Immunohistochemical and molecular pathology of ocular uveal melanocytoma: Evidence for somatic GNAQ mutations

National Specialist Ophthalmic Pathology Service (NSOPS), Department of Histopathology, Royal Hallamshire Hospital, Sheffield, UK.
The British journal of ophthalmology (Impact Factor: 2.98). 05/2013; 97(7). DOI: 10.1136/bjophthalmol-2013-303291
Source: PubMed


Intraocular melanocytoma is a rare naevus variant that can be located at the optic disc or within the uvea, and belongs to the group of non-epithelial-associated melanocytic lesions. We wanted to gain an understanding of the role of GNAQ, GNA11 and BRAF V600E in the pathogenesis of uveal melanocytoma and in cases of transformation to uveal melanoma and also to perform a differential immunohistochemical study comparing melanocytoma with uveal melanoma.

Methods and results:
Two patients were identified with melanocytoma, one of which had transformed to melanoma. In the latter case, the melanocytoma exhibited an immunophenotype that featured nuclear p27 and no HMB45 staining, with very low Cyclin D1 expression compared with the melanoma that featured little nuclear but more cytoplasmic p27 positivity, much higher Cyclin D1 expression and HMB45 positivity. The melanocytomas were negative for CD68 allowing distinction from melanophages. Both melanocytomas and the melanoma harboured mutations in GNAQ, with no mutations of GNA11 or BRAF V600E.

GNAQ mutations are present in uveal melanocytomas and in a case of transformation to melanoma, implicating GNAQ-dependent mitogen activation signals, in the pathogenesis of uveal melanocytoma. This assists in explaining why a proportion of uveal melanocytoma can transform to uveal melanoma, known to harbour high-frequency GNAQ mutations at exon 5, codon 209.

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    • "Specific windows for GNAQ signaling in terms of location, cell type and developmental time were previously reported, such as that GNAQ mutations induced melanocytic proliferations spared epithelial structures [16]. Our study did not allow addressing whether the association between GNAQ/11 mutations and uveal melanomas involving optic disc would be a parallel to the previous observation of two choroidal melanocytomas harboring mutations in GNAQ with one of them transforming into uveal melanoma [25]. "
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    ABSTRACT: Purpose To examine whether GNAQ and GNA11 somatic mutations previously identified in uveal melanomas of Caucasians are associated with uveal melanomas in Chinese patients. Methods Uveal melanomas treated by primary enucleation in Chinese patients underwent a mutation analysis of GNAQ and GNA11 with sequencing of exon 5 and exon 4. Results The study included 50 patients with uveal melanoma and with a mean age of 47.6±13.0 years. During the follow-up of at least 3 years, 20 (40%) patients developed extraocular metastases. The frequencies of GNAQ and GNA11 somatic mutations in uveal melanoma were 18% (9/50) and 20% (10/50), respectively. The mutations occurred exclusively in codon 209 of exon 5. No mutations were detected in exon 4. Mutations affecting codon 209 in GNAQ were c.626A>C(Q209P) (78%) and c.626A>T(Q209L) (22%). Mutations affecting codon 209 in GNA11 were exclusively c.626A>T(Q209L) (100%). In none of the tumors, mutations of BRAF and NRAS were detected. GNAQ/11 mutations were marginally (P = 0.045) associated with optic disc involvement. In Kaplan-Meier analysis, metastasis-free survival was not significantly (P = 0.94) associated with GNAQ/11 mutations. Conclusions Mutations of GNAQ and GNA11 can be found in Chinese patients as in Caucasian patients with uveal melanoma, with a higher frequency reported for Caucasian patients.
    PLoS ONE 10/2014; 9(10):e109699. DOI:10.1371/journal.pone.0109699 · 3.23 Impact Factor
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    • "This method is also suitable for pathological tissues that require melanin depigmentation for morphological features that are often obscured, such as in the diagnosis of uveal melanoma. [6], [32] Cryosections are best air-dried overnight and post-fixed in 4% PFA prior to bleaching and staining in order to enhance tissue adherence and to preserve tissue architecture. The use of subbed slides, especially for cryosections, prevents the tissues from detaching from glass slides during vigorous treatment with highly alkaline solutions, such as H2O2. "
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    ABSTRACT: Purpose The removal of excessive melanin pigments that obscure ocular tissue morphology is important to address scientific questions and for differential diagnosis of ocular tumours based on histology. Thus, the goal of the present study was to establish an effective and fast melanin bleaching method for paraffin and frozen mouse and human ocular tissues. Methods Paraffin-embedded and frozen ocular specimens from mice and human donors were subjected to bleaching employing two methods. The first employed potassium permanganate (KMnO4) with oxalic acid, and the second 10% hydrogen peroxide (H2O2). To determine optimal bleaching conditions, depigmentation was carried out at various incubation times. The effect of diluents used for 10% H2O2 was assessed using phosphate-buffered saline (PBS), and deionized water. Three different slide types and two fixatives, which were ice-cold acetone with 80% methanol, and 4% paraformaldehyde (PFA) were used to determine the optimal conditions for better tissue adherence during bleaching. All tissues were stained in hematoxylin and eosin for histological evaluation. Results Optimal bleaching was achieved using warm 10% H2O2 diluted in PBS at 65°C for 120 minutes. Chromium-gelatin-coated slides prevented tissue detachment. Adherence of cryosections was also improved with post-fixation using 4% PFA and overnight air-drying at RT after cryosectioning. Tissue morphology was preserved under these conditions. Conversely, tissues bleached in KMnO4/oxalic acid demonstrated poor depigmentation with extensive tissue damage. Conclusions Warm dilute H2O2 at 65°C for 120 minutes rapidly and effectively bleached both cryo- and paraffin sections of murine and human ocular tissues.
    PLoS ONE 07/2014; 9(7):e102512. DOI:10.1371/journal.pone.0102512 · 3.23 Impact Factor
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    ABSTRACT: Purpose: The aim of this study was to discuss the clinical and translational content of the literature as well as advancement in our knowledge pertaining to retinoblastoma and uveal melanoma that were published from January to December 2013. Design: This study is a literature review. Methods: The search terms retinoblastoma and uveal melanoma were used in a MEDLINE literature search. Abstracts were studied, and the most relevant articles were selected for inclusion and further in-depth review. Results: In retinoblastoma, fewer eyes are lost because of the expanded use of ophthalmic artery chemosurgery and intravitreal melphalan, and the past year marks a deepening in our understanding of these modalities. Knowledge on the genetic underpinnings of uveal melanoma has broadened to include genes associated with a favorable prognosis. This is accompanied by promising results in the treatment of metastatic uveal melanoma. Conclusions: This past year, there were important advancements in our knowledge of retinoblastoma and uveal melanoma.
    01/2014; 3(4):241-256. DOI:10.1097/APO.0000000000000079
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