CONICET | Buscador de Institutos y Recursos Humanos

We have previously demonstrated that TAP7f, a synthetic derivative formed by penicillin linked to the dipeptide Leu-Phe through a triazole group, inhibited melanoma metastasis by downregulating β-catenin. Since it has been reported that melanoma cells resistant to BRAF inhibitors (BRAFi) express higher levels of β-catenin, blocking Wnt/ β-catenin signaling could be considered a useful strategy to improve cell response. In this study we examined the mechanism of action of TAP7f in melanoma cells sensitive (A375, BRAFV600E) and resistant (SB2, NRASQ61K) to BRAFi. The cytotoxic potency of TAP7f on these cell lines was first determined, being IC50 values of 10.0 ± 1 µM and 9.0 ± 3 µM for A375 and SB2 cells, respectively (72 h, n=3). In addition, IC50 values of 7.7 ± 0.5 nM and 2.1 ± 0.9 nM were obtained for dabrafenib (D, BRAFi) and Trametinib (T, MEKi) in A375 cells, whereas SB2 cells were more resistant to these inhibitors (IC50 D: > 30 µM; IC50 T: > 20 nM). Western Blot assays revealed that TAP7f (20 µM) reduced 67.5 ± 0.6 % and 59.1 ± 0.1 % the expression levels of β-catenin in A375 and SB2 cells, respectively, after 24 h of incubation. Under similar experimental conditions, TAP7f increased the phosphorylation of Ser 33/37 of β-catenin in both cell lines, a target region for β-catenin degradation. Furthermore, when A375 and SB2 melanoma cells were incubated with TAP7f in the presence of MG132, a proteasome inhibitor, a significant reversion of TAP7f inhibitory effect on β-catenin levels was observed. In summary, our results suggest that TAP7f downregulates β-catenin levels by promoting its degradation via proteasome in melanoma cell lines with different genetic profile, regardless of sensitivity to BRFA inhibitors. This behavior positions this penicillin derivative as a promising agent for the treatment of BRAFi-resistant cells.

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