This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Parrella, P. Articles by Merbs, S. L. Search for Related Content PubMed PubMed Citation Articles by Parrella, P. Articles by Merbs, S. L.

Allelotype of Posterior Uveal Melanoma

Implications for a Bifurcated Tumor Progression Pathway¹

Dipartimento Ricerca BioMedica, Section for Molecular Medicine and Gene Therapy, University Campus Bio Medico, 00155 Rome, Italy [P. P.], and Department of Otolaryngology, Head & Neck Surgery, Division of Head and Neck Cancer Research [P. P., D. S.] and Wilmer Ophthalmological Institute, Department of Ophthalmology [S. L. M.], John Hopkins University School of Medicine, Baltimore, Maryland 21287

	ABSTRACT

Top ABSTRACT Introduction Materials and Methods Results Discussion REFERENCES

 
To further elucidate the somatic genetic alterations leading to acquired choroidal and ciliochoroidal melanoma, we screened every autosomal arm and the X chromosome of 50 primary posterior melanomas (31 choroidal tumors and 19 ciliochoroidal tumors). A minimum of two microsatellite markers were used to achieve at least 90% informativity (excluding X). Twenty-eight of 47 informative tumors (59%) showed allelic loss of all informative markers on chromosome 3, consistent with monosomy 3 (M3). Allelic imbalance of 8q was observed in 60% of tumors. A total of 28% of tumors displayed allelic loss of 6p. We then compared these genetic alterations with the status of chromosome 3 and found a relative absence of 6p alteration in tumors with M3 (P = 0.0005). Additionally, all observed 8q imbalance was associated with either M3 or alteration of 6p, suggesting that 8q alterations occur later in tumor progression. The mutual exclusivity of M3 and 6p alterations suggests a bifurcated tumor progression model. In this model, M3 or 6p loss identify distinct pathways, both followed by 8q loss in tumor progression.

	Introduction

Top ABSTRACT Introduction Materials and Methods Results Discussion REFERENCES

 
Although the incidence of uveal melanoma is about six to seven cases per one million people in North America, it is the most common primary intraocular malignancy in adults and represents 80% of all noncutaneous melanomas (1) . Posterior uveal melanoma, which includes ciliary body and choroidal melanoma, can result in significant visual morbidity from local tumor control and can pose a serious threat to life. The overall 5-year tumor-related mortality from metastases after enucleation ranges between 16% and 53%, depending on tumor size (2) .

Uveal melanomas are believed to arise from the neuroectodermally derived melanocytes present in the choroid, ciliary body, or iris. Iris melanomas comprise less then 10% of uveal melanomas and are typically associated with a good prognosis (3) . In contrast, ciliary body or choroidal melanomas are associated with poor prognosis, with at least 30% of affected patients developing metastases within 10 years of successful local management of the primary tumor (4) . The average life expectancy after identification of metastases is only 7 months (5) .

Advances in molecular genetics have made possible the identification of genetic changes and particular mutant genes in human tumors. To date, many chromosomal loci have been implicated in the development of human cancer, and over a dozen genes have now been isolated that are specifically mutated in primary tumors of various types (6) . Studies have also shown that tumor induction and progression are characterized by the accumulation of multiple genetic alterations in the long and complex evolution of cells from a normal to a malignant phenotype (7) . The frequent loss of a specific chromosome in cancer is thought to represent one step in the inactivation of a tumor suppressor gene residing on the lost chromosome. Tumorigenesis can also result from the amplification or mutation of an oncogene.

Although current histopathological and cytogenetic methods can provide some prognostic information, relatively little is known about the molecular genetic alterations and their temporal relationship leading to the development of uveal melanoma. Many cytogenetic investigations of cultured uveal melanoma cells have revealed that the majority of choroidal and ciliary body melanomas are characterized by nonrandom alterations in chromosomes 3, 6, and 8 (8) .

These observations have been confirmed by CGH³ (9, 10, 11) and, for chromosome 3, by FISH analysis (12) . M3 (either true or functional) and trisomy 8, partial duplication of 8q, or isochromosome 8q are the most frequent karyotypic abnormalities present in approximately 50% of cases (8 , 13) . Abnormalities in chromosomes 3 and 8 often coexist in the same tumor, whereas chromosome 6 alterations are detected in about 40% of cases (8, 9, 10, 11) .

To further investigate the molecular genetic aberrations that underlie the development of uveal melanoma and to uncover areas of chromosomal loss that may contain tumor suppressor genes, we performed a comprehensive allelotype of 50 posterior uveal melanomas. Every autosomal arm and the X chromosome were screened with highly informative microsatellite markers for genetic alterations. The highest frequency of LOH was found on chromosome 3 (64%), followed by chromosomal arm 8q (60%). Chromosome 6p allelic imbalance was found in 28% of tumors. LOH at all informative loci on chromosome 3 (likely M3) and 6p alterations were almost mutually exclusive, suggesting the presence of two distinct pathways in the development of uveal melanoma.

