Chromosome Research

, Volume 25, Issue 2, pp 129–143 | Cite as

Genomic profiling of canine mast cell tumors identifies DNA copy number aberrations associated with KIT mutations and high histological grade

  • Hiroyuki Mochizuki
  • Rachael Thomas
  • Scott Moroff
  • Matthew Breen
Original Article


Mast cell tumor (MCT) is the most common skin malignancy of domestic dogs and presents with a widely variable clinical behavior. Although activating KIT mutations are present in approximately 20% of canine MCTs, molecular etiology is largely unknown for the majority of this cancer. Characterization of genomic alterations in canine MCTs may identify genomic regions and/or genes responsible for their development and progression, facilitating the discovery of new therapeutic targets and improved clinical management of this heterogeneous cancer. We performed genome-wide DNA copy number analysis of 109 primary MCTs derived from three popular canine breeds (the Boxer, Labrador Retriever, and Pug) as well as nontarget breeds using oligonucleotide array comparative genomic hybridization (oaCGH). We demonstrated a stepwise accumulation of numerical DNA copy number aberrations (CNAs) as tumor grade increases. DNA sequencing analysis revealed that KIT mutations were found less frequently in the Pug tumors and were strongly associated with high histological grade. Tumors with KIT mutations showed genome-wide aberrant copy number profiles, with frequent CNAs involving genes in the p53 and RB pathways, whereas CNAs were very limited in tumors with wild-type KIT. We evaluated the presence of four CNAs to predict aggressive tumor phenotypes. This approach predicted aggressive tumors with a sensitivity of 78–94% and specificity of 88–93%, when using oaCGH and droplet digital PCR platforms. Further investigation of genome regions identified in this study may lead to the development of a molecular tool for classification and prognosis, as well as identification of therapeutic target molecules.


Cancer Comparative genomic hybridization Digital PCR Dog Mastocytosis Pug 



Canis familiaris (also used as a prefix to chromosome numbers)


Confidence interval


Copy number aberration


Droplet digital PCR


Formalin fixed paraffin embedded


Genomic Identification of Significant Targets in Cancer


Internal tandem duplication


Mast cell tumor


Mutant KIT


Oligonucleotide array comparative genomic hybridization


Odds ratio


Receiver operating characteristic


Wild-type KIT



This study was funded in part by a grant from Antech Diagnostics (awarded to MB) and by the NCSU Cancer Genomics fund (MB). HM was supported in part by a Morris Animal Foundation Fellowship (awarded to HM, study ID: D14CA-401) and a Postdoctoral Fellowship for Research Abroad, provided by the Japan Society for the Promotion of Science (HM).

Compliance with ethical standards

Ethical standards

The experiment complies with the current laws of the country, the USA, in which they were performed.

Studies of human or animal subjects

This article does not contain any studies with human or animal subjects performed by the any of the authors.

Supplementary material

10577_2016_9543_MOESM1_ESM.xls (236 kb)
ESM 1 (XLS 235 kb)


