temperatures, controlled release, and enhanced duration of contact with pathogens than oral rinse. In this study, we optimize in vitro release kinetics by simulating human chewing using gum tablets containing different doses of target proteins.
Methods
The feasibility of topical drug delivery was tested on two drug-containing gum tablets: the green fluorescent protein (GFP) and the angiotensin converting enzyme-2 (ACE-2). The GFP gum contained 448 µg GFP, and the two doses of ACE-2 gum were comprised of 70 µg and 140 µg ACE-2. Plant cells expressing GFP were incorporated into gum, and release kinetics after mechanical stimulation were quantified via fluorescence. The ACE-2 protein was impregnated in gum, and release was quantified using Western Blot. Simulated chewing of the gum was accomplished using an Instron machine (model 5564) at varying timepoints.
Results
Release was linear until 20 minutes of mechanical chewing for all tablets, and a sustained release of protein material was observed. After 20 minutes of simulated chewing, 30% GFP release for the GFP-impregnated tablet was recorded and 33% and 42% ACE-2 release was observed for the 70 µg and 140 µg ACE-2 tablets, respectively. Therefore, drug release was not time-dependent but dose-dependent, and release continued up to 60 minutes when high dose gum tablet was evaluated.
Conclusion
We were able to optimize and effectively simulate human chewing patterns using the Instron machine. This mode of testing will be useful for future experimental design. When comparing varying doses of ACE-2 chewing gum, the lower-dosed gum released less ACE-2 at each timepoint than the higher-dosed gum. Chewing time was not paramount; instead, the release was dose-dependent. Since the gums showed sustained release of the plant protein, longer chewing ensures optimal results.