	Materials and Methods

Top ABSTRACT Introduction Materials and Methods Results Discussion REFERENCES

 
Specimens and DNA Extraction.
We evaluated 50 cases of posterior uveal melanoma treated by enucleation between 1993 and 1998 as paraffin-embedded tissue archived by the Eye Pathology Laboratory of the Wilmer Ophthalmological Institute. Twenty-six samples (52%) were from men, and 24 samples (48%) were from from women. Nineteen of 50 tumors involved the ciliary body (38%), and the remaining 31 tumors (62%) involved only the choroid. A wide range of tumor sizes was included in the study: (a) 6 small tumors (AH < 3 mm; BD < 16 mm); (b) 34 medium tumors (AH, 3–10 mm; BD 16 mm); and (c ) 10 large tumors (AH > 10 mm or BD > 16 mm; 14 ). The BD ranged from 5–18 mm (mean, 12 ± 3 mm), and the AH ranged from 2–19 mm (mean, 7 ± 4 mm). Tumors were classified by the modified Callendar classification (15) . Five tumors (10%) were classified as spindle cell tumors, 43 tumors (86%) were classified as mixed cell tumors, and 2 tumors (4%) were classified as epithelioid cell tumors.

DNA from the 50 choroidal and ciliochoroidal tumors was extracted from paraffin-embedded tissue as described previously (16) . Briefly, unstained paraffin sections of each block were microdissected into tumor and normal tissue. The normal ocular tissue was used as the source of normal control DNA for each patient. Normal tissue and tumor tissue were digested with SDS-proteinase K for 24 h at 48°C, and samples were extracted with phenol/chloroform. Pigments, which coisolated with the DNA after the extraction, tended to inhibit the PCR. Therefore, DNA samples were additionally purified using Geneclean II according to the manufacturer’s instructions (Bio 101, Vista, CA).

For sample UM58, two distinct regions of the tumor, one pigmented and one unpigmented, were microdissected and processed separately as tumor samples A (unpigmented) and B (pigmented). Both regions were classified as mixed cell tumors composed predominantly of spindle B cells with some epithelioid cells. For the allelotype and all statistical calculations, the results for UM58A were used (see "Results").

Allelotyping.
DNA from normal and tumor tissue was analyzed for LOH by amplification of microsatellite markers using PCR and the conditions described below (17) . Primer pairs designed to amplify microsatellite markers were obtained from Research Genetics (Huntsville, AL). As an initial screen, two markers per chromosomal arm were used: 1p, D1S214 and D1S219; 1q, D1S305 and D1S237; 2p, D2S162 and D2S358; 2q, D2S126 and D2S125; 3p, D3S1597 and D3S1284; 3q, D3S1292 and D3S1268; 4p, D4S394 and D4S404; 4q, D4S430 and D4S171; 5p, D5S417 and D5S455; 5q, D5S421 and D5S429; 6p, D6S470 and D6S265; 6q, D6S268 and D6S264; 7p, D7S481 and D7S507; 7q, D7S479 and D7S495; 8p, D8S262 and D8S261; 8q, D8S285 and D8S257; 9p, D9S171 and D9S104; 9q, D9S176 and D9S170; 10p, D10S249 and D10S191; 10q, D10S185 and D10S221; 11p, d11S907 and D11S929; 11q, INT2 and D11S873; 12p, D12S341 and D12S77; 12q, D12S329 and D12S86; 13q, D13S263 and D13S170; 14q, D14S283 and D14S67; 15q, D15S117 and D15S87; 16p, D16S404 and D16S412; 16q, D16S514 and D16S413; 17p, D17S796 and CHRNB1; 17q, D17S579 and D17S789; 18p, D18S52 and D18S453; 18q, D18S67 and D18S61; 19p, D19S424 and D19S177; 19q, D19S412 and D19S926; 20p, D20S95 and D20S104; 20q, D20S119 and D20S171; 21q, D21S11 and D21S262; 22q, D22S283 and IL2RB; Xp, DXS996 and DXS451; and Xq, DXS990 and DXS1192. Markers were chosen so that each member of a pair was widely separated on its chromosomal arm. The vast majority of chromosomal arms had at least 90% informativity for all tumors using only these two markers per chromosomal arm. An additional marker was used for the four chromosomal arms with less than 90% informativity, for those samples which were noninformative for both standard markers. The additional markers were D1S255 (1p), D1S158 (1q), D8S265 (8p), and D9S196 (9q).

