Tyrosinase is a copper-containing type I membrane glycoprotein essential for melanin synthesis. Tyrosinase catalyzes the hydroxylation of tyrosine to dihydroxyphenylalanine, which is considered the rate-limiting step in melanin production.1 In humans, tyrosinase is expressed in epidermal melanocytes as well as the pigmented epithelia of the retina, iris, and ciliary body of the eye.2,3 This expression appears to be tightly controlled both spatially and temporally through a variety of cis-acting and trans-acting elements.4 In brief, tyrosinase expression is upregulated in developing melanocytes and downregulated in mature and quiescent melanocytes.1,4 In contrast, in neoplastic tissues, tyrosinase expression appears constitutively increased in all malignant melanocytic tumors.4,5 Because of the tight temporal and spatial regulation in normal tissues (and the high expression in tumor tissues), tyrosinase has proven to be a useful target for immunotherapeutic approaches in humans with melanocytic tumors.6
Much of the information regarding tyrosinase expression has been derived from human and rodent cell lines and histologic samples.1–5 Although a commercially available xenogenic tyrosinase vaccine for the treatment of dogs with melanoma has had encouraging results, minimal published information exists on the tissue-specific expression of canine or equine tyrosinase.7,8 Gene and protein expression studies9,10 have identified detectable expression of tyrosinase in canine and equine tissues, respectively. A genetic study11 has identified mutations associated with development of melanocytic tumors in gray horses; these mutations are thought to result in upregulation of genes such as tyrosinase. However, no large-scale or comparative tyrosinase gene expression has been described in either species. Further information on the expression of canine tyrosinase may be useful to understand the role of targeted immunotherapy in dogs with melanocytic tumors. Furthermore, data on the expression of tyrosinase in equine melanocytic tumors may support the use of this immunologic modality in a different species.
The MHC I gene complex is a component of the antigen-processing machinery that is commonly dysregulated in tumor tissues.12 Downregulation of this gene may result in the development of resistance to targeted immunotherapies.12,13 Correlations between MHC I expression and tissue type may thus prove useful in further understanding the response to treatment in patients treated with tyrosinase-targeted immunotherapy. The primary objective of the study reported here was to determine the relative expression of tyrosinase mRNA in a series of canine and equine melanocytic tumors. The secondary objective was to determine the relative expression of antigen presentation gene MHC I mRNA in this series of tissue samples.
Formalin-fixed and paraffin embedded
Major histocompatibility complex
SurePrep RNA Isolation kit, catalog No. BP-2816–50, Fisher Lifescience, Waltham, Mass.
TaqMan gene expression assays, Applied Biosystems, Foster City, Calif.
TaqMan Custom Assay Design Tool, Applied Biosystems, Foster City, Calif.
TaqMan RNA-to-Ct 1-Step Kit, Applied Biosystems, Foster City, Calif.
BioRad MyiQ Real-Time PCR Detection System, BioRad, Berkeley, Calif.
iQ 5 Real-time PCR Detection Optical System Software, version 2.0, BioRad, Berkeley, Calif.
STATA, version 11.0, Data Analysis and Statistical Software, College Station, Tex.
Oncept, Canine Melanoma Vaccine, Merial Ltd, Duluth, Ga.
Wang N, Hebert D. Tyrosinase maturation through the mammalian secretory pathway: bringing color to life. Pigment Cell Res 2006; 19:3–18.
Schraermeyer U, Kopitz J, Peters S, et al. Tyrosinase biosynthesis in adult mammalian retinal pigment epithelial cells. Exp Eye Res 2006; 83:315–321.
Hayasaka S, Nakazawa M, Ishiguro S, et al. Presence of tyrosinase activity in human ciliary body. Jpn J Ophthalmol 1986; 30:32–35.
Vourc'h-Jourdain M, Volteau C, Nguyen J, et al. Melanoma gene expression and clinical course. Arch Dermatol Res 2009; 301:673–679.
Ferguson A, Nichols L, Zarling A, et al. Strategies and challenges in eliciting immunity to melanoma. Immunol Rev 2008; 222:28–42.
Bergman P, McKnight J, Novosad A, et al. Long-term survival of dogs with advanced malignant melanoma after DNA vaccination with xenogenic human tyrosinase: a phase I trial. Clin Cancer Res 2003; 9:1284–1290.
