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Mutational Analysis of GNAQ and GNA11 to Aid Therapy Management of a Choroidal Melanoma Metastatic to the Contralateral Orbit

Mutational Analysis of GNAQ and GNA11 to Aid Therapy Management of a Choroidal Melanoma... Metastasis to the contralateral orbital cavity in uveal melanoma (UM) is extremely rare.1 Only 8 cases of such metastases have been reported, and none included mutational analysis. For the first time, to our knowledge, we report data from a mutational analysis of GNAQ and GNA11 supporting the diagnosis of a single metastatic tumor. Report of a Case A 53-year-old man had right-sided retrobulbar pain persisting for 4 weeks. The patient had undergone enucleation of his left eye 3 years previously for choroidal melanoma (27 × 12 × 10 mm). At this time, no other malignant neoplasms were observed by chest radiography, abdominal ultrasonography, and dermatological examinations. Findings on yearly magnetic resonance imaging of the orbital cavities with a T2-weighted turbo spin-echo sequence were unremarkable until 12 months prior to presentation (Figure 1A). Visual acuity of the remaining eye was 20/20. Findings on further ophthalmological examinations were normal. No periorbital hyperpigmentation or axial proptosis was present. An axial T2-weighted turbo spin-echo sequence magnetic resonance image demonstrated a right-sided, upper intraconal, hypointense lesion (20 × 11 × 12 mm) (Figure 1B). View LargeDownload Figure 1. Axial T2-weighted turbo spin-echo sequence magnetic resonance images of each orbit. A, Regular retrobulbar morphology 12 months prior to presentation. B, Upper intraconal, hypointense lesion (20 × 11 × 12 mm) detected at presentation. Tumor closely adjacent to the optic nerve and with infiltration of the rectus superior muscle was seen. A brown-colored tissue sample (10 × 10 mm) without pigmentation of the unaffected orbital tissue was obtained by performing a biopsy via the transcutaneous medial orbitotomy route. Histological analysis revealed weakly pigmented epithelioid cells and strongly pigmented spindle cells, leading to the diagnosis of melanoma (Figure 2A). An increased mitotic count (>5/10 high-power fields) and no connective tissue loops were observed. Immunohistochemical staining demonstrated tumor cells positive for S-100 protein and HMB-45, while Ki-67 (MIB-1) showed a growth fraction of 20% (data not shown). View LargeDownload Figure 2. Histopathological analysis and mutational analysis. Orbital specimen showed weakly pigmented epithelioid cells, strongly pigmented spindle cells, and scattered melanophages (hematoxylin-eosin, original magnification ×10) (A). Original sections of the enucleated eye showed comparable histopathological findings (hematoxylin-eosin, original magnification ×20 [B] and ×2 [C]). Sequence analysis of exon 4 showed identical mutations in GNA11 (c.547C > T) at codon 183 (p.R183C) in both metastasis (D) and primary choroidal melanoma (E). Evaluation of the patient's blood (F) and an adult control (G) revealed the wild type (WT) of GNA11 at codon 183. The DNA from tumor and peripheral blood cells was extracted using the QIAamp DNA Micro Kit (Qiagen) according to the manufacturer's instructions. For molecular karyotyping, array comparative genomic hybridization using the Agilent Technologies platform was performed according to the manufacturer's instructions. It revealed loss of chromosome 3, partial loss of 1p, 2q33-qter, 6q, and 16q, and gain of 8q and 16p. Sequencing of exons 4 and 5 of GNA11 revealed a mutation in exon 4 at c.547C > T (R183C) (Figure 2D). No mutations had been detected in GNA11 in exon 5 at amino acid position Q209 or in GNAQ at amino acid positions R183 and Q209, respectively. Evaluation of the patient's DNA from white blood cells revealed no mutations in GNA11 or GNAQ (Figure 2F). Mutation in BRAF (codon 600) was not detectable. Reevaluation of the primary UM revealed comparable histopathological findings with no evidence for extraocular spread, necrotic foci, or infiltration of the ciliary body (Figure 2B and C). Array comparative genomic hybridization analysis demonstrated loss of chromosome 3, partial loss of 1p, and gain of 16p, also seen in the orbital specimen. Identical molecular pathological results were found for GNA11, GNAQ, and BRAF (Figure 2E). Routine follow-up detected no abnormalities suspicious for widespread malignant disease in positron emission tomography and abdominal ultrasonography. In summary, detection of the same mutation in GNA11 at R183C and a missing mutation in GNAQ at Q209L in