Using T4 kinase (Life Technologies, Inc., Gaithersburg, MD), 500 ng of one primer from each pair were end-labeled with [32 P]ATP (20 mCi; Amersham, Piscataway, NJ) in a total volume of 20 µl (16) . Approximately 50 ng of DNA in 1x PCR buffer [16.6 mM ammonium sulfate, 67 mM Tris (pH 8.8), 6.7 mM magnesium chloride, 10 mM ß-mercaptoethanol, 1% DMSO, and 1.25 mM deoxynucleotide triphosphates) were amplified using 25 ng of end-labeled primer, 60 ng of each unlabeled primer, and 2 units of Taq DNA polymerase (New England Biolabs, Beverly, MA). PCR conditions were as follows: 35 cycles of 95°C for 1 min, 54°C for 1 min, and 72°C for 1 min, followed by a final 4-min extension at 72°C. About one-fourth of the PCR product was separated on an 8 M urea-formamide-polyacrylamide gel and exposed to film.

Cases were considered informative if two alleles were present in the normal sample. For informative cases, allelic loss was scored if the intensity of one allele was at least 30% reduced in tumor DNA as compared with the normal DNA, as determined visually by two independent observers (D. S. and S. L. M.).

Tumors with LOH of all informative markers on chromosome 3 and at least one informative marker per arm were considered to be M3. We did not perform cytogenetic studies or FISH analysis to distinguish cases of true M3 from cases of functional M3 (acquired isodisomy) (13) . Because both allelic loss and allelic amplification can be detected by microsatellite analysis, and because duplications of 6p and 8q have been observed in uveal melanoma by cytogenetic studies, we refer to apparent LOH on 6p and 8q only as allelic imbalance.

Statistical Analysis.
Statistical analyses were performed using SigmaStat program 1.02. The associations between the discrete variables were assessed using Fisher’s exact test. Mean values were compared using the two-tailed t test. Differences were considered statistically significant for P < 0.05.

	Results

Top ABSTRACT Introduction Materials and Methods Results Discussion REFERENCES

 
We screened 50 uveal melanomas (31 choroidal tumors and 19 ciliochoroidal tumors) for LOH at 86 highly informative microsatellite marker loci representing every chromosomal arm as shown in Table 1 . For each chromosomal locus, the total number of informative cases and the number of cases with LOH are shown. All chromosomal arms had at least 90% informativity for all tumors, and the majority had greater than 95% informativity.

View larger version (19K): [in this window] [in a new window]   Fig. 1. Total frequency of allelic loss for autosomes and the X chromosome in choroidal and ciliochoroidal melanoma. Allelotyping was performed using the microsatellite markers listed in Table 1 . , p; , q.

For further analysis, the tumors were divided into two groups: (a) those with retention of markers from both arms of chromosome 3 (R3); and (b) those with LOH of all markers from both arms of chromosome 3 (M3). Table 2 compares the frequencies of allelic imbalance for these two groups. Only chromosomal arms for which the frequencies were not identical are shown (Table 2) . Most striking was the relative absence of 6p alterations in tumors with M3. In fact, only 1 of 26 informative cases (4%) had both LOH of all chromosome 3 markers (M3) and allelic imbalance of markers on 6p, whereas 8 of 15 cases (53%) with retention of all chromosome 3 markers (R3) showed a 6p imbalance (P = 0.0005).

Allelic imbalance of chromosome 3 or chromosome 6p was found in 43 of 50 tumors (86%), and in 28 of 41 tumors (2 tumors were noninformative for 8q), 8q alteration was also present (68%). In those few tumors without alterations in 3, 6p, or 8q, no alternative common regions of LOH were noted (data not shown). Every tumor had at least one marker that displayed LOH. Representative results for two tumors, UM3 and UM14, are shown in Fig. 2A . Only 12 tumors showed microsatellite instability of at least one but no more than two microsatellite markers (data not shown).

We found no statistically significant difference in the frequency of chromosome 3 loss between those tumors with ciliary body involvement. M3 with associated alteration of 8q was found more often in tumors with ciliary body involvement (80%) than in tumors restricted to the choroid (66%), but the difference was not statistically significant.

A weak association was found between AH and allelic imbalance of chromosomal arm 8q. AH was 7.9 mm ± 4.2 in tumors with 8q alterations and 5.8 mm ± 2.9 in tumors without 8q alterations (P = 0.05).

