Oncogene (2013) 32, 2121–2130 & 2013 Macmillan Publishers Limited All rights reserved 0950-9232/13 www.nature.com/onc REVIEW The RUNX family in breast cancer: relationships with estrogen signaling N-O Chimge1 and B Frenkel2 The three RUNX family members are lineage specific master regulators, which also have important, context-dependent roles in carcinogenesis as either tumor suppressors or oncogenes. Here we review evidence for such roles in breast cancer (BCa). RUNX1, the predominant RUNX family member in breast epithelial cells, has a tumor suppressor role reflected by many somatic mutations found in primary tumor biopsies. The classical tumor suppressor gene RUNX3 does not consist of such a mutation hot spot, but it too seems to inhibit BCa; it is often inactivated in human BCa tumors and its haploinsufficiency in mice leads to spontaneous BCa development. The tumor suppressor activities of RUNX1 and RUNX3 are mediated in part by antagonism of estrogen signaling, a feature recently attributed to RUNX2 as well. Paradoxically, however RUNX2, a master osteoblast regulator, has been implicated in various aspects of metastasis in general and bone metastasis in particular. Reciprocating the anti-estrogenic tumor suppressor activity of RUNX proteins, inhibition of RUNX2 by estrogens may help explain their context-dependent anti-metastatic roles. Such roles are reserved to non-osseous metastasis, because ERa is associated with increased, not decreased skeletal dissemination of BCa cells. Finally, based on diverse expression patterns in BCa subtypes, the successful use of future RUNX-based therapies will most likely require careful patient selection. Oncogene (2013) 32, 2121–2130; doi:10.1038/onc.2012.328; published online 8 October 2012 Keywords: RUNX1; RUNX2; RUNX3; tumor suppressor; oncogene; ERa The mammalian RUNX family of transcription factors comprises diverse origins have been extensively reviewed,21–24 and Table 1 three members, RUNX1, RUNX2 and RUNX3, which have pivotal summarizes reported abnormalities implicating each of the RUNX roles in both normal development and neoplasia.1 Despite their proteins as a tumor suppressor in various cancer types. structural similarities, RUNX proteins have divergent physiological roles; RUNX1 is required for definitive hematopoiesis,2 RUNX2 has fundamental roles in the differentiation of osteoblasts and RUNX1 AS A BCa SUPPRESSOR chondrocytes,3–5 and RUNX3 is involved in gastrointestinal and 6–8 In addition to their most notable roles in hematopoietic cells, bone neuronal development. Inherited mutations in RUNX1 are the cells and neurons, the three RUNX genes are also expressed in the cause of familial platelet disorder with associated myeloid normal mammary gland, and their mRNA levels fluctuate during malignancy,9 and RUNX2 mutations cause the dominant 25 10 cycles of pregnancy, lactation and involution. RUNX1 is the autosomal skeletal disease cleidocranial dysplasia. predominant RUNX family member expressed in human breast In cancer, RUNX act as either tumor suppressors or oncogenes 13,26,27 1,11 epithelial cells (Figure 1a) and the protein is readily detected depending on cellular context. RUNX1 is a frequent target in both the basal and the luminal cell layers.26 The most of translocations and other mutations in hematopoietic maligna- compelling evidence for its role in inhibiting BCa is the somatic ncies, and RUNX3, located on chromosome 1p36, resides mutations recently observed in RUNX1 (and not RUNX2 or RUNX3) in a deletion hotspot found in diverse cancers of epithelial, 28 12 in tumors from the Washington University BRC77 study and in hematopoietic and neuronal origins. Tumor suppressor activity the cohort of BCa patients studied in The Cancer Genome Atlas in breast cancer (BCa) has been attributed to all three RUNX pilot project29 (Figure 1b). Furthermore, RUNX1 was one of 17 proteins,13–15 whereas RUNX2 is increasingly recognized as pro- 30 11,16–20 genes whose expression pattern predicted BCa metastasis; its metastatic in breast (and prostate) cancer progression. Here expression is less abundant in BCa compared to normal breast we review the dual functions of RUNX proteins in BCa and their epithelial cells, and it progressively decreases with increasing relationships with estrogen signaling. BCa tumor aggression.31 Finally, experimental Ras-mediated transformation of basal-like MCF10A cells was associated with loss of RUNX1;31 and RUNX1 silencing in another study with RUNX PROTEINS ARE TUMOR SUPPRESSORS MCF10A cells led to the formation of abnormal, hyperproliferative All three mammalian RUNX family members have been implicated acinar structures in 3D cultures.13 in tumor suppression. The frequent functional inactivation of How RUNX1 suppresses BCa remains to be