TTK/hMPS1 Is an Attractive Therapeutic Target for Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) represents a subgroup of breast cancers (BC) associated with the most aggressive clinical behavior. No targeted therapy is currently available for the treatment of patients with TNBC. In order to discover potential therapeutic targets, we searched for protein kinases that are overexpressed in human TNBC biopsies and whose silencing in TNBC cell lines causes cell death. A cohort including human BC biopsies obtained at Institut Curie as well as normal tissues has been analyzed at a gene-expression level. The data revealed that the human protein kinase monopolar spindle 1 (hMPS1), also known as TTK and involved in mitotic checkpoint, is specifically overexpressed in TNBC, compared to the other BC subgroups and healthy tissues. We confirmed by immunohistochemistry and reverse phase protein array that TNBC expressed higher levels of TTK protein compared to the other BC subgroups. We then determined the biological effects of TTK depletion by RNA interference, through analyses of tumorigenic capacity and cell viability in different human TNBC cell lines. We found that RNAi-mediated depletion of TTK in various TNBC cell lines severely compromised their viability and their ability to form colonies in an anchorage-independent manner. Moreover, we observed that TTK silencing led to an increase in H2AX phosphorylation, activation of caspases 3/7, sub-G1 cell population accumulation and high annexin V staining, as well as to a decrease in G1 phase cell population and an increased aneuploidy. Altogether, these data indicate that TTK depletion in TNBC cells induces apoptosis. These results point out TTK as a protein kinase overexpressed in TNBC that may represent an attractive therapeutic target specifically for this poor prognosis associated subgroup of breast cancer.

[1]  O. Kallioniemi,et al.  Analysis of Kinase Gene Expression Patterns across 5681 Human Tissue Samples Reveals Functional Genomic Taxonomy of the Kinome , 2010, PloS one.

[2]  E. Rogakou,et al.  Megabase Chromatin Domains Involved in DNA Double-Strand Breaks in Vivo , 1999, The Journal of cell biology.

[3]  R. Myers,et al.  Evolving gene/transcript definitions significantly alter the interpretation of GeneChip data , 2005, Nucleic acids research.

[4]  V. Cryns,et al.  Minireview: Basal-like breast cancer: from molecular profiles to targeted therapies. , 2011, Molecular endocrinology.

[5]  L. Saal,et al.  Recurrent gross mutations of the PTEN tumor suppressor gene in breast cancers with deficient DSB repair , 2008, Nature Genetics.

[6]  E. Salmon,et al.  The spindle-assembly checkpoint in space and time , 2007, Nature Reviews Molecular Cell Biology.

[7]  Yusuke Nakamura,et al.  Detection of novel cancer‐testis antigen‐specific T‐cell responses in TIL, regional lymph nodes, and PBL in patients with esophageal squamous cell carcinoma , 2008, Cancer Science.

[8]  Wen-Lin Kuo,et al.  A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. , 2006, Cancer cell.

[9]  A. Vincent-Salomon,et al.  A New Model of Patient Tumor-Derived Breast Cancer Xenografts for Preclinical Assays , 2007, Clinical Cancer Research.

[10]  K. Hess,et al.  Response to neoadjuvant therapy and long-term survival in patients with triple-negative breast cancer. , 2008, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[11]  P. Pandolfi,et al.  Essential Role for Nuclear PTEN in Maintaining Chromosomal Integrity , 2007, Cell.

[12]  Chao Zhang,et al.  Chemical Genetic Inhibition of Mps1 in Stable Human Cell Lines Reveals Novel Aspects of Mps1 Function in Mitosis , 2010, PloS one.

[13]  Alan Mackay,et al.  Functional viability profiles of breast cancer. , 2011, Cancer discovery.

[14]  Christian A. Rees,et al.  Molecular portraits of human breast tumours , 2000, Nature.

[15]  Zhi Hu,et al.  An integrative genomic and proteomic analysis of PIK3CA, PTEN, and AKT mutations in breast cancer. , 2008, Cancer research.

[16]  R. Medema,et al.  Elevating the frequency of chromosome mis-segregation as a strategy to kill tumor cells , 2009, Proceedings of the National Academy of Sciences.

[17]  Ju-Hyung Woo,et al.  Increased Expression of Mitotic Checkpoint Genes in Breast Cancer Cells with Chromosomal Instability , 2006, Clinical Cancer Research.