Fig. 2B demonstrates the two histologically distinct regions of tumor UM58, which were microdissected separately. Although both the unpigmented region (UM58A) and the pigmented region (UM58B) had loss of the same allele for all informative markers on chromosome 3, UM58A and UM58B had relative loss of different alleles for a marker on 8q (Fig. 2C) , consistent with the hypothesis that 8q alteration follows the alteration of chromosome 3. Both regions had retention of 6p markers (Fig. 2C) . On chromosome 22, LOH of the same allele was also seen for both regions (data not shown). UM58A had a total of 10 additional chromosomal arms with LOH when compared to UM58B (data not shown). We chose to include the results from UM58A in the allelotype because it had the more complex pattern of LOH.

View larger version (39K): [in this window] [in a new window]   Fig. 2. LOH analysis of posterior uveal melanoma. A, autoradiographs depicting microsatellite analysis of representative tumors UM3 and UM14 are shown. Markers are labeled above each normal (N) and corresponding tumor (T) DNA lanes. Arrows indicate relative loss of alleles in tumor samples. From left to right, for marker D3S1597 (3p), UM3 demonstrates loss of the lower allele, whereas UM14 shows retention of both alleles; for marker D6S470 (6p), UM3 shows retention of both alleles, whereas UM14 shows relative loss of the bottom allele; and for marker D8S257 (8q), both tumors show relative loss of the top allele. B, H&E-stained paraffin section of tumor UM58, demonstrating the two distinct regions of the tumor, TA (unpigmented) and TB (pigmented). C, analysis of UM58 shows loss of the upper allele in both tumor regions (TA and TB) for marker D3S1597 (3p; ) and retention of alleles in both tumor regions for marker D6S265 (6p). For marker D8S285, TA shows relative LOH of the upper allele (), whereas TB shows relative LOH of the lower allele ().

	Discussion

Top ABSTRACT Introduction Materials and Methods Results Discussion REFERENCES

We found that 28 of 50 (59%) posterior uveal melanomas showed LOH at all informative loci on both arms of chromosome 3, consistent with the karyotypic detection of M3 in 50–60% of cases (8) . The exact mechanism by which M3 may lead to uveal melanoma is not known, but many candidate genes implicated in carcinogenesis (e.g., VHL, FHIT, and THRB) have been mapped to chromosome 3 (19, 20, 21) .

A total of 60% of tumors studied showed allelic imbalance of chromosomal arm 8q. Allelic imbalance, as detected by microsatellite analysis, can derive from the deletion of one of two chromosomal homologues (true LOH) or from the amplification of one of two chromosomal homologues (amplification). Because duplication of chromosomal arm 8q has been demonstrated in approximately 50% of uveal melanomas by many cytogenetic studies, our detected imbalance most likely represents duplication, but this remains to be confirmed by CGH or FISH analysis. The smallest duplicated region appears to be 8q21ter, spanning the c-myc oncogene locus (22) .

Loss of chromosomal arm 6q and gain of chromosomal arm 6p have also been reported in several karyotypic and CGH studies with different frequencies of occurrence (8, 9, 10, 11) . We found more 6p alterations (28%) than 6q alterations (18%). Interestingly, alterations in 6p occurred much more frequently in tumors with retention of chromosome 3 (P = 0.0005). As with the 8q alterations we detected, the 6p alterations most likely represent amplification when the results from karyotypic and CGH studies are considered. WAF1/CIP1, a cyclin-dependent kinase inhibitor, has been mapped to 6p and may be involved in the development of cutaneous melanoma (23, 24, 25, 26) . The genes encoding MHC class I molecules are also located on 6p, and expression of HLA molecules has been found to be altered in uveal melanoma by immunohistochemistry (27) .

Solid tumor induction and progression are characterized by the accumulation of multiple genetic alterations in the evolution of a normal cell to a malignant cell (7) . It has been suggested that loss of chromosome 3 is an early event in the neoplastic transformation of uveal melanomas (22) , and we found eight cases of M3 without chromosome 8 abnormalities. Our observation that M3 and alterations of 6p are essentially mutually exclusive in our population has led us to propose a bifurcated tumor progression model for uveal melanoma. As shown in Fig. 3 , the most common pathway would involve the loss of one copy of chromosome 3, with a divergent pathway characterized by alteration of 6p.

View larger version (8K): [in this window] [in a new window]   Fig. 3. A proposed genetic tumor progression model of posterior uveal melanoma. Most tumors undergo either loss of chromosome 3 (M3) or alteration of chromosomal arm 6p, with both groups subsequently undergoing alteration of chromosomal arm 8q. In our population, 28 tumors were M3 (65%), and 8 tumors had 6p alterations (20%). Approximately 60–70% of tumors go on to accumulate alteration in 8q.