elucidated. Among RUNX1 and RUNX3 observed in hematopoietic and solid tumors of possible mechanisms, RUNX1 may stimulate E-cadherin to inhibit 1Department of Biochemistry and Molecular Biology, Institute for Genetic Medicine, Keck School of Medicine of the University of Southern California, Los Angeles, CA, USA and 2Departments of Orthopaedic Surgery and Biochemistry and Molecular Biology, Institute for Genetic Medicine, Keck School of Medicine of the University of Southern California, Los Angeles, CA, USA. Correspondence: Dr N-O Chimge or Dr B Frenkel, Department of Biochemistry and Molecular Biology, University of Southern California, Institute for Genetic Medicine, 2250 Alcazar Street, CSC-240, Los Angeles, CA 90033, USA. E-mail: [email protected] or [email protected] Received 3 May 2012; revised 20 June 2012; accepted 20 June 2012; published online 8 October 2012 RUNX1,2,3 in breast cancer N-O Chimge and B Frenkel 2122 Table 1. Tumor suppressor activities of the RUNX family in various cancers RUNX Types of cancer Related mechanisms References RUNX1 AML-M2 RUNX1-ETO: t(8;21)(q22;q22) 137–140 RUNX1-LRP16: t(11;21)(q13;q22) RUNX1-MTG16: t(16;21)(q24;q22) RUNX1-others Mut 141,142 MDS RUNX1-EVI: t(3;21)(q26.2;q22) 140,143–149 RUNX1-MDS1: t(3;21)(q26.2;q22) RUNX1-EAP: t(3;21)(q26.2;q22) RUNX1-PRDM6: t(1;21)(p36;q22) RUNX1-others Mut 150–156 B-ALL TEL-RUNX1: t(12;21)(p13;q22) 157–160 FPDMM Hemizygous deletion, SNP 9,161 AML-M1 Mut 162 AML-M0 LOH, biallelic RUNX1 mut 162–164 CMML Mut 165,166 Colorectal Mut 167 Gastointestinal tract Mut 168,169 Esophageal squamous-cell carcinoma SNP 170,171 Prostate SNP 172 Breast Mut 28,29 RUNX2 Osteosarcoma Inhibition of proliferation 173,174 Breast Antagonism of estrogen signaling 15 RUNX3 Gastric Hemizygous deletion, Mut, 8,167,169,175–177 hypermethylation, cytoplasmic mislocalization Bladder Mut, hypermethylation 178–180 Prostate Hypermethylation 181 Lung Hemizygous deletion, hypermethylation 182 Pancreatic Hemizygous deletion, hypermethylation 183 Colorectal Hypermethylation, Mut, cytoplasmic mislocalization 184–187 Brain Hypermethylation 188 Squamous cell carcinoma Cytoplasmic mislocalization 189 Hepatocellular carcinoma Hypermethylation 37,190 Bile duct Hemizygous deletion 183 Renal cell carcinoma Inhibition of proliferation 191 Bone Hypermethylation 192 Breast Hypermethylation, 14,34,35 cytoplasmic mislocalization Abbreviations: AML, acute myeloblastic leukemia; AML-M0, minimally differentiated acute myeloblastic leukemia; AML-M1, acute myeloblastic leukemia without maturation; CMML, chronic myelomonocytic leukemia; FPDMM, familial platelet disorder with associated myeloid malignancy; LOH, loss of heterozygosity; MDS, myelodysplastic syndrome; Mut, mutations other than SNPs or translocations; SNP, single-nucleotide polymorphism; t, translocation. Studies of breast cancer are in bold text. epithelial-to-mesenchymal transition (EMT)32 and it may increased estradiol signaling owing to partial loss of ERa cooperate with FOXO signaling to protect ER-negative BCa cells destabilization by RUNX314 (Figure 2a). Expression of RUNX3 from oxidative stress.13 In ER-positive cells, however, we believe decreases during human BCa progression and high RUNX3 RUNX1 attenuates oncogenic effects of estrogens (Figure 2a) levels are associated with favorable prognosis.35 As in other mainly through physical interaction with ERa.33 In fact, all but one malignancies, RUNX3 is frequently inactivated in BCa by RUNX1 somatic mutations found so far are in ERa-positive promoter hypermethylation, genetic deletion, and protein tumors.28,29 mislocalization.34,35,37–42 RUNX3 promoter hypermethylation in response to estradiol has been observed in mammosphere- derived cells, which may contribute to the oncogenic activity of estradiol in breast epithelial cells.43 RUNX3 AS A BCa SUPPRESSOR Evidence suggesting a role for RUNX3 in inhibiting various malignancies is cited in Table 114,34–36 and the literature on the tumor suppressor role of RUNX3 specifically in BCa has been RUNX2 AS A BCa SUPPRESSOR recently reviewed.24 Most notably, RUNX3 hemizygous female RUNX2 is expressed in the developing mammary gland during mice develop spontaneous ductal carcinoma,14 attributable to embryogenesis5 and puberty, and its mRNA levels in glands of Oncogene (2013) 2121 – 2130 & 2013 Macmillan Publishers Limited RUNX1,2,3 in breast cancer N-O Chimge and B Frenkel 2123 RUNX1: Chr.21q22.3 Scale: 200 kb hg19 RUNX1 RUNX1 RUNX1 Brain Heart Lymph Node Breast HMEC RUNX2: Chr.6p21 Scale: 200 kb hg19 RUNX2 RUNX2 RUNX2 Brain Heart Lymph Node Breast HMEC RUNX3: Chr.1p36 Scale: 200 kb hg19 RUNX3 RUNX3 Brain Heart Lymph Node Breast HMEC frame shift deletion frame shift insertion RUNX1 in-frame deletion missense nonsense F259fs Q266* L288fs L317F 322e7+2