Breast tumor kinase (Brk/PTK6) is a mediator of hypoxia-associated breast cancer progression.

Basal-type triple-negative breast cancers (TNBC) are aggressive and difficult to treat relative to luminal-type breast cancers. TNBC often express abundant Met receptors and are enriched for transcriptional targets regulated by hypoxia-inducible factor-1α (HIF-1α), which independently predict cancer relapse and increased risk of metastasis. Brk/PTK6 is a critical downstream effector of Met signaling and is required for hepatocyte growth factor (HGF)-induced cell migration. Herein, we examined the regulation of Brk by HIFs in TNBC in vitro and in vivo. Brk mRNA and protein levels are upregulated strongly in vitro by hypoxia, low glucose, and reactive oxygen species. In HIF-silenced cells, Brk expression relied upon both HIF-1α and HIF-2α, which we found to regulate BRK transcription directly. HIF-1α/2α silencing in MDA-MB-231 cells diminished xenograft growth and Brk reexpression reversed this effect. These findings were pursued in vivo by crossing WAP-Brk (FVB) transgenic mice into the MET(Mut) knockin (FVB) model. In this setting, Brk expression augmented MET(Mut)-induced mammary tumor formation and metastasis. Unexpectedly, tumors arising in either MET(Mut) or WAP-Brk × MET(Mut) mice expressed abundant levels of Sik, the mouse homolog of Brk, which conferred increased tumor formation and decreased survival. Taken together, our results identify HIF-1α/2α as novel regulators of Brk expression and suggest that Brk is a key mediator of hypoxia-induced breast cancer progression. Targeting Brk expression or activity may provide an effective means to block the progression of aggressive breast cancers.

[1]  Seung-Taek Lee,et al.  PTK6 promotes degradation of c-Cbl through PTK6-mediated phosphorylation. , 2013, Biochemical and biophysical research communications.

[2]  M. Rosemann,et al.  Effects of Simultaneous Knockdown of HER2 and PTK6 on Malignancy and Tumor Progression in Human Breast Cancer Cells , 2013, Molecular Cancer Research.

[3]  K. Liang,et al.  Brk/PTK6 cooperates with HER2 and Src in regulating breast cancer cell survival and epithelial-to-mesenchymal transition , 2013, Cancer biology & therapy.

[4]  G. Semenza,et al.  Hypoxia-inducible factor 1-dependent expression of platelet-derived growth factor B promotes lymphatic metastasis of hypoxic breast cancer cells , 2012, Proceedings of the National Academy of Sciences.

[5]  S. Fuqua,et al.  Phosphorylated and sumoylation-deficient progesterone receptors drive proliferative gene signatures during breast cancer progression , 2012, Breast Cancer Research.

[6]  G. Kristiansen,et al.  Non-canonical HIF-2α function drives autonomous breast cancer cell growth via an AREG–EGFR/ErbB4 autocrine loop , 2012, Oncogene.

[7]  J. Ingels,et al.  Hypoxia-inducible factor 1α promotes primary tumor growth and tumor-initiating cell activity in breast cancer , 2012, Breast Cancer Research.

[8]  G. Mills,et al.  Brk/PTK6 Sustains Activated EGFR Signaling through Inhibiting EGFR Degradation and Transactivating EGFR , 2011, Oncogene.

[9]  C. Ge,et al.  Physical and functional interactions between Runx2 and HIF‐1α induce vascular endothelial growth factor gene expression , 2011, Journal of cellular biochemistry.

[10]  C. Lange,et al.  Mechanisms of HGF/Met Signaling to Brk and Sam68 in Breast Cancer Progression , 2011, Hormones and Cancer.

[11]  Mark T. W. Ebbert,et al.  Tumor grafts derived from women with breast cancer authentically reflect tumor pathology, growth, metastasis and disease outcomes , 2011, Nature Medicine.

[12]  J. Ostrander,et al.  Mammary gland specific expression of Brk/PTK6 promotes delayed involution and tumor formation associated with activation of p38 MAPK , 2011, Breast Cancer Research.

[13]  G. Semenza,et al.  Hypoxia-inducible factor 1 is a master regulator of breast cancer metastatic niche formation , 2011, Proceedings of the National Academy of Sciences.

[14]  G. Semenza,et al.  HIF-1-dependent Expression of Angiopoietin-like 4 and L1CAM Mediates Vascular Metastasis of Hypoxic Breast Cancer Cells to the Lungs , 2011, Oncogene.

[15]  Jiannis Ragoussis,et al.  High-resolution genome-wide mapping of HIF-binding sites by ChIP-seq. , 2011, Blood.

[16]  Michael L. Gatza,et al.  Analysis of tumor environmental response and oncogenic pathway activation identifies distinct basal and luminal features in HER2-related breast tumor subtypes , 2011, Breast Cancer Research.

[17]  C. Lange,et al.  ck2-Dependent Phosphorylation of Progesterone Receptors (PR) on Ser81 Regulates PR-B Isoform-Specific Target Gene Expression in Breast Cancer Cells , 2011, Molecular and Cellular Biology.

[18]  D. Hanahan,et al.  Hallmarks of Cancer: The Next Generation , 2011, Cell.

[19]  J. Ostrander,et al.  Brk/PTK6 signaling in normal and cancer cell models. , 2010, Current opinion in pharmacology.

[20]  C. Lange,et al.  Breast tumor kinase and extracellular signal-regulated kinase 5 mediate Met receptor signaling to cell migration in breast cancer cells , 2010, Breast Cancer Research.