An association between cytogenetic abnormalities on chromosomes 3, 6, and 8 in uveal melanoma and patient outcomes has been reported (29, 30, 31) . Most recently, White et al. (31) found a correlation between poor prognosis and the presence of M3 and amplification of 8q together and a correlation between better prognosis and the presence of chromosome 6 abnormalities. It is tempting to consider that the observations of White et al. may represent the two arms of our proposed progression model.

Although we did not find an association between M3 and ciliary body involvement (29) , we did find M3 with alterations of 8q more often in ciliochoroidal melanomas, consistent with a prior observation (31) . However, in our population, the difference was not statistically significant. Several reasons could explain this discrepancy. The most obvious explanation is the difference in technique used to detect genetic alterations. Microsatellite analysis, like CGH, is performed using DNA isolated directly from many tumor cells without cell culture and more accurately represents the tumor cell population in vivo. Differences in the frequency of genetic alterations have previously been observed when both CGH and cytogenetics were used to study a group of tumors (9) . As an example, higher frequencies of chromosome 6 alterations were found by CGH when compared to chromosome banding analysis of the same tumors (9) . Additionally, using CGH, Gordon et al. (10) did not find an association between abnormalities of chromosomes 3 and 8 and tumor location or cell type.

Another possibility to explain the discrepancy is a difference in our tumor population. We included a wide range of tumor sizes in our analysis to get a broad view of the genetic changes that lead to uveal melanoma. In contrast, most karyotypic studies for which tumor size has been provided have only included medium to very large tumors.

To date, only a few studies have analyzed uveal melanomas using microsatellite analysis, and these studies tested only chromosomes 3 and 9 (13 , 16 , 32) . We were able to detect LOH in 86% of tumors using a total of six microsatellite markers for chromosome 3 and 6p. Microsatellite analysis of tissue from fine needle aspirate biopsies or enucleations could be used to detect the genetic alterations, which correlate with prognosis (33) . This method might also be useful in identifying metastatic disease because microsatellite alterations have been found in the serum of patients with metastases from head and neck cancer, small cell lung carcinoma, and colon cancer (34, 35, 36) and, most recently, in the plasma of patients with cutaneous melanoma (37) .

	ACKNOWLEDGMENTS

 
We thank W. Richard Green (The Eye Pathology Laboratory, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD) for providing access to the banked paraffin specimens and Vicente Resto for comments on the manuscript.

	FOOTNOTES

 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ Supported by National Eye Institute Grant K08 EY00378-01 and the Niuta and Roy Titus Faculty Development Award (to S. L. M.) and by Specialized Program of Research Excellence in Lung Cancer Grant CA58-184 and National Institute for Dental and Craniofacial Research Grant R01 DE012588-01 (to D. S.). P. P. was supported by a fellowship from Progetto Sanita 96/97, Fondazione Cassa di Risparmio di Verona, Vicenza, Belluno, ed Ancona, Italy.

² To whom requests for reprints should be addressed, at Wilmer Ophthalmological Institute, Johns Hopkins University School of Medicine, 600 North Wolfe Street, Maumenee 127, Baltimore, MD 21287. Phone: (410) 955-1112; Fax: (410) 614-9987; E-mail: smerbs{at}jhmi.edu

³ The abbreviations used are: CGH, comparative genomic hybridization; FISH, fluorescence in situ hybridization; LOH, loss of heterozygosity; AH, apical height; BD, basal diameter; M3, monosomy 3; R3, disomy 3.

Received 4/ 8/99. Accepted 5/17/99.

	REFERENCES

Top ABSTRACT Introduction Materials and Methods Results Discussion REFERENCES

 