both tumor samples was interpreted to support the metastatic spread of the primary UM. The actual diagnosis prompted stereotactic, fractionated radiation therapy of the orbital tumor at 60 Gy (to convert to rad, multiply by 100) using a multileaf collimator. One year later, the patient presented with stable disease. Written consent for mutational and chromosomal analysis was given by the patient. Comment The clinical distinction between metastatic melanoma and second primary melanoma is hampered by both the rarity of orbital melanomas and the similarity of the histological features in the tumors.2 To our knowledge, only 8 cases of orbital metastases due to primary contralateral UM1 have been reported. Orbital melanomas are frequently associated with primary cutaneous melanomas.3 Only single cases of orbital melanoma without visible pigmentation or bilateral onset of UM have been described.4,5 The novelty of our case is in the association with mutational analysis of GNA11 and GNAQ. The recently described mutations of GNA11 and GNAQ affect amino acids Q209 in exon 5 and R183 in exon 4 in UM. While these 2 mutations have been found in both genes, they do not occur together in the same individual.6,7 Although these mutations are insufficient for full progression of melanoma, mitogen-activated protein kinase activation by oncogenes BRAF and NRAS appears to be an early event in neoplasms involving GNAQ mutations.8 The Q209 mutation is frequent in GNA11 and GNAQ in primary UM and metastases, while the R183 mutation is less frequent.7 In our case, both melanocytic tumors exhibited almost identical histopathology, immunohistochemistry, and proliferation rate. Based on our data and the presence of the identical GNA11 mutation in both tumors of the patient, the appearance of a tumor 3 years after successful eradication of the primary tumor from the orbital cavity was considered to favor a metastatic event and not a second primary tumor. This assumption prompted treatment with radiotherapy rather than radical surgical excision of the metastatic tumor, as would have been discussed as a curative treatment option for a second primary tumor. Hence, knowledge of mutations in GNA11 and GNAQ in UMs can help to distinguish between primary and metastatic melanoma lesions and to select the optimal therapeutic regimen. Performing GNA11 and GNAQ mutational analysis is recommended to direct therapy in future similar cases. Back to top Article Information Correspondence: Dr Böhm, Department of Ophthalmology, University Hospital of Muenster, Albert Schweitzer Campus D1, Bldg D15, 48149 Muenster, Germany (michael.boehm@ukmuenster.de). Published Online: April 9, 2013. doi:10.1001/jamaophthalmol.2013.1921 Author Contributions: Drs Böhm and Tsianakas share first authorship. Conflict of Interest Disclosures: None reported. Additional Contributions: The manuscript was edited by English Science Editing, Auckland, New Zealand. Solon Thanos, MD, PhD, provided comments that greatly improved the manuscript. References 1. Coupland SE, Sidiki S, Clark BJ, McClaren K, Kyle P, Lee WR. Metastatic choroidal melanoma to the contralateral orbit 40 years after enucleation. Arch Ophthalmol. 1996;114(6):751-7568639093PubMedGoogle ScholarCrossref 2. Liarikos S, Rapidis AD, Roumeliotis A, Angelopoulos AP. Secondary orbital melanomas: analysis of 15 cases. J Craniomaxillofac Surg. 2000;28(3):148-15210964550PubMedGoogle ScholarCrossref 3. Orcutt JC, Char DH. Melanoma metastatic to the orbit. Ophthalmology. 1988;95(8):1033-10373231441PubMedGoogle Scholar 4. Rice CD, Brown HH. Primary orbital melanoma associated with orbital melanocytosis. Arch Ophthalmol. 1990;108(8):1130-11342383203PubMedGoogle ScholarCrossref 5. Bhouri L, Lumbroso L, Levy C, et al. Bilateral uveal melanomas: five case reports [in French]. J Fr Ophtalmol. 2003;26(2):149-15312660588PubMedGoogle Scholar 6. Van Raamsdonk CD, Bezrookove V, Green G, et al. Frequent somatic mutations of GNAQ in uveal melanoma and blue naevi. Nature. 2009;457(7229):599-60219078957PubMedGoogle ScholarCrossref 7. Van Raamsdonk CD, Griewank KG, Crosby MB, et al. Mutations in GNA11 in uveal melanoma. N Engl J Med. 2010;363(23):2191-219921083380PubMedGoogle ScholarCrossref 8. Pollock PM, Harper UL, Hansen KS, et al. High frequency of BRAF mutations in nevi. Nat Genet. 2003;33(1):19-2012447372PubMedGoogle ScholarCrossref http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png JAMA Ophthalmology American Medical Association

Mutational Analysis of GNAQ and GNA11 to Aid Therapy Management of a Choroidal Melanoma Metastatic to the Contralateral Orbit

