Reaction on "Ocular ultrasound versus MRI in the detection of extrascleral extension in a patient with choroidal melanoma".

BACKGROUND: In the recently published article entitled "Ocular ultrasound versus MRI in the detection of extrascleral extension in a patient with choroidal melanoma" Jacobsen et al. describe a case in which a hyper-intense extra-ocular lesion on MRI was erroneously diagnosed as an extrascleral extension of the tumor. Based upon this the authors conclude "the superiority of ocular ultrasound in the diagnostic management of extra scleral extension in choroidal melanoma". In our view, there are numerous flaws in the investigation that cast doubt on this message. MAIN: First of all, this is quite a bold statement when only one patient has been evaluated. Secondly, the manuscript only presents a post-contrast T1-weighted image, whereas multiple MRI-sequences need to be included to determine if a hyperintense region is an extrascleral invasion. Moreover, no modern MRI-techniques such Dynamic Contrast Enhanced (DCE) or Diffusion Weighted Imaging (DWI) have been included in the evaluation of this patient, making it hard to use this single case to compare the efficacy of MRI and Ultrasound. The presented data do, however, give clear clues that the hyperintense lesion is likely to be inflammatory. CONCLUSION: Although the study falls short in providing a comprehensive comparison between current MRI techniques and ultrasound, it does show that the evaluation of ocular MR-images should be made in a multi-disciplinary setting involving both ophthalmologist and radiologists, since the field of ocular MRI is continuously progressing.

CT and MR imaging of orbital inflammation.

PURPOSE: Orbital inflammation can be idiopathic or in the context of a specific disease and it can involve different anatomical orbital structures. On imaging, inflammatory disease is frequently mistaken for infection and malignant tumors, and its underlying cause is often not determined. Through this article we aim to improve orbital inflammation diagnosis and underlying inflammatory diseases recognition. METHODS: The imaging protocols and characteristics of orbital inflammation were reviewed. RESULTS: A decision tree for the evaluation of these patients is provided. First, a combination of clinical and radiological clues is used to recognize inflammation, in particular to differentiate it both from orbital infection and tumor. Subsequently, different radiological patterns are recognized, often allowing the differentiation of the several orbital inflammatory diseases. CONCLUSION: The use of adequate imaging protocols and subsequent evaluation allow the recognition of an orbital lesion as inflammatory and the diagnosis of the underlying inflammatory disease. All in all, a proper treatment can be established, and at times, a biopsy can be avoided.

High-resolution MRI of uveal melanoma using a microcoil phased array at 7 T.

High-field MRI is a promising technique for the characterisation of ocular tumours, both in vivo and after enucleation. For in vivo imaging at 7 T, a dedicated three-element microcoil array was constructed as a high-sensitivity receive-only device. Using a dedicated blink/fixation protocol, high-resolution in vivo images could be acquired within 3 min in volunteers and patients with no requirement for post-acquisition image registration. Quantitative measures of axial length, aqueous depth and lens thickness in a healthy volunteer were found to agree well with standard ocular biometric techniques. In a patient with uveal melanoma, in vivo MRI gave excellent tumour/aqueous body contrast. Ex vivo imaging of the enucleated eye showed significant heterogeneity within the tumour.

Three-dimensional MRI-based treatment planning approach for non-invasive ocular proton therapy.