1. Scotto J., Fraumeni J. F., Lee J. A. H. Melanomas of the eye and other noncutaneous sites: epidemiologic aspects. J. Natl. Cancer Inst., 56: 489-461, 1976.
2. Diener-West M., Hawkins B. S., Markowitz J. A., Schachat A. P. A review of mortality from choroidal melanoma. II. A meta-analysis of 5-year mortality rates following enucleation, 1966 through 1988. Arch. Ophthalmol., 77: 245-250, 1993.
3. Brown D., Boniuk M., Font R. Diffuse malignant melanoma of iris with metastases. Surv. Ophthalmol., 34: 357-364, 1990.[Medline]
4. Jensen O. A. Malignant melanomas of the human uvea: 25-year follow-up of cases in Denmark, 1943–1952. Acta Ophthalmol., 60: 161-182, 1981.
5. Kath R., Hayungs J., Bornfeld N., Sauerwein W., Höffken K., Seeber S. Prognosis and treatment of disseminated uveal melanoma. Cancer (Phila.), 72: 2219-2223, 1993.[Medline]
6. Vogelstein B. Kinzler K. W. eds. . The Genetic Basis of Human Cancer, : McGraw-Hill New York 1998.
7. Fearon E. R., Vogelstein B. A genetic model for colorectal tumorigenesis. Cell, 61: 759-767, 1990.[Medline]
8. Becher R., Korn W. M., Prescher G. Use of fluorescent in situ hybridization and comparative genomic hybridization in the cytogenetic analysis of testicular germ cell tumors and uveal melanomas. Cancer Genet. Cytogenet., 93: 22-28, 1997.[Medline]
9. Speicher M. R., Prescher G., du Manoir S., Jauch A., Horsthemke B., Bornfeld N., Becher R., Cremer T. Chromosomal gains and losses in uveal melanomas detected by comparative genomic hybridization. Cancer Res., 54: 3817-3823, 1994.[Abstract/Free Full Text]
10. Gordon K. B., Thompson C. T., Char D. H., O’Brien J. M., Kroll S., Ghazvini S., Gray J. W. Comparative genomic hybridization in the detection of DNA copy number abnormalities in uveal melanoma. Cancer Res., 54: 4764-4768, 1994.[Abstract/Free Full Text]
11. Ghazvini S., Char D. H., Kroll S., Waldman F. M., Pinkel D. Comparative genomic hybridization analysis of archival formalin-fixed paraffin-embedded uveal melanomas. Cancer Genet. Cytogenet., 90: 95-101, 1996.[Medline]
12. McNamara M., Felix C., Davison E. V., Fenton M., Kennedy S. M. Assessment of chromosome 3 copy number in ocular melanoma using fluorescent in situ hybridization. Cancer Genet. Cytogenet., 98: 4-8, 1997.[Medline]
13. White V. A., McNeil K., Horsman D. Acquired homozygosity (isodisomy) of chromosome 3 in uveal melanoma. Cancer Genet. Cytogenet., 102: 40-45, 1998.[Medline]
14. The Collaborative Ocular Melanoma Study Group. Design and methods of a clinical trial for a rare condition: the collaborative ocular melanoma study. COMS Report No. 3. Controlled Clin. Trials, 14: 362-391, 1993.[Medline]
15. Mclean I. W., Foster W. D., Zimmerman L. E. Uveal melanoma: location, size, cell type and enucleation as risk factor in metastasis. Hum. Pathol., 13: 123-132, 1983.
16. Merbs S. M., Sidransky D. Analysis of p16 (CDKN2/MTS-1/INK4A) alterations in primary sporadic uveal melanoma. Invest. Ophthalmol. Visual Sci., 40: 779-83, 1999.[Abstract/Free Full Text]
17. van der Riet P., Nawroz H., Hruban R. H., Corio R., Tokino K., Koch W., Sidransky D. Frequent loss of chromosome 9p21–22 early in head and neck cancer progression. Cancer Res., 54: 1156-1158, 1994.[Abstract/Free Full Text]
18. Healy E., Belgaid C. E., Takata M., Vahlquist A., Rehman I., Rigby H., Rees J. L. Allelotypes of primary cutaneous melanoma and benign melanocytic nevi. Cancer Res., 56: 589-593, 1996.[Abstract/Free Full Text]
19. Latif F., Tory K., Gnarra J., Yao M., Duh F. M., Orcutt M. L., Stachouse T., Kuzmin I., Modi W., Geil L., Schmidt L., Zhou F., Li H., Wei M. H., Chen F., Glenn G., Choyke P., Walther M. M., Weng Y., Duan D. S. R., Dean M., Glavac D., Richards F. M., Crossey P. A., Ferguson-Smith M. A., Le Paslier D., Chumakov I., Cohen D., Chinault A. C., Maher E. R., Linehan W. M., Zbar B., Lerman M. I. Identification of the von Hippel-Lindau disease tumor suppressor gene. Science (Washington DC), 260: 1317-1320, 1993.[Abstract/Free Full Text]