[21]  W. Hahn,et al.  PTK6 Regulates IGF-1-Induced Anchorage-Independent Survival , 2010, PloS one.

[22]  R. Wenger,et al.  Regulated oxygen sensing by protein hydroxylation in renal erythropoietin-producing cells. , 2010, American journal of physiology. Renal physiology.

[23]  T. Sørlie,et al.  Triple‐negative breast cancer: Present challenges and new perspectives , 2010, Molecular oncology.

[24]  G. Semenza.  Defining the role of hypoxia-inducible factor 1 in cancer biology and therapeutics , 2010, Oncogene.

[25]  A. Kung,et al.  Preferential binding of HIF-1 to transcriptionally active loci determines cell-type specific response to hypoxia , 2009, Genome Biology.

[26]  Karl J. Dykema,et al.  Met induces diverse mammary carcinomas in mice and is associated with human basal breast cancer , 2009, Proceedings of the National Academy of Sciences.

[27]  A. Krasnitz,et al.  Brk is coamplified with ErbB2 to promote proliferation in breast cancer , 2008, Proceedings of the National Academy of Sciences.

[28]  H. Iwase,et al.  Hypoxia-inducible factor 1α is closely linked to an aggressive phenotype in breast cancer , 2008, Breast Cancer Research and Treatment.

[29]  J. Ostrander,et al.  Breast tumor kinase (protein tyrosine kinase 6) regulates heregulin-induced activation of ERK5 and p38 MAP kinases in breast cancer cells. , 2007, Cancer research.

[30]  Zohar Yakhini,et al.  Polycomb-mediated methylation on Lys27 of histone H3 pre-marks genes for de novo methylation in cancer , 2007, Nature Genetics.

[31]  R. Johnson,et al.  Hypoxia-Inducible Factor-1α Is a Key Regulator of Metastasis in a Transgenic Model of Cancer Initiation and Progression , 2007 .

[32]  N. Ahn,et al.  Global Gene Expression Analysis of ERK5 and ERK1/2 Signaling Reveals a Role for HIF-1 in ERK5-mediated Responses* , 2006, Journal of Biological Chemistry.

[33]  J. Pouysségur,et al.  Hypoxia signalling in cancer and approaches to enforce tumour regression , 2006, Nature.

[34]  G. Camenisch,et al.  Integration of Oxygen Signaling at the Consensus HRE , 2005, Science's STKE.

[35]  P. Bonnier,et al.  Overexpression of hypoxia‐inducible factor HIF‐1α predicts early relapse in breast cancer: Retrospective study in a series of 745 patients , 2005, International journal of cancer.

[36]  Patrick H. Maxwell,et al.  Contrasting Properties of Hypoxia-Inducible Factor 1 (HIF-1) and HIF-2 in von Hippel-Lindau-Associated Renal Cell Carcinoma , 2005, Molecular and Cellular Biology.

[37]  H. Braselmann,et al.  Simultaneous over‐expression of the Her2/neu and PTK6 tyrosine kinases in archival invasive ductal breast carcinomas , 2005, The Journal of pathology.

[38]  K. Hunter,et al.  Modeling metastasis in vivo. , 2004, Carcinogenesis.

[39]  Brian Keith,et al.  Differential Roles of Hypoxia-Inducible Factor 1α (HIF-1α) and HIF-2α in Hypoxic Gene Regulation , 2003, Molecular and Cellular Biology.

[40]  P. V. van Diest,et al.  Levels of hypoxia‐inducible factor‐1α independently predict prognosis in patients with lymph node negative breast carcinoma , 2003, Cancer.

[41]  Yudong D. He,et al.  Gene expression profiling predicts clinical outcome of breast cancer , 2002, Nature.

[42]  V. Vasioukhin,et al.  BRK/Sik expression in the gastrointestinal tract and in colon tumors. , 1999, Clinical cancer research : an official journal of the American Association for Cancer Research.

[43]  B. Gusterson,et al.  Brk, a Breast Tumor-derived Non-receptor Protein-tyrosine Kinase, Sensitizes Mammary Epithelial Cells to Epidermal Growth Factor* , 1996, The Journal of Biological Chemistry.

[44]  G. Semenza,et al.  Hypoxia-inducible factor 1 is a basic-helix-loop-helix-PAS heterodimer regulated by cellular O2 tension. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[45]  V. Vasioukhin,et al.  A novel intracellular epithelial cell tyrosine kinase is expressed in the skin and gastrointestinal tract. , 1995, Oncogene.

[46]  B. Gusterson,et al.  Cloning and characterisation of cDNAs encoding a novel non-receptor tyrosine kinase, brk, expressed in human breast tumours. , 1994, Oncogene.

[47]  R. Johnson,et al.  Hypoxia-inducible factor-1alpha is a key regulator of metastasis in a transgenic model of cancer initiation and progression. , 2007, Cancer research.

[48]  A. Tyner,et al.  Brk, Srm, Frk, and Src42A form a distinct family of intracellular Src-like tyrosine kinases. , 2003, Oncology research.

[49]  Brian Keith,et al.  Differential roles of hypoxia-inducible factor 1alpha (HIF-1alpha) and HIF-2alpha in hypoxic gene regulation. , 2003, Molecular and cellular biology.

[50]  V. Vasioukhin,et al.  Sik Expression in the Gastrointestinal Tract and in Colon Tumors 1 , 1999 .