Skin cancer, including squamous cell carcinoma (SCC) and cutaneous melanoma (CM), is highly prevalent and strongly associated with ultraviolet radiation exposure. Limitations of conventional therapies highlight the need for new therapeutic strategies, and metal-based drugs have emerged as promising alternatives due to their anticancer potential. However, their topical application is limited by the skin barrier, and the association with the monoterpene carvone, a well-described skin permeation enhancer, represents a strategy to improve local therapeutic efficacy.

To evaluate the in vitro combination index (CI), apoptosis and the cutaneous permeation of a silver complex derived from nucleobases (Ag-1) associated with carvone.

The Ag-1 complex was synthesized and quantified at the Institute of Chemistry, University of Campinas, and carvone was obtained commercially. At the School of Medical Sciences, University of Campinas, SCC hypopharynx (FaDu), tongue (SCC-9) and melanoma (SK-MEL-28) cells were treated with Ag-1 (0.8–12 µM), alone or combined with carvone. Cell viability was assessed by the sulforhodamine B assay, and the CI was calculated as the ratio between the 50% growth inhibition (GI50) of the Ag-1 + carvone/GI50 of the Ag-1 alone, using the Chou-Talalay method. The phosphatidylserine externalization analysis was performed using the Annexin V Apoptosis Kit, which includes annexin V and 7-AAD conjugates, followed by flow cytometric analysis using a BD FACSVerse™ system. In vitro permeation studies were conducted using Franz diffusion cells with poly (vinyl alcohol) (PVA, 3% w/v) films applied onto polytetrafluoroethylene (PTFE) membranes. Samples were collected at 0h, 12h, 24h, and 48h, and silver permeation was quantified by inductively coupled plasma mass spectrometry (ICP-MS), monitoring the 107Ag isotope.

Ag-1 in combination with carvone resulted in synergistic effect in the FaDu cell line with GI50 < 2 µM, CI = 0.8, SCC-9 GI50 < 4 µM, CI = 0.3 and SK-MEL-28 with GI50 < 6 µM, CI = 0.6). The combination of Ag-1 with carvone increased early apoptotic cell death (annexin V?/7-AAD?) compared to untreated group in FaDu, SK-MEL-28 and SCC-9 (26.7 ± 12.6 versus 55.5 ± 11.8, 27.9 ± 11.6 versus 59.8 ± 13.6, 12.2 ± 10.8 versus 57.3 ± 11.6, respectively; p < 0.05). Permeated silver concentrations were higher for the carvone-containing (CC) formulation compared to the carvone-free (CF) formulation at T0 (114.04 ± 1.52 µg L-1 versus 53.39 ± 3.52 µg L-1, respectively; p < 0.05), T24 (230.76 ± 11.93 µg L-1 versus 130.65 ± 43.46 µg L-1, respectively; p < 0.01) and T48 (359.17 ± 46.53 µg L-1 versus 164.43 ± 40.55 µg L-1, respectively; p < 0.001) hours.

Ag-1 combined with carvone showed synergistic anticancer activity, promoted controlled early apoptotic cell death and enhanced silver permeation, supporting its potential as a topical therapeutic strategy for skin cancer. Further in vivo analyses are necessary.