Avaliação da eficácia e segurança de formulações tópicas contendo ativos moduladores das enzimas LOX

Abstract

Lysyl oxidase (LOX) and lysyl oxidase like (LOXL) are enzymes involved in the skin regeneration process. Nanotechnology makes it possible to formulate efficient delivery systems for topical application such as nanostructured lipid carriers (NLC). The present study aimed to explore the application of compounds that modulate LOXL2 enzymes to control skin regeneration. For this purpose, the impact of LOXL2 inhibitor compounds identified by the CBIOS group (8α,9α-epoxycoleon-U-quinone - EPCU and Pimarane 7, 15-isopimaradien-19-ol - PN1) on the viability of human keratinocytes was evaluated. The impact of PN1, EPCU and BAPN on the viability of human keratinocytes (HaCaT cells) was tested using the Crystal Violet method. The IC50 of LOXL2 inhibition of EPCU is 58.3 uM, and in the present study, keratinocytes subjected to a concentration of 50 nM suffered a significant reduction in their viability. Regarding PN1, its LOXL2 inhibition IC50 is 104.4 uM, and from the results obtained in this study, with a maximum concentration of 200 nM, the compound does not impact cell viability, which is promising for future studies. In addition, the carrier compatibility of a commercially available LOXL2 inhibitor compound, β-aminopropionitrile (BAPN) was verified by NLC. Regarding the characterization of nanoparticles (NPs) with and without BAPN, the hydrodynamic diameter (Dh) indicates that the NPs without BAPN were smaller, their polydispersion index (PDI) did not exceed 0.3 in both, and the zeta potential was situated between -18 and -22 mV. NPs with BAPN have a higher pH value compared to NPs without BAPN, and this value slightly increased with storage time. Stability evaluation tests demonstrated that the formulations remained unchanged. The safety and efficacy of NPs without BAPN was tested in a group of human volunteers. With regard to the measurements of skin parameters after topical application, the parameters transepidermal water loss (TEWL), erythema and hydration revealed that the NPs have good compatibility, but the increase in hydration was small. The work led to the conclusion that the maximum concentrations of the tested inhibitors that do not induce changes in cell viability are respectively: 500 nM for BAPN, 20 nM for EPCU and 200 nM for PN1. BAPN was successfully incorporated in NLC formulation, but because it is ionized it reduces its zeta potential, which may have an impact in stability over time. NPs can be considered compatible with the skin. These data show the potential of these compounds as possible modulators of the healing process, which may be conveyed by NPs. Keywords: HaCat cells, BAPN, cell viability, NLC, skin regeneration

Date of Award	2023
Original language	Portuguese
Supervisor	Nuno Saraiva (Supervisor) & Catarina Rosado (Supervisor)