Prevention and treatment of dermatological diseases: optimizing the protective action and efficacy of phenylpropanoid glycosides delivered through the skin by inclusion in biodegradable nanovesicles

The skin, the human body’s largest organ, serves critical roles in defense and physiological regulation. Maintaining its integrity is essential, yet it is constantly challenged by environmental factors such as ultraviolet radiation (UVR), extreme temperatures, and chemical pollutants. As a "complete carcinogen," UVR has the unique capacity to both initiate and promote tumor development, making it a significant contributor to skin cancer, with melanoma being the most aggressive and lethal form. The global incidence of melanoma continues to rise. Although targeted therapies and immunotherapies have advanced, clinical outcomes remain suboptimal due to tumor resistance and adverse effects, underscoring the need for innovative treatment strategies. Natural antioxidants, including Phenylpropanoid Glycosides (PPGs) and Hyperforin (HPF), show promising antitumor potential among emerging thera peutic agents. PPGs, a group of over 200 plant-derived polyphenols, and HPF, an acylphloroglucinol extracted from Hypericum perforatum, exhibit comple mentary pharmacological properties, such as antioxidant, anti-inflammatory, and antitumor activities. This thesis examines the dual role of the PPGs, particularly Echinacoside and Forsythoside B, in preventing skin tumors and treating melanoma and other skin cancers. To explore the protective properties of PPGs against UVR, normal hu man epidermal melanocytes (NHEM) and normal human dermal fibroblasts (NHDF) were pre-treated with these compounds. Solar UV radiation was then simulated by exposing cultured cells to a UV lamp. The protective effects of PPGs pre-treatment were analyzed by assessing cell viability and the activation of multiple stress signaling pathways. Our findings suggest that PPGs exert a mild UV protective effect in normal cells, although statistical significance was not consistently observed. To improve their stability and bioavailability under UV and temperature exposure, PPGs were nanoencapsulated. In collaboration with the company “Nanomnia”, a natural, biodegradable polymer-based process was employed, eliminating microplastics and enhancing targeted delivery and controlled release. This approach can also mitigate the environmental impact of conven tional sunscreen components, which are associated with marine ecosystem disruption. Preliminary studies revealed a mild UV protective effect in normal cells from encapsulated PPGs. Particular attention is given to the antitumor properties of Echinacoside and Forsythoside B against BRAF-mutated melanoma cell line (FO-1) and a human skin tumor cell line (A-431). The results reveal that these compounds selectively induce cytotoxicity in malignant cells without affecting normal f ibroblasts, keratinocytes, or melanocytes, even at concentrations four times higher than their EC50 values in cancer cells. Within 48 h of in vitro administration, PPGs promote apoptosis by cleaving Poly (ADP-ribose) Polymerase-1 (PARP-1), activating Ataxia-Telangiectasia Mutated (ATM) kinase, and increasing phosphorylated H2A histone family member X (γH2AX) levels, indicating DNA damage. Additionally, the phosphorylation of p38 mitogen-activated protein kinases (MAPK) suggests that stress response signaling pathways are involved. These findings demonstrate a slight protective effect on normal skin cells following UV stress, as well as the selective cytotoxicity and apoptotic potential of Echinacoside and Forsythoside B in skin cancer cell lines, highlighting their therapeutic promise in melanoma and other skin cancer. Encapsulated PPGs were evaluated to assess their protective activity on NHDF exposed to UV-A radiation. Preliminary results indicate that lower concentrations of encapsulated PPGs revealed a remarkable protective response through morphological analysis. However, quantitative analysis need to be carried out. Furthermore, Hyperforin (HPF), an acylphloroglucinol derived from Hy pericum perforatum, exhibits a variety of pharmacological activities, including antidepressant, anti-inflammatory, antimicrobial, and antitumor effects. Our f indings demonstrate that HPF exerts potent antitumor activity against BRAF-mutated melanoma cell lines (A375, FO-1, SK-Mel-28), including inhibition of proliferation, motility, colony formation, and induction of apop tosis. Its cytotoxic mechanisms involve lipid peroxidation and disruption of iron homeostasis through downregulation of glutathione peroxidase-4 (GPX 4), cystine transporter SLC7A11, and ferritin, thereby increasing cellular susceptibility to oxidative damage. Concurrently, HPF upregulates heme oxygenase-1 (HO-1), which promotes the release of free iron. Cytofluorimetric analysis confirmed an increase in lipid peroxidation, as evidenced by enhanced BODIPY-C11 fluorescence. HPF also induces autophagy, as indicated by elevated LC3B expression. Moreover, HPF exerts cytostatic effects by modulating key cell cycle regulators, including downregulation of cyclins D1 and A2, CDK4, and phos phorylated Rb, and upregulation of P21/waf1 and activated P53. Apoptotic effects are further enhanced by reductions in Bcl-2 and Bcl-xL levels and increased cleaved PARP-1. Furthermore, HPF alters the metabolism of melanoma cells inhibiting mitochondrial enzymes expression and compro mising the mitochondrial membrane potential. Although HPF modulates TRPC6 channels and elevates cytosolic Ca2+ and Zn2+ levels, its cytostatic effects appear largely independent of these pathways. Furthermore, HPF suppresses survival signaling by reducing the activity of transcription factors nuclear factor-kappa B (NF-κB) P65 and Signal Transducer and Activator of Transcription 3 (STAT3). HPF’s antimetastatic properties further reinforce its therapeutic potential. It downregulates many invasion markers, consistent with observed reductions in cell migration and colony formation. Collectively, these findings underscore HPF’s pleiotropic antitumor properties, includ ing cytostatic, cytotoxic, and antimetastatic effects, thereby supporting its potential as a therapeutic agent in melanoma management. These findings collectively highlight the therapeutic potential of PPGs and HPF. While PPGs demonstrate a slight UV-protective effect on normal f ibroblasts and melanocytes and a selective cytotoxic effect on malignant skin cells, HPF exerts broader cytostatic, cytotoxic, and antimetastatic activities on melanoma cell lines, providing a strong foundation for future translational research and therapeutic applications.