(W0930-01-02) Transdermal Delivery of Ultradeformable Cationic Liposomes Complexed with microRNA Prevents Transformation of Melanocytes to Melanoma

Purpose: Melanoma is a malignant skin cancer resulting from the uncontrolled growth of melanocytes. Upon acquiring an oncogenic driver mutation, BRAFV600E, melanocytes proliferate to form melanocytic nevi, which are nests of growth-arrested melanocytes [1]. The upregulation of miR211-5p, which is a microRNA inhibiting the expression of Aurora Kinase B (AURKB), the mitosis-regulating kinase, has recently been identified to be a key mechanism in preventing the malignant transformation of nevi melanocytes to melanoma [2]. We hypothesize that topical delivery of miR211-5p may serve as a novel chemoprevention strategy to block the transformation of nevi melanocytes to melanoma cells by inhibiting AURKB and melanoma growth. To this end, a transdermal delivery system consisting of ultradeformable cationic liposomes (UCLs)[3] complexed with miR211-5p (UCL-211) as a chemopreventive agent was developed and validated.

Methods: UCLs were synthesized by thin film hydration technique and then complexed with miR211-5p (UCL-211). UCL-211 were characterized and evaluated in vitro for cytotoxicity and uptake in different skin cell lines. Several orthogonal approaches were used to validate the delivery of biologically active miR211-5p, including a sponge reporter assay, live-cell microscopy, and absolute qPCR. Ex vivo permeation studies using a Franz diffusion cell system with de-identified human skin biopsies were performed to assess the permeability of UCLs in the epidermis. A transgenic nevus growth-arrested mouse model was developed to evaluate the efficacy of UCL-211 delivery to melanocytes through topical application.

Results: UCL-211 exhibits an average particle size of 108±0.6nm and an average encapsulation efficiency of 74%. UCLs were cytocompatible with melanocytes, keratinocytes, and fibroblasts, and did not induce any cellular toxicity. Melanocytes showed the highest uptake rate compared to the other two cell lines. In vitro studies were performed to confirm biological activity of miR211-5p delivered by UCL-211 delivery system. Cells treated with UCL-211 showed morphological changes and accumulation of miR211-5p. Ex vivo studies confirmed UCL-211 permeation and localization in skin epidermis. Successful topical application and delivery of UCL-211 to melanocytes were demonstrated in vivo. We observed an increase in nevi in groups treated with UCL-211 compared to controls, and there was no melanoma development in any group.

Conclusion: We showed that UCL-211 liposomal delivery system is able to efficiently deliver miR211-5p to target cells in in vitro, ex vivo, and in vivo models. The animal studies also demonstrated that miR211-5p levels promoted growth of benign nevi.

References: 1. Damsky, W.E. and M. Bosenberg, Melanocytic nevi and melanoma: unraveling a complex relationship. Oncogene, 2017. 36(42): p. 5771-5792.
2. McNeal, A.S., et al., BRAFV600E induces reversible mitotic arrest in human melanocytes via microRNA-mediated suppression of AURKB. eLife, 2021. 10: p. e70385.
3. Dorrani, M., et al., Development of edge-activated liposomes for siRNA delivery to human basal epidermis for melanoma therapy. J Control Release, 2016. 228: p. 150-158.

Acknowledgements: This research was funded by the HCI Melanoma Research Program Seed Grant and the National Cancer Institute (R01CA229896).


Figure 1.a. Synthesis of UCL-211 by thin film hydration technique. b. Particle size analysis by nanoparticle tracking analysis (NTA). c. Transmission electron microscopy (TEM) for UCL-211. d. Quantification of miR211 levels and miR16 levels (control miRNA) using Absolute qPCR in normal human melanocytes. e. Ex vivo permeation studies with deidentified human biopsy- Control and UCL-211. Blue- Hoechst 33342, green- UCL-211, red- Melan-A (melanocyte marker). Yellow arrows: presence of melanocytes. Orange arrows: permeation of UCL-211.


Figure 2. a. BRAFCA: TYR-CREER transgenic nevus growth-arrested mouse model. b. Study design to evaluate the efficacy of UCL-211 delivery to melanocytes through topical application. P- age of pups, T- treatment day, 4HT- 4-hydroxytamoxifen application. c. Images of mice skin collected 21-days after treatment. Control (skin only treated with 4-HT), UCL-complexed with non-targeting miRNA (UCL-NT) and UCL-211.