Ramos-Vara JA, Beissenherz ME, Miller MA, et al. Retrospective study of 338 canine oral melanomas with clinical, histologic, and immunohistochemical review of 129 cases. Vet Pathol 2000; 37:597–608.
Stell AJ, Dobson JM, Scase TJ, et al. Evaluation of variants of melanoma-associated antigen genes and mRNA transcripts in melanomas of dogs. Am J Vet Res 2009; 70:1512–1520.
Seltenhammer M, Heere-Ress E, Brandt S, et al. Comparative histopathology of grey-horse-melanoma and human malignant melanoma. Pigment Cell Res 2004; 17:674–681.
Rosengren G, Pielberg G, Golovko A, et al. A cis-acting regulatory mutation causes premature hair graying and susceptibility to melanoma in the horse. Nat Genet 2008; 40:1004–1009.
Seliger B. Molecular mechanisms of MHC class I abnormalities and APM components in human tumors. Cancer Immunol Immunother 2008; 57:1719–1726.
Garrido F, Algarra I, Garcia-Lora AM. The escape of cancer from T lymphocytes: immunoselection of MHC class I loss variants harboring structural-irreversible “hard” lesions. Cancer Immunol Immunother 2010; 59:1601–1606.
Proulx DR, Ruslander DM, Dodge RK, et al. A retrospective analysis of 140 dogs with oral melanoma treated with external beam radiation. Vet Radiol Ultrasound 2003; 44:352–359.
Bonin S, Hlubek F, Benhattar J, et al. Multicentre validation study of nucleic acids extraction from FFPE tissues. Virchows Arch 2010; 457:309–317.
Farragher SM, Tanney A, Kennedy RD, et al. RNA expression analysis from formalin fixed paraffin embedded tissues. Histochem Cell Biol 2008; 130:435–445.
Jager D, Taverna C, Zippelius A, et al. Identification of tumor antigens as potential target antigens for immunotherapy by serological expression cloning. Cancer Immunol Immunother 2004; 53:144–147.
Jaanson N, Moll K, Kulla A, et al. Identification of the immunodominant regions of the melanoma antigen tyrosinase by anti-tyrosinase monoclonal antibodies. Melanoma Res 2003; 13:473–482.
Chen YT, Stockert E, Tsang S, et al. Immunophenotyping of melanomas for tyrosinase: implications for vaccine development. Proc Natl Acad Sci USA 1995; 92:8125–8129.
Willers J, Lucchese A, Mittelman A, et al. Definition of anti-tyrosinase MAb T311 linear determinant by proteome-based similarity analysis. Exp Dermatol 2005; 14:543–550.
Cangul IT, van Garderen E, van der Poel HJ, et al. Tyrosinase gene expression in clear cell sarcoma indicates a melanocytic origin: insight from the first reported canine case. APMIS 1999; 107:982–988.
Catchpole B, Gould SM, Kellett-Gregory LM, et al. Development of a multiple-marker polymerase chain reaction assay for detection of metastatic melanoma in lymph node aspirates of dogs. Am J Vet Res 2003; 64:544–549.
Oetting WS. The tyrosinase gene and oculocutaneous albinism type 1 (OCA1): a model for understanding the molecular biology of melanin formation. Pigment Cell Res 2000; 13:320–325.
Gradilone A, Cigna E, Agliano A, et al. Tyrosinase expression as a molecular marker for investigating the presence of circulating tumor cells in melanoma patients. Curr Cancer Drug Targets 2010; 10:529–538.
Quaglino P, Osella-Abate S, Cappello N, et al. Prognostic relevance of baseline and sequential peripheral blood tyrosinase expression in 200 consecutive advanced metastatic melanoma patients. Melanoma Res 2007; 17:75–82.
Gkalpakiotis S, Arenberger P, Kremen J, et al. Quantitative detection of melanoma-associated antigens by multimarker realtime RT-PCR for molecular staging: results of a 5 years study. Exp Dermatol 2010; 19:994–999.
Arenberger P, Arenbergerova M, Vohradnikova O, et al. Early detection of melanoma progression by quantitative real-time RT-PCR analysis for multiple melanoma markers. Keio J Med 2008; 57:57–64.