  1. Angstadt AY, Motsinger-Reif A, Thomas R et al (2011) Characterization of canine osteosarcoma by array comparative genomic hybridization and RT-qPCR: signatures of genomic imbalance in canine osteosarcoma parallel the human counterpart. Genes Chromosomes Cancer 50:859–874CrossRefPubMedGoogle Scholar
  2. Arendt ML, Melin M, Tonomura N et al (2015) Genome-wide association study of golden retrievers identifies germ-line risk factors predisposing to mast cell tumours. PLoS Genet 11:e1005647CrossRefPubMedPubMedCentralGoogle Scholar
  3. Beroukhim R, Getz G, Nghiemphu L et al (2007) Assessing the significance of chromosomal aberrations in cancer: methodology and application to glioma. Proc Natl Acad Sci U S A 104:20007–20012CrossRefPubMedPubMedCentralGoogle Scholar
  4. Bhat S, Curach N, Mostyn T, Bains GS, Griffiths KR, Emslie KR (2010) Comparison of methods for accurate quantification of DNA mass concentration with traceability to the international system of units. Anal Chem 82:7185–7192CrossRefPubMedGoogle Scholar
  5. Bhat S, Mclaughlin JL, Emslie KR (2011) Effect of sustained elevated temperature prior to amplification on template copy number estimation using digital polymerase chain reaction. Analyst 136:724–732CrossRefPubMedGoogle Scholar
  6. Bonkobara M (2015) Dysregulation of tyrosine kinases and use of imatinib in small animal practice. Vet J 205:180–188CrossRefPubMedGoogle Scholar
  7. Downing S, Chien MB, Kass PH, Moore PE, London CA (2002) Prevalence and importance of internal tandem duplications in exons 11 and 12 of c-kit in mast cell tumors of dogs. Am J Vet Res 63:1718–1723CrossRefPubMedGoogle Scholar
  8. Elvers I, Turner-Maier J, Swofford R et al (2015) Exome sequencing of lymphomas from three dog breeds reveals somatic mutation patterns reflecting genetic background. Genome Res 25(11):1634–1645CrossRefPubMedPubMedCentralGoogle Scholar
  9. Finnie JW, Bostock DE (1979) Skin neoplasia in dogs. Aust Vet J 55:602–604CrossRefPubMedGoogle Scholar
  10. Frohling S, Dohner H (2008) Chromosomal abnormalities in cancer. N Engl J Med 359:722–734CrossRefPubMedGoogle Scholar
  11. Giantin M, Granato A, Baratto C et al (2014) Global gene expression analysis of canine cutaneous mast cell tumor: could molecular profiling be useful for subtype classification and prognostication? PLoS One 9:e95481CrossRefPubMedPubMedCentralGoogle Scholar
  12. Haenisch B, Nöthen MM, Molderings GJ (2012) Systemic mast cell activation disease: the role of molecular genetic alterations in pathogenesis, heritability and diagnostics. Immunology 137:197–205CrossRefPubMedPubMedCentralGoogle Scholar
  13. Hahn KA, Legendre AM, Shaw NG et al (2010) Evaluation of 12- and 24-month survival rates after treatment with masitinib in dogs with nonresectable mast cell tumors. Am J Vet Res 71:1354–1361CrossRefPubMedGoogle Scholar
  14. Hanahan D, Weinberg RA (2011) Hallmarks of cancer: the next generation. Cell 144:646–674CrossRefPubMedGoogle Scholar
  15. Hedan B, Thomas R, Motsinger-Reif A et al (2011) Molecular cytogenetic characterization of canine histiocytic sarcoma: a spontaneous model for human histiocytic cancer identifies deletion of tumor suppressor genes and highlights influence of genetic background on tumor behavior. BMC Cancer 11(201)Google Scholar
  16. Kiupel M, Webster JD, Bailey KL et al (2011) Proposal of a 2-tier histologic grading system for canine cutaneous mast cell tumors to more accurately predict biological behavior. Vet Pathol 48:147–155CrossRefPubMedGoogle Scholar
  17. Letard S, Yang Y, Hanssens K et al (2008) Gain-of-function mutations in the extracellular domain of KIT are common in canine mast cell tumors. Mol Cancer Res 6:1137–1145CrossRefPubMedGoogle Scholar
  18. Lin TY, Thomas R, Tsai PC, Breen M, London CA (2009) Generation and characterization of novel canine malignant mast cell line CL1. Vet Immunol Immunopathol 127:114–124CrossRefPubMedGoogle Scholar
  19. London CA, Malpas PB, Wood-Follis SL et al (2009) Multi-center, placebo-controlled, double-blind, randomized study of oral toceranib phosphate (SU11654), a receptor tyrosine kinase inhibitor, for the treatment of dogs with recurrent (either local or distant) mast cell tumor following surgical excision. Clin Cancer Res 15:3856–3865CrossRefPubMedGoogle Scholar
  20. Mcniel EA, Prink AL, O'brien TD (2006) Evaluation of risk and clinical outcome of mast cell tumours in pug dogs. Vet Comp Oncol 4:2–8CrossRefPubMedGoogle Scholar
  21. Mitelman F, Johansson B, Mertens F (2007) The impact of translocations and gene fusions on cancer causation. Nat Rev Cancer 7:233–245CrossRefPubMedGoogle Scholar
  22. Mochizuki, H, Motsinger-Reif, A, Bettini, C, Moroff, S, Breen, M (2016a) Association of breed and histopathological grade in canine mast cell tumours. Veterinary and Comparative Oncology. Google Scholar