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Publisher
American Medical Association
Copyright
Copyright © 2013 American Medical Association. All Rights Reserved.
ISSN
2168-6165
eISSN
2168-6173
DOI
10.1001/jamaophthalmol.2013.1921
Publisher site
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Abstract

Metastasis to the contralateral orbital cavity in uveal melanoma (UM) is extremely rare.1 Only 8 cases of such metastases have been reported, and none included mutational analysis. For the first time, to our knowledge, we report data from a mutational analysis of GNAQ and GNA11 supporting the diagnosis of a single metastatic tumor. Report of a Case A 53-year-old man had right-sided retrobulbar pain persisting for 4 weeks. The patient had undergone enucleation of his left eye 3 years previously for choroidal melanoma (27 × 12 × 10 mm). At this time, no other malignant neoplasms were observed by chest radiography, abdominal ultrasonography, and dermatological examinations. Findings on yearly magnetic resonance imaging of the orbital cavities with a T2-weighted turbo spin-echo sequence were unremarkable until 12 months prior to presentation (Figure 1A). Visual acuity of the remaining eye was 20/20. Findings on further ophthalmological examinations were normal. No periorbital hyperpigmentation or axial proptosis was present. An axial T2-weighted turbo spin-echo sequence magnetic resonance image demonstrated a right-sided, upper intraconal, hypointense lesion (20 × 11 × 12 mm) (Figure 1B). View LargeDownload Figure 1. Axial T2-weighted turbo spin-echo sequence magnetic resonance images of each orbit. A, Regular retrobulbar morphology 12 months prior to presentation. B, Upper intraconal, hypointense lesion (20 × 11 × 12 mm) detected at presentation. Tumor closely adjacent to the optic nerve and with infiltration of the rectus superior muscle was seen. A brown-colored tissue sample (10 × 10 mm) without pigmentation of the unaffected orbital tissue was obtained by performing a biopsy via the transcutaneous medial orbitotomy route. Histological analysis revealed weakly pigmented epithelioid cells and strongly pigmented spindle cells, leading to the diagnosis of melanoma (Figure 2A). An increased mitotic count (>5/10 high-power fields) and no connective tissue loops were observed. Immunohistochemical staining demonstrated tumor cells positive for S-100 protein and HMB-45, while Ki-67 (MIB-1) showed a growth fraction of 20% (data not shown). View LargeDownload Figure 2. Histopathological analysis and mutational analysis. Orbital specimen showed weakly pigmented epithelioid cells, strongly pigmented spindle cells, and scattered melanophages (hematoxylin-eosin, original magnification ×10) (A). Original sections of the enucleated eye showed comparable histopathological findings (hematoxylin-eosin, original magnification ×20 [B] and ×2 [C]). Sequence analysis of exon 4 showed identical mutations in GNA11 (c.547C > T) at codon 183 (p.R183C) in both metastasis (D) and primary choroidal melanoma (E). Evaluation of the patient's blood (F) and an adult control (G) revealed the wild type (WT) of GNA11 at codon 183. The DNA from tumor and peripheral blood cells was extracted using the QIAamp DNA Micro Kit (Qiagen) according to the manufacturer's instructions. For molecular karyotyping, array comparative genomic hybridization using the Agilent Technologies platform was performed according to the manufacturer's instructions. It revealed loss of chromosome 3, partial loss of 1p, 2q33-qter, 6q, and 16q, and gain of 8q and 16p. Sequencing of exons 4 and 5 of GNA11 revealed a mutation in exon 4 at c.547C > T (R183C) (Figure 2D). No mutations had been detected in GNA11 in exon 5 at amino acid position Q209 or in GNAQ at amino acid positions R183 and Q209, respectively. Evaluation of the patient's DNA from white blood cells revealed no mutations in GNA11 or GNAQ (Figure 2F). Mutation in BRAF (codon 600) was not detectable. Reevaluation of the primary UM revealed comparable histopathological findings with no evidence for extraocular spread, necrotic foci, or infiltration of the ciliary body (Figure 2B and C). Array comparative genomic hybridization analysis demonstrated loss of chromosome 3, partial loss of 1p, and gain of 16p, also seen in the orbital specimen. Identical molecular pathological results were found for GNA11, GNAQ, and BRAF (Figure 2E). Routine follow-up detected no abnormalities suspicious for widespread malignant disease in positron emission tomography and abdominal ultrasonography. In summary, detection of the