20. Ohta M., Inoue H., Cotticelli M. G., Kastury K., Baffa R., Palazzo J., Siprashvili Z., Mori M., McCue P., Druck T., Croce C. M., Huebner K. The FHIT gene, spanning the chromosome 3p14.2 fragile site and renal carcinoma-associated t(3;8) breakpoint, is abnormal in digestive tract cancers. Cell, 84: 587-597, 1996.[Medline]
21. Weinberg C., Thompson C. C., Ong E. S., Lebo R., Gruol D. J., Evans R. M. The c-erb-A gene encodes a thyroid hormone receptor. Nature (Lond.), 324: 641-646, 1986.[Medline]
22. Prescher G., Bornfeld N., Becher R. Nonrandom abnormalities in primary uveal melanoma. J. Natl. Cancer Inst., 82: 1765-1769, 1990.[Abstract/Free Full Text]
23. Harper J. W., Adami G. R., Wei N., Keyomarsi K., Elledge S. J. The p21 CdK-interacting protein cip1 is a potent inhibitor of G₁ cyclin-dependent kinase. Cell, 75: 805-816, 1993.[Medline]
24. El-Deiry W. S., Tokino T., Velculescu V. E., Levy D. B., Parson R., Trent J. M., Lin D., Mercer W. E., Kinzler K. W., Vogelstein B. WAF1 a potential mediator of p53 tumor suppression. Cell, 75: 817-825, 1993.[Medline]
25. Gorospe M., Cirielli C., Wang X., Seth P., Capogrossi M. C., Holbrook N. J. p21(Waf1/Cip1) protects against p53-mediated apoptosis of human melanoma cells. Oncogene, 14: 929-935, 1997.[Medline]
26. Karjalainen J. M., Eskelinen M. J., Kellokoski J. K., Reinikainen M., Alhava E. M., Kosma V. M. p21(WAF1/CIP1) expression in stage I cutaneous malignant melanoma: its relationship with p53, cell proliferation and survival. Br. J. Cancer, 79: 895-902, 1999.[Medline]
27. Blom D. J. R., Schurmans L. R. H. M., de Waard-Siebinga I., De Wolff-Rouendaal D., Keunen J. E. E., Jager M. J. HLA expression in a primary uveal melanoma, its cell line and four of its metastases. Br. J. Ophthalmol., 81: 989-993, 1997.[Abstract/Free Full Text]
28. Horsman D. E., Sroka H., Rootman J., White V. A. Monosomy 3 and isochromosome 8q in uveal melanoma. Cancer Genet. Cytogenet., 45: 249-253, 1990.[Medline]
29. Prescher C., Bornfeld N., Hirche H., Horsthemke B., Jöckel K-H., Becher R. Prognostic implications of monosomy 3 in uveal melanoma. Lancet, 347: 1222-1225, 1996.[Medline]
30. Sisley K., Rennie I. G., Parsons A., Jacques R., Hammond D. W., Bell S. M., Potter A. M., Rees R. C. Abnormalities of chromosomes 3 and 8 in posterior uveal melanoma correlate with prognosis. Genes Chromosomes Cancer, 19: 22-28, 1997.[Medline]
31. White V. A., Chambers J. D., Courtright P. D., Chang W. Y., Horsman D. E. Correlation of cytogenetic abnormalities with the outcome of patients with uveal melanoma. Cancer (Phila.), 83: 354-359, 1998.[Medline]
32. Ohta M., Berd D., Shimizu M., Nagai H., Cotticelli M-G., Mastrangelo M., Shields J. A., Sheilds C. L., Croce C. M., Huebner K. Deletion mapping of chromosome region 9p21–p22 surrounding the CDNK2 locus in melanoma. Int. J. Cancer, 65: 762-767, 1996.[Medline]
33. Sisley K., Nichols C., Parsons M. A., Farr R., Rees R. C., Rennie I. G. Clinical applications of chromosome analysis, from fine needle aspiration biopsies, of posterior uveal melanomas. Eye (Lond.), 12: 203-207, 1998.
34. Nawroz H., Koch W., Anker P., Stroun M., Sidransky D. Microsatellite alterations in serum DNA of head and neck cancer patients. Nat. Med., 2: 1035-1037, 1996.[Medline]
35. Chen X. Q., Stroun M., Manenat J. L., Nicod L. P., Kurt A. M., Lyautey J., Lederrey C., Anker P. Microsatellite alterations in plasma DNA of small cell lung cancer patients. Nat. Med., 2: 1033-1034, 1996.[Medline]
36. Hibi K., Robinson C. R., Booker S., Wu L., Hamilton S. R., Sidransky D., Jen J. Molecular detection of genetic alterations in the serum of colorectal cancer patients. Cancer Res., 58: 1405-1407, 1998.[Abstract/Free Full Text]
37. Fujiwara Y., Chi D. D. J., Wang H., Keleman P., Morton D. L., Turner R., Hoon D. S. B. Plasma DNA microsatellites as tumor-specific markers and indicators of tumor progression in melanoma patients. Cancer Res., 59: 1567-1571, 1999.[Abstract/Free Full Text]

