Runx2 is required for activity of CD44+/CD24-/low breast cancer stem cell in breast cancer development.

Runx2, a master regulator of osteogenesis, is abnormally expressed in advanced breast cancer. Here we addressed Runx2 contribution to breast cancer cell growth and metastasis. We found that CD44 and Runx2 were both elevated in breast cancer tissues compared with the adjacent normal tissues in breast cancer patients. Runx2 expression was significantly correlated with tumor TNM stage, metastasis and poor prognosis. We then screened several breast cancer cell lines and found that Runx2 expression level was positively related to the malignant level of the cells screened. Knockdown of Runx2 in high metastatic cell line MDA-MB-231 could inhibit breast cancer cell vitality, invasion and clone formation capacity, while overexpression of Runx2 in low metastatic cell line MCF-7 could increase those malignant behaviors. The mechanism might be due to Runx2 positively regulating cancer stem cell properties, as CD44 expression level and CD44+/CD24-/low breast cancer stem cell population were both significantly decreased in Runx2 knockdown cells. Cancer stem cell renewal ability such as soft agar clone formation, mammospheres formation and tumor formation ability in null mice were all decreased after knockdown of Runx2. On the contrary, overexpression of Runx2 could enhance all above stem cell renewal ability. Lastly, we explored how Runx2 changes cancer stem cell population. We found it could affect epithelial mesenchymal transition (EMT). Runx2 could regulate mesenchymal marker and epithelial marker expression and affect activation of Wnt/β-catenin signaling pathway. These results together strongly suggest that Runx2 can promote CD44+/CD24-/low breast cancer stem cell properties and breast cancer tumorigenesis through EMT process.

[1]  Maryam Zare,et al.  Aberrant miRNA promoter methylation and EMT‐involving miRNAs in breast cancer metastasis: Diagnosis and therapeutic implications , 2018, Journal of cellular physiology.

[2]  Hao-Yu Hsieh,et al.  Targeting breast cancer stem cells by novel HDAC3-selective inhibitors. , 2017, European journal of medicinal chemistry.

[3]  B. Arumugam,et al.  Role of Runx2 in breast cancer-mediated bone metastasis. , 2017, International journal of biological macromolecules.

[4]  J. Pratap,et al.  Role of Runx2 in IGF-1Rβ/Akt- and AMPK/Erk-dependent growth, survival and sensitivity towards metformin in breast cancer bone metastasis , 2016, Oncogene.

[5]  Peng Zhang,et al.  Transcription factor RUNX2 up-regulates chemokine receptor CXCR4 to promote invasive and metastatic potentials of human gastric cancer , 2016, Oncotarget.

[6]  L. Fabris,et al.  Epithelial-to-Mesenchymal Transition and Cancer Invasiveness: What Can We Learn from Cholangiocarcinoma? , 2015, Journal of clinical medicine.

[7]  Yan Sun,et al.  ITGBL1 Is a Runx2 Transcriptional Target and Promotes Breast Cancer Bone Metastasis by Activating the TGFβ Signaling Pathway. , 2015, Cancer research.

[8]  R. Weinberg,et al.  Distinct EMT programs control normal mammary stem cells and tumour-initiating cells , 2015, Nature.

[9]  G. Christofori,et al.  The relevance of EMT in breast cancer metastasis: Correlation or causality? , 2015, FEBS letters.

[10]  A. Jemal,et al.  Global cancer statistics, 2012 , 2015, CA: a cancer journal for clinicians.

[11]  Jianjiang Fu,et al.  Codonolactone, a sesquiterpene lactone isolated from Chloranthus henryi Hemsl, inhibits breast cancer cell invasion, migration and metastasis by downregulating the transcriptional activity of Runx2. , 2014, International journal of oncology.

[12]  Annette Meeson,et al.  Epithelial-to-mesenchymal transition: what is the impact on breast cancer stem cells and drug resistance. , 2014, Cancer treatment reviews.

[13]  Samy Lamouille,et al.  Molecular mechanisms of epithelial–mesenchymal transition , 2014, Nature Reviews Molecular Cell Biology.

[14]  Joanna S Morris,et al.  RUNX2 in mammary gland development and breast cancer , 2013, Journal of cellular physiology.

[15]  V. Castronovo,et al.  Both osteopontin‐c and osteopontin‐b splicing isoforms exert pro‐tumorigenic roles in prostate cancer cells , 2012, The Prostate.