same mutation in GNA11 at R183C and a missing mutation in GNAQ at Q209L in both tumor samples was interpreted to support the metastatic spread of the primary UM. The actual diagnosis prompted stereotactic, fractionated radiation therapy of the orbital tumor at 60 Gy (to convert to rad, multiply by 100) using a multileaf collimator. One year later, the patient presented with stable disease. Written consent for mutational and chromosomal analysis was given by the patient. Comment The clinical distinction between metastatic melanoma and second primary melanoma is hampered by both the rarity of orbital melanomas and the similarity of the histological features in the tumors.2 To our knowledge, only 8 cases of orbital metastases due to primary contralateral UM1 have been reported. Orbital melanomas are frequently associated with primary cutaneous melanomas.3 Only single cases of orbital melanoma without visible pigmentation or bilateral onset of UM have been described.4,5 The novelty of our case is in the association with mutational analysis of GNA11 and GNAQ. The recently described mutations of GNA11 and GNAQ affect amino acids Q209 in exon 5 and R183 in exon 4 in UM. While these 2 mutations have been found in both genes, they do not occur together in the same individual.6,7 Although these mutations are insufficient for full progression of melanoma, mitogen-activated protein kinase activation by oncogenes BRAF and NRAS appears to be an early event in neoplasms involving GNAQ mutations.8 The Q209 mutation is frequent in GNA11 and GNAQ in primary UM and metastases, while the R183 mutation is less frequent.7 In our case, both melanocytic tumors exhibited almost identical histopathology, immunohistochemistry, and proliferation rate. Based on our data and the presence of the identical GNA11 mutation in both tumors of the patient, the appearance of a tumor 3 years after successful eradication of the primary tumor from the orbital cavity was considered to favor a metastatic event and not a second primary tumor. This assumption prompted treatment with radiotherapy rather than radical surgical excision of the metastatic tumor, as would have been discussed as a curative treatment option for a second primary tumor. Hence, knowledge of mutations in GNA11 and GNAQ in UMs can help to distinguish between primary and metastatic melanoma lesions and to select the optimal therapeutic regimen. Performing GNA11 and GNAQ mutational analysis is recommended to direct therapy in future similar cases. Back to top Article Information Correspondence: Dr Böhm, Department of Ophthalmology, University Hospital of Muenster, Albert Schweitzer Campus D1, Bldg D15, 48149 Muenster, Germany (michael.boehm@ukmuenster.de). Published Online: April 9, 2013. doi:10.1001/jamaophthalmol.2013.1921 Author Contributions: Drs Böhm and Tsianakas share first authorship. Conflict of Interest Disclosures: None reported. Additional Contributions: The manuscript was edited by English Science Editing, Auckland, New Zealand. Solon Thanos, MD, PhD, provided comments that greatly improved the manuscript. References 1. Coupland SE, Sidiki S, Clark BJ, McClaren K, Kyle P, Lee WR. Metastatic choroidal melanoma to the contralateral orbit 40 years after enucleation. Arch Ophthalmol. 1996;114(6):751-7568639093PubMedGoogle ScholarCrossref 2. Liarikos S, Rapidis AD, Roumeliotis A, Angelopoulos AP. Secondary orbital melanomas: analysis of 15 cases. J Craniomaxillofac Surg. 2000;28(3):148-15210964550PubMedGoogle ScholarCrossref 3. Orcutt JC, Char DH. Melanoma metastatic to the orbit. Ophthalmology. 1988;95(8):1033-10373231441PubMedGoogle Scholar 4. Rice CD, Brown HH. Primary orbital melanoma associated with orbital melanocytosis. Arch Ophthalmol. 1990;108(8):1130-11342383203PubMedGoogle ScholarCrossref 5. Bhouri L, Lumbroso L, Levy C, et al. Bilateral uveal melanomas: five case reports [in French]. J Fr Ophtalmol. 2003;26(2):149-15312660588PubMedGoogle Scholar 6. Van Raamsdonk CD, Bezrookove V, Green G, et al. Frequent somatic mutations of GNAQ in uveal melanoma and blue naevi. Nature. 2009;457(7229):599-60219078957PubMedGoogle ScholarCrossref 7. Van Raamsdonk CD, Griewank KG, Crosby MB, et al. Mutations in GNA11 in uveal melanoma. N Engl J Med. 2010;363(23):2191-219921083380PubMedGoogle ScholarCrossref 8. Pollock PM, Harper UL, Hansen KS, et al. High frequency of BRAF mutations in nevi. Nat Genet. 2003;33(1):19-2012447372PubMedGoogle ScholarCrossref

Journal

JAMA OphthalmologyAmerican Medical Association

Published: Jun 1, 2013

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