[18]  John W M Martens,et al.  Subtypes of breast cancer show preferential site of relapse. , 2008, Cancer research.

[19]  Chao Zhang,et al.  Chemical Genetics Reveals a Role for Mps1 Kinase in Kinetochore Attachment during Mitosis , 2005, Current Biology.

[20]  Anthony Rhodes,et al.  American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer. , 2006, Archives of pathology & laboratory medicine.

[21]  M. Kruhøffer,et al.  Identification of gene expression patterns in superficial and invasive human bladder cancer. , 2001, Cancer research.

[22]  C. Perou,et al.  The Triple Negative Paradox: Primary Tumor Chemosensitivity of Breast Cancer Subtypes , 2007, Clinical Cancer Research.

[23]  R. Medema,et al.  Chromosome Segregation Errors as a Cause of DNA Damage and Structural Chromosome Aberrations , 2011, Science.

[24]  M. Moynahan,et al.  BRCA Gene Structure and Function in Tumor Suppression: A Repair-Centric Perspective , 2010, Cancer journal.

[25]  Duane A. Compton,et al.  Proliferation of aneuploid human cells is limited by a p53-dependent mechanism , 2010, The Journal of cell biology.

[26]  A. Vincent-Salomon,et al.  High sensitivity and specificity of immunohistochemistry for the detection of hormone receptors in breast carcinoma: comparison with biochemical determination in a prospective study of 793 cases , 2000, Histopathology.

[27]  Eric L. Weiss,et al.  The Saccharomyces cerevisiae spindle pole body duplication gene MPS1 is part of a mitotic checkpoint , 1996, The Journal of cell biology.

[28]  Chen-Yang Shen,et al.  TTK/hMps1 Participates in the Regulation of DNA Damage Checkpoint Response by Phosphorylating CHK2 on Threonine 68* , 2005, Journal of Biological Chemistry.

[29]  Ka-Wo Lee,et al.  Arecoline arrests cells at prometaphase by deregulating mitotic spindle assembly and spindle assembly checkpoint: implication for carcinogenesis. , 2010, Oral oncology.

[30]  V. Adamo,et al.  The role of Aurora-A inhibitors in cancer therapy. , 2007, Annals of oncology : official journal of the European Society for Medical Oncology.

[31]  Antonella Isacchi,et al.  Targeting the mitotic checkpoint for cancer therapy with NMS-P715, an inhibitor of MPS1 kinase. , 2010, Cancer research.

[32]  Geert J. P. L. Kops,et al.  On the road to cancer: aneuploidy and the mitotic checkpoint , 2005, Nature Reviews Cancer.

[33]  Ian O Ellis,et al.  Basal-like breast cancer: a critical review. , 2008, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[34]  R. Tibshirani,et al.  Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[35]  Taebo Sim,et al.  Small Molecule Kinase Inhibitors Provide Insight into Mps1 Cell Cycle Function , 2010, Nature chemical biology.

[36]  F. Bertucci,et al.  Sixteen-kinase gene expression identifies luminal breast cancers with poor prognosis. , 2008, Cancer research.

[37]  S. Wacholder,et al.  Gene Expression Signature of Cigarette Smoking and Its Role in Lung Adenocarcinoma Development and Survival , 2008, PloS one.

[38]  Harry Bartelink,et al.  Gene expression profiling and histopathological characterization of triple-negative/basal-like breast carcinomas , 2007, Breast Cancer Research.

[39]  A. Nobel,et al.  Concordance among Gene-Expression – Based Predictors for Breast Cancer , 2011 .

[40]  Geert J P L Kops,et al.  Lethality to human cancer cells through massive chromosome loss by inhibition of the mitotic checkpoint. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[41]  S. Shieh,et al.  TTK/hMps1 Mediates the p53-Dependent Postmitotic Checkpoint by Phosphorylating p53 at Thr18 , 2009, Molecular and Cellular Biology.

[42]  M. Alaoui-Jamali,et al.  Molecular insights on basal-like breast cancer , 2012, Breast Cancer Research and Treatment.

[43]  A. Ashworth,et al.  Hallmarks of 'BRCAness' in sporadic cancers , 2004, Nature Reviews Cancer.

[44]  A. Vincent-Salomon,et al.  KIT is highly expressed in adenoid cystic carcinoma of the breast, a basal-like carcinoma associated with a favorable outcome , 2005, Modern Pathology.

