FAK inhibition suppresses breast cancer progression via DNA methylation-mediated DAB2 gene reactivation

Abstract

Epigenetic silencing of tumor suppressor genes is one of the main drivers of tumor progression. Without these tumor suppressors to reduce proliferation, tumor cells proliferate unchecked. Focal adhesion kinase (FAK) is a tyrosine kinase which is often upregulated in various tumors and promotes cell proliferation and migration. Recent studies have demonstrated that pharmacological or genetic FAK inhibition can reduce suppressive DNA methylation in vascular cells. Mechanistically, this is through nuclear FAK-mediated ubiquitination and proteasomal degradation of DNA methyltransferase 3A (DNMT3A). Treatment of breast cancer cell lines with FAK inhibitor (FAK-I) was able to reduce both FAK activity and DNMT3A protein expression. Further, global DNA methylation was reduced in breast cancer cell lines treated with FAK-I. This decrease in DNA methylation was correlated with decreased cell proliferation. We further showed that FAK-I reduced DNMT3A expression in breast cancer cells and that treatment with the proteasome inhibitor MG132 prevented loss of DNTM3A protein stability. To identify how FAK-I and DNMT3A loss could reduce breast cancer cell growth we compared RNA sequencing data from breast cancer cells treated with or without FAK-I or in shRNA DNMT3A knockdown. We have identified a potential tumor suppressor, DAB2, as being regulated by the nuclear FAK-DNMT3A axis. DAB2 is often downregulated in cancers and has been shown to play a vital role in switching TGF{beta} signaling from proliferative to apoptotic by altering TGF{beta}RI binding partners. Immunoblotting and immunostaining indeed revealed that FAK-I and shDNMT3A could induce DAB2 protein expression. Further, FAK-I treatment showed efficacy in reducing tumor growth in vivo using the murine 4T1 tumor model. Immunostaining of 4T1 tumors showed FAK-I decreased DNMT3A, DNA methylation (5-methylcytosine, 5-mC), and increased DAB2 expression. Taken together, these data suggest that nuclear FAK-mediated regulation of DNMT3A can alter the epigenetic landscape and induce tumor suppressor gene expression.