PURPOSE: To develop a high-resolution three-dimensional (3D) magnetic resonance imaging (MRI)-based treatment planning approach for uveal melanomas (UM) in proton therapy. MATERIALS/METHODS: For eight patients with UM, a segmentation of the gross tumor volume (GTV) and organs-at-risk (OARs) was performed on T1- and T2-weighted 7 Tesla MRI image data to reconstruct the patient MR-eye. An extended contour was defined with a 2.5-mm isotropic margin derived from the GTV. A broad beam algorithm, which we have called πDose, was implemented to calculate relative proton absorbed doses to the ipsilateral OARs. Clinically favorable gazing angles of the treated eye were assessed by calculating a global weighted-sum objective function, which set penalties for OARs and extreme gazing angles. An optimizer, which we have named OPT'im-Eye-Tool, was developed to tune the parameters of the functions for sparing critical-OARs. RESULTS: In total, 441 gazing angles were simulated for every patient. Target coverage including margins was achieved in all the cases (V95%  > 95%). Over the whole gazing angles solutions space, maximum dose (Dmax ) to the optic nerve and the macula, and mean doses (Dmean ) to the lens, the ciliary body and the sclera were calculated. A forward optimization was applied by OPT'im-Eye-Tool in three different prioritizations: iso-weighted, optic nerve prioritized, and macula prioritized. In each, the function values were depicted in a selection tool to select the optimal gazing angle(s). For example, patient 4 had a T2 equatorial tumor. The optimization applied for the straight gazing angle resulted in objective function values of 0.46 (iso-weighted situation), 0.90 (optic nerve prioritization) and 0.08 (macula prioritization) demonstrating the impact of that angle in different clinical approaches. CONCLUSIONS: The feasibility and suitability of a 3D MRI-based treatment planning approach have been successfully tested on a cohort of eight patients diagnosed with UM. Moreover, a gaze-angle trade-off dose optimization with respect to OARs sparing has been developed. Further validation of the whole treatment process is the next step in the goal to achieve both a non-invasive and a personalized proton therapy treatment.

Episodic Src activation in uveal melanoma revealed by kinase activity profiling.

BACKGROUND: The RAS/RAF/MEK/ERK pathway is involved in the balance between melanocyte proliferation and differentiation. The same pathway is constitutively activated in cutaneous and uveal melanoma (UM) and related to tumour growth and survival. Whereas mutant BRAF and NRAS are responsible for the activation of the RAS/RAF/MEK/ERK pathway in most cutaneous melanoma, mutations in these genes are usually absent in UM. METHODS: We set out to explore the RAS/RAF/MEK/ERK pathway and used mitogen-activated protein kinase profiling and tyrosine kinase arrays. RESULTS: We identified Src as a kinase that is associated with ERK1/2 activation in UM. However, low Src levels and reduced ERK1/2 activation in metastatic cell lines suggest that proliferation in metastases can become independent of Src and RAS/RAF/MEK/ERK signalling. Inhibition of Src led to the growth reduction of primary UM cultures and cell lines, whereas metastatic cell line growth was only slightly reduced. CONCLUSION: We identified Src as an important kinase and a potential target for treatment in primary UM. Metastasis cell lines seemed largely resistant to Src inhibition and indicate that in metastases treatment, a different approach may be required.

[A woman with iris heterochromia]. / Een vrouw met irisheterochromie.

A 20-year-old woman with congenital iris heterochromia presented with loss of vision of her right eye. We made de diagnosis of a large 'uvea melanoma' and enucleated the eye. Pathological examination showed an underlying oculodermal melanocytosis (ODM). The life-time risk of uveal melanoma in the general population is 0.7:100,000, but 1:400 in patients with ODM. Therefore, annual fundoscopy is recommended in these patients.

Biocompatibility of a fish scale-derived artificial cornea: Cytotoxicity, cellular adhesion and phenotype, and in vivo immunogenicity.

PURPOSE: To determine whether a fish scale-derived collagen matrix (FSCM) meets the basic criteria to serve as an artificial cornea, as determined with in vitro and in vivo tests. METHODS: Primary corneal epithelial and stromal cells were obtained from human donor corneas and used to examine the (in)direct cytotoxicity effects of the scaffold. Cytotoxicity was assessed by an MTT assay, while cellular proliferation, corneal cell phenotype and adhesion markers were assessed using an EdU-assay and immunofluorescence. For in vivo-testing, FSCMs were implanted subcutaneously in rats. Ologen(®) Collagen Matrices were used as controls. A second implant was implanted as an immunological challenge. The FSCM was implanted in a corneal pocket of seven New Zealand White rabbits, and compared to sham surgery. RESULTS: The FSCM was used as a scaffold to grow corneal epithelial and stromal cells, and displayed no cytotoxicity to these cells. Corneal epithelial cells displayed their normal phenotypical markers (CK3/12 and E-cadherin), as well as cell-matrix adhesion molecules: integrin-α6 and ß4, laminin 332, and hemi-desmosomes. Corneal stromal cells similarly expressed adhesion molecules (integrin-α6 and ß1). A subcutaneous implant of the FSCM in rats did not induce inflammation or sensitization; the response was comparable to the response against the Ologen(®) Collagen Matrix. Implantation of the FSCM in a corneal stromal pocket in rabbits led to a transparent cornea, healthy epithelium, and, on histology, hardly any infiltrating immune cells. CONCLUSION: The FSCM allows excellent cell growth, is not immunogenic and is well-tolerated in the cornea, and thus meets the basic criteria to serve as a scaffold to reconstitute the cornea.

