Abstract 2977: Malignant transformation associated with repetitive Ras/Rac1 activation, increased production of superoxide anion, and global DNA methylation alterations.

Abstract Driver mutations in N-Ras and BRAF, and more recently in Rac1, were described in melanomas. Using a malignant transformation model of murine melanocytes developed in our laboratory, in which melanoma cell lines were obtained after submitting the melanocyte lineage melan-a to sequential cycles of anchorage blockade, we identified dysfunction in both N-Ras and Rac1 during stressful condition. Our group has already showed increased global DNA methylation as well increased DNA methyltransferase 1 (Dnmt1) protein level few hours after anchorage blockade. This increase seems to be related with elevated amounts of superoxide anion. Previous works have showed that both Ras may regulate superoxide level as Ras activation may affect DNA methylation. Therefore, we aimed to study the relation among Ras/MEK/ERK signaling pathway, oxidative stress and DNA methylation during melan-a anchorage blockade. We showed that Ras, Rac1 and ERK are activated few hours after melan-a anchorage blockade and inhibiting this activation abrogates superoxide increase. Furthermore, global DNA methylation levels decrease during anchorage blockade after treating cells with superoxide scavenger or Ras inhibitor (FTI). Dnmt1 protein level is also decreased after treating melan-a cells during anchorage blockade with FTI or ERK inhibitor (U0126). Ras/MEK/ERK pathway seems to be regulated by and regulates superoxide production during melan-a anchorage restriction. In this way, superoxide anion depletion impairs Ras and ERK activation and FTI or U0126 treatment decreases superoxide during adhesion impediment. This aberrant signaling seems to have a significant impact in melanoma genesis since the malignant transformation was drastically compromised when melan-a melanocytes were pre-treated with superoxide scavenger before each anchorage impediment cycle. Elucidating the relation among oxidative stress, epigenetic changes and melanocyte malignant transformation may provide novel avenues to develop alternative therapeutic approaches. Citation Format: Fernanda Molognoni, Fabiana M. Melo, Camila Tainah, Miriam G. Jasiulionis. Malignant transformation associated with repetitive Ras/Rac1 activation, increased production of superoxide anion, and global DNA methylation alterations. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2977. doi:10.1158/1538-7445.AM2013-2977