Tumor Suppressor Mir-16 Mediates Trastuzumab Therapeutic Effects in Breast Cancer Via Its Novel Targets, Cyclin J (CCNJ) and Fuse Binding Protein 1 (FUBP1)

LEANDRO VENTURUTTI; ROSALIA INES CORDO RUSSO; PATRICIO YANKILEVICH; ROBERT H OAKLEY; TIM H-M HUANG; ROXANA SCHILLACI; JOHN A CIDLOWSKI; PATRICIA VIRGINIA ELIZALDE

Chicago

Encuentro; Endocrine Society's 96th Annual Meeting and Expo; 2014

Endocrine Society

Este poster obtuvo los premios "ICE Travel Fellowship Award" y "Presidential Poster Competition Winner". Breast cancer (BC) displaying gene amplification and/or overexpression of ErbB-2 (ErbB-2-positive subtype), a member of the ErbBs family of receptor tyrosine kinases, is associated with increased metastatic potential and poor prognosis. The most commonly used therapeutic option for ErbB-2-positive BC is the antibody trastuzumab (TZ). However, many patients show de novo or acquired TZ resistance. On the other hand, increasing evidence has revealed the involvement of microRNAs (miRNAs) in BC cancer growth and metastasis. We previously identified a novel role for miR-16 as a tumor suppressor in progestin-induced BC progression (1). We also reported that miR-16 is up-regulated upon TZ treatment in TZ-sensitive, but not in TZ-resistant BC cell lines (2). In addition, we showed that miR-16 overexpression could serve as an alternative therapy for TZ-resistant BC, and that miR-16-induced cyclin E (CCNE) down-regulation could account for some of its antiproliferative effects (2). Here, our first aim was to elucidate the mechanism by which TZ up-regulates miR-16 expression in TZ-sensitive BC cells. Recruitment of c-Myc to its response elements (E-boxes) in the DLEU2 (miR-16 host gene) proximal promoter has been found by us and others to be a mechanism underlying miR-16 repression. Our chromatin immunoprecipitation studies showed that TZ blocked c-Myc loading at the miR-16 promoter, thereby increasing miR-16 expression. In addition, TZ repressed c-Myc expression. Implicated in these effects was inhibition by TZ of the Phosphatidylinositol 3-phosphate (PI-3K)/AKT and p42/p44 mitogen-activated kinases (MAPK)pathways, which control c-Myc transcriptional activity and stability by modulating its phosphorylation status. Interestingly, TZ was unable to block c-Myc recruitment to the DLEU2promoter or to induce c-Myc downregulation in cells with intrinsic or acquired resistance. Our second aim was to identify novel miR-16 targets which, along with CCNE, could account for miR-16 effects on BC growth. We performed a whole-genome gene expression microarray using ErbB-2 overexpressing BC cells transfected with a miR-16 precursor. Among the top down-regulated genes, we found cyclin J (CCNJ) and FUSE Binding Protein 1 (FUBP1), both of which were predicted as miR-16 targets by bioinformatics analysis. TZ induced CCNJ and FUBP1 down-regulation in TZ-sensitive BC cells, but not in unresponsive cells. In addition, silencing CCNJ or FUBP1 by using siRNAs abrogated proliferation of TZ-sensitive and -resistant BC cells. Our study demonstrates that TZ enhancement of miR-16 expression is mediated by c-Myc, and identifies two novel miR-16 targets underlying its antiproliferative effects.