Biodegradable Polylactide-co-Glycolide-Chitosan Janus Nanoparticles (JNP) for the Local Delivery of the IL-6R Receptor Antagonist, Tocilizumab, for Oral Squamous Cell Carcinoma (OSCC) Chemoprevention

Abstract

<h3>Introduction</h3> Risk factor behavior e.g. smoking with/without alcohol or diseases associated with DNA repair deficits e.g. Fanconi anemia (FA) render the entire oral cavity at risk to develop OSCC. While systemically- delivered chemopreventives should conceivably provide oral mucosal field-coverage, bioavailability issues and drug-related toxicities generated disappointing outcomes. In contrast, local delivery formulations can deliver therapeutically-relevant levels without systemic side-effects. This proof of concept study determined whether Janus nanoparticles are a viable field-coverage chemopreventive strategy. Due to IL6's vital role in OSCC de- velopment, tocilizumab was selected as the payload chemopreventive.Methods:1) Formulation of tocilizumab- containing PLGA-chitosan nanoparticles (electrohydrodynamic co-jetting), 2) Determination of keratinocyte [FA- OSCC and E6/E7 transduced (epi) human cell lines] Internalization via confocal microscopy and fluorescent- activated cell sorting (FACS), 3)Assessment of nanoparticle-delivered tocilizumab immunoreactivity (tocilizumab ELISA), 4)Evaluation of fluorescenttagged nanoparticle penetration of oral mucosal explants (fluorescent mi- croscopy). Results:1) JNP were successfully formulated to achieve 65% tocilizumab loading and subsequent re- lease of immuno-reactive tocilizumab (ELISA confirmed)., 2) Qualitative (confocal microscopy) and quantitative (FACS) analyses confirmed human keratinocyte nanoparticle internalization in a time-dependent fashion. Notably, even the shortest co-incubation FACS time point (1 h) revealed appreciable internalization (75% and 73% of epi and FA-OSCC cell populations, respectively)., 3) Fluorescent-tagged nanoparticle studies revealed 85% of the mucosal explants (11/13) demonstrated particle penetration past the stratum corneum while 38% (5/13) contained nanopar- ticles in the basilar third of the epithelium. <h3>Conclusions</h3> One of the mechanisms by which nanoparticles penetrate stratified epithelium is energy dependent (transcytosis). It is therefore probable that JNP optimization combined with in vivo analyses in ATP-replete tissues will enhance nanoparticle transport to the targeted, proliferative ep- ithelial cells. Previously, our lab has shown that local injections of tocilizumab significantly reduced OSCC tumori- genesis in vivo. Studies are currently ongoing to determine the OSCC chemoprevention efficacy of locally injected, sustained-release tocilizumab JNP.