This article has been cited by other articles:

				J. P. Ehlers, L. Worley, M. D. Onken, and J. W. Harbour Integrative Genomic Analysis of Aneuploidy in Uveal Melanoma Clin. Cancer Res., January 1, 2008; 14(1): 115 - 122. [Abstract] [Full Text] [PDF]

				M. D. Onken, L. A. Worley, E. Person, D. H. Char, A. M. Bowcock, and J. W. Harbour Loss of Heterozygosity of Chromosome 3 Detected with Single Nucleotide Polymorphisms Is Superior to Monosomy 3 for Predicting Metastasis in Uveal Melanoma Clin. Cancer Res., May 15, 2007; 13(10): 2923 - 2927. [Abstract] [Full Text] [PDF]

				T. Sandinha, M. Farquharson, I. McKay, and F. Roberts Correlation of Heterogeneity for Chromosome 3 Copy Number with Cell Type in Choroidal Melanoma of Mixed-Cell Type Invest. Ophthalmol. Vis. Sci., December 1, 2006; 47(12): 5177 - 5180. [Abstract] [Full Text] [PDF]

				N. Goldenberg-Cohen, Y. Cohen, E. Rosenbaum, Z. Herscovici, I. Chowers, D. Weinberger, J. Pe'er, and D. Sidransky T1799A BRAF Mutations in Conjunctival Melanocytic Lesions Invest. Ophthalmol. Vis. Sci., September 1, 2005; 46(9): 3027 - 3030. [Abstract] [Full Text] [PDF]

				J. P. Ehlers, L. Worley, M. D. Onken, and J. W. Harbour DDEF1 Is Located in an Amplified Region of Chromosome 8q and Is Overexpressed in Uveal Melanoma Clin. Cancer Res., May 15, 2005; 11(10): 3609 - 3613. [Abstract] [Full Text] [PDF]

				M T Sandinha, M A Farquharson, and F Roberts Identification of monosomy 3 in choroidal melanoma by chromosome in situ hybridisation Br. J. Ophthalmol., December 1, 2004; 88(12): 1527 - 1532. [Abstract] [Full Text] [PDF]

				P. Parrella, M. L. Poeta, A. P. Gallo, M. Prencipe, M. Scintu, A. Apicella, R. Rossiello, G. Liguoro, D. Seripa, C. Gravina, et al. Nonrandom Distribution of Aberrant Promoter Methylation of Cancer-Related Genes in Sporadic Breast Tumors Clin. Cancer Res., August 15, 2004; 10(16): 5349 - 5354. [Abstract] [Full Text] [PDF]

				P. Parrella, V. M. Fazio, A. P. Gallo, D. Sidransky, and S. L. Merbs Fine Mapping of Chromosome 3 in Uveal Melanoma: Identification of a Minimal Region of Deletion on Chromosomal Arm 3p25.1-p25.2 Cancer Res., December 1, 2003; 63(23): 8507 - 8510. [Abstract] [Full Text] [PDF]

				Y. Cohen, N. Goldenberg-Cohen, P. Parrella, I. Chowers, S. L. Merbs, J. Pe'er, and D. Sidransky Lack of BRAF Mutation in Primary Uveal Melanoma Invest. Ophthalmol. Vis. Sci., July 1, 2003; 44(7): 2876 - 2878. [Abstract] [Full Text] [PDF]

				P. Parrella, Y. Xiao, M. Fliss, M. Sanchez-Cespedes, P. Mazzarelli, M. Rinaldi, T. Nicol, E. Gabrielson, C. Cuomo, D. Cohen, et al. Detection of Mitochondrial DNA Mutations in Primary Breast Cancer and Fine-Needle Aspirates Cancer Res., October 1, 2001; 61(20): 7623 - 7626. [Abstract] [Full Text] [PDF]

				P. Parrella, O. L. Caballero, D. Sidransky, and S. L. Merbs Detection of c-myc Amplification in Uveal Melanoma by Fluorescent In Situ Hybridization Invest. Ophthalmol. Vis. Sci., July 1, 2001; 42(8): 1679 - 1684. [Abstract] [Full Text] [PDF]

				F. Tschentscher, G. Prescher, D. E. Horsman, V. A. White, H. Rieder, G. Anastassiou, H. Schilling, N. Bornfeld, K. U. Bartz-Schmidt, B. Horsthemke, et al. Partial Deletions of the Long and Short Arm of Chromosome 3 Point to Two Tumor Suppressor Genes in Uveal Melanoma Cancer Res., April 1, 2001; 61(8): 3439 - 3442. [Abstract] [Full Text]

				Y. Aalto, L. Eriksson, S. Seregard, O. Larsson, and S. Knuutila Concomitant Loss of Chromosome 3 and Whole Arm Losses and Gains of Chromosome 1, 6, or 8 in Metastasizing Primary Uveal Melanoma Invest. Ophthalmol. Vis. Sci., February 1, 2001; 42(2): 313 - 317. [Abstract] [Full Text]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Parrella, P. Articles by Merbs, S. L. Search for Related Content PubMed PubMed Citation Articles by Parrella, P. Articles by Merbs, S. L.