Phase I and pharmacokinetic study of the oral farnesyl transferase inhibitor SCH 66336 given twice daily to patients with advanced solid tumors.

PURPOSE: A single-agent dose-escalating phase I and pharmacokinetic study on the farnesyl transferase inhibitor SCH 66336 was performed to determine the safety profile, maximum-tolerated dose, and recommended dose for phase II studies. Plasma and urine pharmacokinetics were determined. PATIENTS AND METHODS: SCH 66336 was given orally bid without interruption to patients with histologically or cytologically confirmed solid tumors. Routine antiemetics were not prescribed. RESULTS: Twenty-four patients were enrolled onto the study. Dose levels studied were 25, 50, 100, 200, 400, and 300 mg bid. Pharmacokinetic sampling was performed on days 1 and 15. At 400 mg bid, the dose-limiting toxicity (DLT) consisted of grade 4 vomiting, grade 4 neutropenia and thrombocytopenia, and the combination of grade 3 anorexia and diarrhea with reversible grade 3 plasma creatinine elevation. After dose reduction, at 300 mg bid, the DLTs consisted of grade 4 neutropenia, grade 3 neurocortical toxicity, and the combination of grade 3 fatigue with grade 2 nausea and diarrhea. The recommended dose for phase II studies is 200 mg bid, which was found feasible for prolonged periods of time. Pharmacokinetic analysis showed a greater than dose-proportional increase in drug exposure and peak plasma concentrations, with increased parameters at day 15 compared with day 1, indicating some accumulation on multiple dosing. Plasma half-life ranged from 4 to 11 hours and seemed to increase with increasing doses. Steady-state plasma concentrations were attained at days 7 through 14. A large volume of distribution at steady-state indicated extensive distribution outside the plasma compartment. CONCLUSION: SCH 66336 can be administered safely using a continuous oral bid dosing regimen. The recommended dose for phase II studies using this regimen is 200 mg bid.

A phase I and pharmacologic study of the MDR converter GF120918 in combination with doxorubicin in patients with advanced solid tumors.

BACKGROUND: Resistance to chemotherapy can partly be explained by the activity of membrane bound P-glycoprotein. Competitive inhibition of P-glycoprotein, by multidrug resistance (MDR) converters, may overcome this MDR. Previously studied MDR converters either have serious intrinsic side effects or considerably influence the pharmacokinetics of cytotoxic agents at concentrations theoretically required to convert MDR. GF120918 is a third-generation MDR converter with high affinity for P-glycoprotein and can be given orally. We performed a phase 1 study with escalating doses of GF120918 in combination with doxorubicin. PATIENTS AND METHODS: The study group comprised 46 patients with advanced solid tumors. Doxorubicin was administered on day 1 (cycle 1), GF120918 on days 22-24 (cycle 2), and on days 29-33 with doxorubicin administered on day 31 (cycle 3). Pharmacokinetics of both GF120918 and doxorubicin were studied. The starting daily dose of GF120918 was 50 mg and was to be increased in subsequent cohorts until a steady state plasma level of 100 ng/ml was reached. The starting dose of doxorubicin was 50 mg/m2 and was to be increased after reaching the target dose level of GF120918. RESULTS: In 37 of the 46 patients, full pharmacokinetic data from the three scheduled cycles were obtained. Pharmacokinetics of GF120918 showed a less than linear increase in Cmax with increasing doses, with considerable interpatient variation. The target steady-state plasma level for GF120918 was exceeded in 12 out of 19 patients who received 400 mg GF120918 alone twice daily and in 12 of 17 patients who received 400 mg GF120918 twice daily in combination with doxorubicin. GF120918 pharmacokinetics were not influenced by coadministration of doxorubicin. The doxorubicin AUC was only marginally influenced by GF120918 and only at the highest dose levels. In these patients there was a significant increase in the AUC of doxorubicinol in cycle 3 as compared to cycle 1. Hematologic toxicity mainly consisted of neutropenia and was more severe in cycle 3 than in cycle 1 (13 vs 5 patients with grade 4 neutropenia, P=0.003). Neutropenic fever was the dose-limiting toxicity at a doxorubicin dose of 75 mg/m2 with 400 mg GF120918 twice daily. The toxicity of GF120918 was limited to somnolence in eight patients and occasional gastrointestinal complaints. CONCLUSION: GF120918 is an MDR converter with only minimal side effects at a dose level yielding concentrations able to convert the action of P-glycoprotein in vitro. A doxorubicin dose of 60 mg/m2 on day 3 in combination with 400 mg GF120918 twice daily on days 1-5 is an acceptable regimen for further clinical trials.

