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Proteomics of Uveal Melanomas Suggests HSP-27 as a Possible Surrogate Marker of Chromosome 3 Loss

From the ¹Department of Pathology, School of Cancer Studies, University of Liverpool, Liverpool, United Kingdom; the ³Department of Ophthalmology, Aarhus University Hospital, Aarhus, Denmark; the ⁴Department of Medical Biochemistry, Aarhus University, Aarhus, Denmark; and the ⁶Ocular Oncology Service, St. Paul's Eye Clinic, Royal Liverpool University Hospital, Liverpool, United Kingdom.

Corresponding author: Sarah E. Coupland, Department of Pathology, University of Liverpool, Liverpool, UK; s.e.coupland{at}liverpool.ac.uk.

Purpose. To compare the proteomic profiles of primary uveal melanomas, with and without loss of chromosome 3.

Methods. Frozen specimens from three uveal melanomas with disomy 3 and from four tumors with monosomy 3, according to fluorescence in situ hybridization (FISH) analysis, were subjected to high-resolution, two-dimensional (2-D) gel electrophoresis. The protein expression profiles of the two uveal melanoma cytogenetic groups were compared: Proteins that differed significantly were excised and analyzed by tandem mass spectrometry. Differentially expressed proteins were further analyzed with Western blot analysis. An independent cohort of 41 formalin-fixed, paraffin-embedded (FFPE) uveal melanomas, whose chromosome 3 status had been determined by multiplex ligation-dependent probe amplification (MLPA), was examined for the appropriate antigens by immunohistochemistry.

Results. Four protein spots were 1.5-fold (Student's t-test, P < 0.05) differentially expressed in the two uveal melanoma types: two spots were overexpressed in the disomy 3 group compared with the monosomy 3 group, whereas two spots were underexpressed. Identification of the four spots yielded nine proteins. Western blot analysis confirmed the results for heat shock protein (HSP)-27, vimentin, and pyruvate dehydrogenase β (PDHB), with a statistical significance for the first two proteins. HSP-27 was significantly downregulated, whereas vimentin was upregulated in the monosomy 3 tumors (Student's t-test, P = 0.003 and P = 0.005, respectively). Immunohistochemistry confirmed low-to-negative HSP-27 protein expression in monosomy 3 uveal melanomas (Student's t-test; P = 0.011).

Conclusions. Low-to-negative HSP-27 protein expression in uveal melanoma correlates strongly with monosomy 3. Further validation is necessary to determine whether immunohistochemical assessment of HSP-27 expression correlates with metastatic mortality.