  23. Mochizuki H, Shapiro SG, Breen M (2016b) Detection of copy number imbalance in canine urothelial carcinoma with droplet digital polymerase chain reaction. Vet Pathol 53:764–772CrossRefPubMedGoogle Scholar
  24. Northrup NC, Harmon BG, Gieger TL et al (2005) Variation among pathologists in histologic grading of canine cutaneous mast cell tumors. J Vet Diagn Investig 17:245–248CrossRefGoogle Scholar
  25. Patnaik AK, Ehler WJ, Macewen EG (1984) Canine cutaneous mast cell tumor: morphologic grading and survival time in 83 dogs. Vet Pathol 21:469–474CrossRefPubMedGoogle Scholar
  26. Poorman K, Borst L, Moroff S et al (2015) Comparative cytogenetic characterization of primary canine melanocytic lesions using array CGH and fluorescence in situ hybridization. Chromosom Res 23:171–186CrossRefGoogle Scholar
  27. Roode, SC, Rotroff, D, Avery, AC, et al. (2015) Genome-wide assessment of recurrent genomic imbalances in canine leukemia identifies evolutionarily conserved regions for subtype differentiation. Chromosome Res.Google Scholar
  28. Roskoski R Jr (2005) Signaling by kit protein-tyrosine kinase—the stem cell factor receptor. Biochem Biophys Res Commun 337:1–13CrossRefPubMedGoogle Scholar
  29. Rothwell TL, Howlett CR, Middleton DJ, Griffiths DA, Duff BC (1987) Skin neoplasms of dogs in Sydney. Aust Vet J 64:161–164CrossRefPubMedGoogle Scholar
  30. Sabattini S, Scarpa F, Berlato D, Bettini G (2015) Histologic grading of canine mast cell tumor: is 2 better than 3? Vet Pathol 52:70–73CrossRefPubMedGoogle Scholar
  31. Shapiro SG, Raghunath S, Williams C et al (2015) Canine urothelial carcinoma: genomically aberrant and comparatively relevant. Chromosom Res 23:311–331CrossRefGoogle Scholar
  32. Shoop S, Marlow S, Church D et al (2015) Prevalence and risk factors for mast cell tumours in dogs in England. Canine Genetics and Epidemiology 2:1–10CrossRefPubMedPubMedCentralGoogle Scholar
  33. Stefanello D, Buracco P, Sabattini S et al (2015) Comparison of 2-and 3-category histologic grading systems for predicting the presence of metastasis at the time of initial evaluation in dogs with cutaneous mast cell tumors: 386 cases (2009–2014). J Am Vet Med Assoc 246:765–769CrossRefPubMedGoogle Scholar
  34. Takeuchi Y, Fujino Y, Watanabe M et al (2013) Validation of the prognostic value of histopathological grading or c-kit mutation in canine cutaneous mast cell tumours: a retrospective cohort study. Vet J 196:492–498CrossRefPubMedGoogle Scholar
  35. Thomas R, Borst L, Rotroff D et al (2014) Genomic profiling reveals extensive heterogeneity in somatic DNA copy number aberrations of canine hemangiosarcoma. Chromosom Res 22:305–319CrossRefGoogle Scholar
  36. Thomas R, Duke SE, Wang HJ et al (2009) ‘Putting our heads together’: insights into genomic conservation between human and canine intracranial tumors. J Neuro-Oncol 94:333–349CrossRefGoogle Scholar
  37. Thomas R, Seiser EL, Motsinger-Reif A et al (2011) Refining tumor-associated aneuploidy through ‘genomic recoding’ of recurrent DNA copy number aberrations in 150 canine non-Hodgkin lymphomas. Leuk Lymphoma 52:1321–1335CrossRefPubMedPubMedCentralGoogle Scholar
  38. Villamil JA, Henry CJ, Bryan JN et al (2011) Identification of the most common cutaneous neoplasms in dogs and evaluation of breed and age distributions for selected neoplasms. J Am Vet Med Assoc 239:960–965CrossRefPubMedGoogle Scholar
  39. Warland J, Dobson J (2013) Breed predispositions in canine mast cell tumour: a single centre experience in the United Kingdom. Vet J 197:496–498CrossRefPubMedGoogle Scholar
  40. Webster JD, Yuzbasiyan-Gurkan V, Kaneene JB, Miller R, Resau JH, Kiupel M (2006) The role of c-kit in tumorigenesis: evaluation in canine cutaneous mast cell tumors. Neoplasia 8:104–111CrossRefPubMedPubMedCentralGoogle Scholar
  41. White CR, Hohenhaus AE, Kelsey J, Procter-Gray E (2011) Cutaneous mcts: associations with spay/neuter status, breed, body size, and phylogenetic cluster. J Am Anim Hosp Assoc 47:210–216CrossRefPubMedGoogle Scholar
  42. Zemke D, Yamini B, Yuzbasiyan-Gurkan V (2002) Mutations in the juxtamembrane domain of c-kit are associated with higher grade mast cell tumors in dogs. Vet Pathol 39:529–535CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2017

Authors and Affiliations

  • Hiroyuki Mochizuki
    • 1
    • 2
  • Rachael Thomas
    • 1
    • 2
  • Scott Moroff
    • 3
  • Matthew Breen
    • 1
    • 2
    • 4
    • 5
  1. 1.Department of Molecular Biomedical Sciences, College of Veterinary MedicineNorth Carolina State UniversityRaleighUSA
  2. 2.Comparative Medicine InstituteNorth Carolina State UniversityRaleighUSA
  3. 3.Antech Diagnostics Inc.New Hyde ParkUSA
  4. 4.Center for Human Health and the EnvironmentNorth Carolina State UniversityRaleighUSA
  5. 5.Lineberger Comprehensive Cancer CenterUniversity of North CarolinaChapel HillUSA

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