[45]  John J Spinelli,et al.  HER-2/neu in Breast Cancer: Interobserver Variability and Performance of Immunohistochemistry with 4 Antibodies Compared with Fluorescent In Situ Hybridization , 2001, Modern Pathology.

[46]  A. Nobel,et al.  The molecular portraits of breast tumors are conserved across microarray platforms , 2006, BMC Genomics.

[47]  A. Vincent-Salomon,et al.  High frequency of TP53 mutation in BRCA1 and sporadic basal-like carcinomas but not in BRCA1 luminal breast tumors. , 2009, Cancer research.

[48]  A. Vincent-Salomon,et al.  Identification of typical medullary breast carcinoma as a genomic sub-group of basal-like carcinomas, a heterogeneous new molecular entity , 2007, Breast Cancer Research.

[49]  M. Bittner,et al.  A cell proliferation and chromosomal instability signature in anaplastic thyroid carcinoma. , 2007, Cancer research.

[50]  Guanjun Chen,et al.  Cellular Abundance of Mps1 and the Role of Its Carboxyl Terminal Tail in Substrate Recruitment* , 2010, The Journal of Biological Chemistry.

[51]  Guillem Rigaill,et al.  Pruned dynamic programming for optimal multiple change-point detection , 2010 .

[52]  T. Jacks,et al.  Characterization of the p53-Dependent Postmitotic Checkpoint following Spindle Disruption , 1998, Molecular and Cellular Biology.

[53]  Aaron Rogers,et al.  Characterization of the Cellular and Antitumor Effects of MPI-0479605, a Small-Molecule Inhibitor of the Mitotic Kinase Mps1 , 2011, Molecular Cancer Therapeutics.

[54]  Rafael A. Irizarry,et al.  A Model-Based Background Adjustment for Oligonucleotide Expression Arrays , 2004 .

[55]  Emmanuel Barillot,et al.  Frequent PTEN genomic alterations and activated phosphatidylinositol 3-kinase pathway in basal-like breast cancer cells , 2008, Breast Cancer Research.

[56]  J. Berlin,et al.  Aurora Kinase Inhibitors - Rising Stars in Cancer Therapeutics? , 2010, Molecular Cancer Therapeutics.

[57]  Jun Qin,et al.  MPS1-dependent mitotic BLM phosphorylation is important for chromosome stability. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[58]  A. Vincent-Salomon,et al.  Ploidy and large-scale genomic instability consistently identify basal-like breast carcinomas with BRCA1/2 inactivation. , 2012, Cancer research.

[59]  Z. Szallasi,et al.  A signature of chromosomal instability inferred from gene expression profiles predicts clinical outcome in multiple human cancers , 2006, Nature Genetics.

[60]  Y. Pommier,et al.  Initiation of DNA Fragmentation during Apoptosis Induces Phosphorylation of H2AX Histone at Serine 139* , 2000, The Journal of Biological Chemistry.

[61]  O. Fadare,et al.  Clinical and pathologic aspects of basal-like breast cancers , 2008, Nature Clinical Practice Oncology.

[62]  R. Tibshirani,et al.  Repeated observation of breast tumor subtypes in independent gene expression data sets , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[63]  E. Gabrielson,et al.  High levels of the Mps1 checkpoint protein are protective of aneuploidy in breast cancer cells , 2011, Proceedings of the National Academy of Sciences.

[64]  Emmanuel Barillot,et al.  Polo-like kinase 1: a potential therapeutic option in combination with conventional chemotherapy for the management of patients with triple-negative breast cancer. , 2013, Cancer research.

[65]  Charles M. Perou,et al.  Triple-Negative Breast Cancer: Risk Factors to Potential Targets , 2008, Clinical Cancer Research.

[66]  Debra L Winkeljohn.  Triple-negative breast cancer. , 2008, Clinical journal of oncology nursing.

[67]  J. Bartek,et al.  Distinct spatiotemporal dynamics of mammalian checkpoint regulators induced by DNA damage , 2003, Nature Cell Biology.

[68]  D. Cleveland,et al.  A chemical tool box defines mitotic and interphase roles for Mps1 kinase , 2010, The Journal of cell biology.

[69]  A. Gown,et al.  Immunohistochemical and Clinical Characterization of the Basal-Like Subtype of Invasive Breast Carcinoma , 2004, Clinical Cancer Research.

[70]  T. Nielsen,et al.  Deconstructing the molecular portrait of basal-like breast cancer. , 2006, Trends in molecular medicine.