ID4 acts as a tumor suppressor in ER+ breast tumors.

LAURITO, SERGIO; URRUTIA, GUILLERMO; ROQUÉ, MARÍA; NASIF, DANIELA; CAMPOY, EMANUEL MARTIN

Mar del Plata

Congreso; LXI Reunión Científica Anual de la SAIC; 2016

Sociedad Argentina de Investigación clinica

Introduction: Inhibitor of differentiation proteins 1, 2, 3 and 4 (ID1?4), are dominant negativeregulators of the basic helix-loop helix (bHLH) family of transcription factors. In human tumors, anincreased expression of ID proteins has been associated with reversion to an embryonic-like state,with loss of differentiation, high rates of proliferation, migration and neo-angiogenesis[1]. In breastcancer there are apparently controversial findings regarding the role of ID4 in tumorigenesis. Forinstance, ID4 silencing by promoter hypermethylation is a frequent event in ER+ (estrogen receptor)breast tumors and is associated with an increased risk of lymph node metastasis [2]. On the contrary,in ER- breast tumors ID4 increased expression has been associated with the ability of breast cancercells to exhibit anchorage-independent growth. Our group has previously shown that ID4 promoterunmethylation is associated with the aggressive Triple Negative Breast cancer subtype. We alsodemonstrated that ID4 unmethylation and expression is associated with BRCAness phenotype anddownregulation of BRCA1 gene [3, 4]. Based on our observations and on literature findings it seemsthat ID4 has a dual role in breast cancer. Here, we hypothesize that ID4 acts as a tumor suppressorin ER+ breast tumors.Methodologies: To test our hypothesis we used two approaches: data mining analyses from the CancerGenome Atlas (TCGA) database and experimental cell culture experiments in MCF-7 and T47D celllines. Data mining analyses were performed by retrieving information from Mexpress web page formethylation analyses and Xena (UCSA) platforms for expression analyses. Cell culture experimentswere performed on MCF-7 and T47D cell lines. Both cell lines are ER+ and have been shown by usand others to have methylated the promoter of ID4 and do not express the gene [4]. Briefly, 3ug ofthe pCMV-ID4 vector and 3ug of control vectors (pCMV alone or GFP) were transfected in subconfluent(80%) cells grown in six well plates using Lipofectamine 2000. Forty-eight hours aftertransfection, the cells with incorporated pCMV-ID4 or control plasmids were treated as previouslydescribed, according to the experiments to be performed: proliferation, apoptosis, cell cycle ormigration [5].Results: Data mining of published microarray databases was used to determine the relative expressionlevels of ID4 in breast tumors. As shown in figure 1A, the expression of ID4 is significantly decreasedin ER+ breast tumors with respect to ER- tumors (p