Transdermal drug delivery offers advantages over oral administration and injection, including non-invasiveness, avoidance of first-pass metabolism, painless application, improved patient compliance, and avoidance of needle phobia. However, efficient transdermal delivery of hydrophilic biomacromolecules like peptides, proteins, and nucleic acids remains a significant challenge. Existing methods, such as using membrane-penetrating peptides or physical enhancement devices (ultrasound, electroporation, microneedles), have limitations, including low efficiency for larger molecules, invasiveness, and potential for skin damage. The development of safe and efficient methods for transdermal delivery of biomacromolecules, particularly for vaccines, is crucial, especially considering the recent COVID-19 pandemic and the need for convenient at-home vaccination strategies. This research aims to overcome this limitation by developing a new transdermal delivery platform using a biocompatible fluorocarbon-modified chitosan (FCS). The researchers hypothesized that FCS, inspired by its successful application in transmucosal delivery, could effectively deliver biomacromolecules through the skin.

The existing literature highlights the limitations of current transdermal drug delivery methods for biomacromolecules. Membrane-penetrating peptides have shown limited efficacy for larger proteins, while physical methods like electroporation and sonophoresis can cause skin damage. Microneedles, though promising, present manufacturing and quality control challenges. Ionic liquids and hyaluronic acids have shown some potential but limited efficacy. Lipid nanocarriers, such as ethosomes, have also been explored but further advancements are needed. The authors of this study leverage prior work on fluorocarbon-modified chitosan's efficacy in transmucosal delivery to propose it as a potential enhancer for transdermal delivery of biomacromolecules.

The study involved the synthesis of fluorocarbon-modified chitosan (FCS) through amide coupling of perfluoroalkyl carboxylic acid to chitosan. FCS nanocomplexes were formed by mixing FCS with various proteins (IgG, OVA, aPDL1, aCTLA4, S1 protein of SARS-CoV-2) at different mass ratios. The physicochemical properties of the nanocomplexes (size, zeta potential, morphology) were characterized using techniques like dynamic light scattering (DLS), transmission electron microscopy (TEM), and circular dichroism (CD). Transdermal delivery efficiency was evaluated using a Franz diffusion cell system with mouse, rabbit, and porcine skin. Confocal microscopy was used to visualize the penetration of nanocomplexes into the skin. The in vitro cytotoxicity of FCS was assessed using HACAT cells. The transdermal mechanism was investigated by studying the effects of FCS/IgG on transepithelial electrical resistance (TEER) of HACAT cell monolayers, analyzing the distribution of tight junction proteins (ZO-1), and assessing myosin light chain (MLC) phosphorylation. In vivo studies involved the treatment of B16F10 melanoma tumors in mice using transdermal delivery of aPDL1, alone or in combination with aCTLA4. Tumor growth, survival rates, and immune cell infiltration (flow cytometry) were analyzed. A proof-of-concept study used transdermal delivery of a SARS-CoV-2 vaccine (FCS/S1/polyIC) in mice, assessing humoral and cellular immune responses. The study also evaluated the transdermal delivery efficiency of FCS/IgG in rabbit and porcine skin models.

The FCS-based nanocomplexes exhibited effective transdermal penetration across mouse, rabbit, and porcine skin, primarily through intercellular and transappendageal routes. The optimal mass ratio of FCS to protein for efficient delivery was found to be 1:1. Transdermal delivery of aPDL1 antibody significantly inhibited melanoma tumor growth in mice, outperforming intravenous injection in terms of efficacy and reduced systemic toxicity. Co-delivery of aPDL1 and aCTLA4 via FCS resulted in enhanced antitumor activity against both primary and distant tumors (abscopal effect). Transdermal delivery of the SARS-CoV-2 vaccine elicited comparable humoral immunity and stronger cellular immunity compared to subcutaneous injection. Mechanistically, FCS was found to temporarily disrupt tight junctions in the epidermis, facilitating intercellular penetration. The transappendageal route also contributed to delivery. The study demonstrated minimal transcellular transport. No significant cytotoxicity was observed for FCS.

The findings demonstrate that the FCS-based transdermal delivery system successfully addresses the limitations of current approaches. The non-invasive nature, high efficiency, and broad applicability to various biomacromolecules and animal models are significant advancements. The superior antitumor efficacy of transdermal aPDL1 delivery compared to intravenous injection highlights the potential for reduced side effects. The enhanced cellular immunity observed with transdermal vaccine delivery suggests improved immune memory. The mechanism of action, involving both intercellular and transappendageal pathways, contributes to the success of the system. The study provides a compelling platform for localized immunotherapy and self-administered vaccines.

This study presents a novel and highly effective transdermal delivery system based on fluorocarbon-modified chitosan (FCS). The system demonstrates superior efficacy for delivering biomacromolecules compared to existing methods, showcasing its potential for localized melanoma immunotherapy and transdermal vaccination. Future research should focus on optimizing the formulation for various biomacromolecules and conducting larger clinical trials to validate the findings and assess long-term efficacy and safety.

The study primarily used murine models. While the rabbit and porcine skin models provide some evidence of broader applicability, further studies in larger animal models and humans are needed. The proof-of-concept study with the SARS-CoV-2 vaccine did not include a virus challenge experiment to assess protective efficacy. Long-term safety studies are also necessary before clinical translation.