[16]  H. Im,et al.  Targeting Runx2 expression in hypertrophic chondrocytes impairs endochondral ossification during early skeletal development , 2012, Journal of cellular physiology.

[17]  Guojun Wu,et al.  Putative function of TAP63α during endochondral bone formation. , 2012, Gene.

[18]  G. Coetzee,et al.  Runx2 transcriptome of prostate cancer cells: insights into invasiveness and bone metastasis , 2010, Molecular Cancer.

[19]  T. Stein,et al.  Runx2 in normal tissues and cancer cells: A developing story. , 2010, Blood cells, molecules & diseases.

[20]  Dianqing Wu,et al.  GSK3: a multifaceted kinase in Wnt signaling. , 2010, Trends in biochemical sciences.

[21]  G. Stein,et al.  Runx2 Association with Progression of Prostate Cancer in Patients: Mechanisms Mediating Bone Osteolysis and Osteoblastic Metastatic Lesions , 2009, Oncogene.

[22]  G. Stein,et al.  Ectopic runx2 expression in mammary epithelial cells disrupts formation of normal acini structure: implications for breast cancer progression. , 2009, Cancer research.

[23]  J. Gordon,et al.  Bone sialoprotein stimulates focal adhesion‐related signaling pathways: Role in migration and survival of breast and prostate cancer cells , 2009, Journal of cellular biochemistry.

[24]  M. Zvelebil,et al.  Transcriptome analysis of mammary epithelial subpopulations identifies novel determinants of lineage commitment and cell fate , 2008, BMC Genomics.

[25]  S. K. Zaidi,et al.  Runx2 deficiency and defective subnuclear targeting bypass senescence to promote immortalization and tumorigenic potential , 2007, Proceedings of the National Academy of Sciences.

[26]  Haiyan I. Li,et al.  Purification and unique properties of mammary epithelial stem cells , 2006, Nature.

[27]  François Vaillant,et al.  Generation of a functional mammary gland from a single stem cell , 2006, Nature.

[28]  G. Stein,et al.  The Runx2 Osteogenic Transcription Factor Regulates Matrix Metalloproteinase 9 in Bone Metastatic Cancer Cells and Controls Cell Invasion , 2005, Molecular and Cellular Biology.

[29]  K. Akashi,et al.  Loss of Runx1 perturbs adult hematopoiesis and is associated with a myeloproliferative phenotype. , 2004, Blood.

[30]  Kazuhiro Yoshida,et al.  Frequent Loss of RUNX3 Expression by Promoter Hypermethylation in Gastric Carcinoma , 2004, Pathobiology.

[31]  Yoram Groner,et al.  Runx3 and Runx1 are required for CD8 T cell development during thymopoiesis , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[32]  J. Lotem,et al.  Phylogenesis and regulated expression of the RUNT domain transcription factors RUNX1 and RUNX3. , 2003, Blood cells, molecules & diseases.

[33]  Y. Oshika,et al.  P-glycoprotein-mediated acquired multidrug resistance of human lung cancer cells in vivo. , 1996, British Journal of Cancer.

[34]  R. Calogero,et al.  Immunotargeting of Antigen xCT Attenuates Stem-like Cell Behavior and Metastatic Progression in Breast Cancer. , 2016, Cancer research.

[35]  D. Jurkovičová,et al.  Evaluation of expression profiles of microRNAs and two target genes, FOXO3a and RUNX2, effectively supports diagnostics and therapy predictions in breast cancer. , 2016, Neoplasma.

[36]  Simon G. Coetzee,et al.  Title: Opposing effects of Runx2 and estradiol on breast cancer cell proliferation: in vitro identification of reciprocally-regulated gene signature related to clinical letrozole responsiveness , 2011 .

[37]  C. Lynch.  Matrix metalloproteinases as master regulators of the vicious cycle of bone metastasis. , 2011, Bone.

[38]  A. Jemal,et al.  Global cancer statistics , 2011, CA: a cancer journal for clinicians.

[39]  Toshihisa Komori,et al.  Regulation of bone development and extracellular matrix protein genes by RUNX2 , 2009, Cell and Tissue Research.

[40]  F. Lo Coco,et al.  The AML1 gene: a transcription factor involved in the pathogenesis of myeloid and lymphoid leukemias. , 1997, Haematologica.