The genetic basis of uveal melanoma.

Uveal melanoma (UM) is the most common intraocular malignancy in adults with an incidence of about 1/100,000 new cases per year in the Western world. Risk factors are having a light skin, blond hair and blue eyes. As some UM patients have a young age at diagnosis or an affected family history for UM or other malignancies, there may be an underlying genetic basis. This review discusses known or suspected risk factors for UM, the cancer risk in UM patients and their family members, and the genes that have been reported to predispose to UM (germline mutations) and tumor development (somatic mutations).

Lack of effect of different cytokines on expression of membrane-bound regulators of complement activity on human uveal melanoma cells.

Tumor cells are protected from antibody-dependent complement-mediated lysis by membrane-bound regulators of complement activation (m-RCA). m-RCA are expressed on uveal melanoma cells. We determined whether cytokine treatment affects expression of m-RCA on these cells in vitro. m-RCA expression on uveal melanoma cell lines was studied by flow cytometry, using monoclonal antibodies directed against CD46, CD55, and CD59. Cytokines studied were interferon-alpha (IFN-alpha), IFN-gamma, interleukin-1B (IL-1B), IL-12, and tumor necrosis factor-alpha (TNF-alpha). All three m-RCA were expressed on the uveal melanoma cell lines (CD59>>CD46>CD55), although in variable amounts. With a few exceptions, the cytokines had no effect on m-RCA expression. CD55 expression was not influenced by any of the cytokines. IFN-gamma downregulated expression of CD46 on one cell line (p < 0.01). TNF-alpha upregulated CD59 expression on two of the five cell lines (p < 0.012 and p < 0.001, respectively), which effect was dose dependent. IFN-alpha, IFN-gamma, IL1-beta, IL12, and TNF-alpha had limited effects on m-RCA expression on uveal melanoma cells in vitro.

A fluorescence staining method for the demonstration and measurement of lysosomal enzyme activities in single cells.

A cytochemical fluorescence method is described that makes possible simple, rapid, and specific demonstration and measurement of the activities of a wide variety of lysosomal enzymes in single cells using 4-methylumbelliferyl derivatives as substrates. The validity of the method and a number of applications using normal and mutant human cells are presented.

Relationship between natural killer cell susceptibility and metastasis of human uveal melanoma cells in a murine model.

PURPOSE: The purpose of this study was to determine the susceptibility of human uveal melanoma cells to in vitro and in vivo natural killer (NK) cell-mediated cytolysis and to determine if NK cells influence metastasis from the eye. METHODS: Four human uveal melanoma cell lines and one melanoma cell line derived from a metastatic lesion from a patient with uveal melanoma were tested for in vitro and in vivo NK cell-mediated lysis in a mouse model. Major histocompatibility complex (MHC) class I antigen expression was evaluated by flow cytometry. The role of NK cells in controlling the metastasis of uveal melanoma cells from the eye to the liver was examined in nude mice. RESULTS: Sensitivity to in vitro and in vivo lysis by human and murine NK cells was correlated with reduced expression of MHC class I antigens. Uveal melanoma lines expressing normal MHC class I antigen expression were insensitive to NK cell-mediated lysis, both in vitro and in vivo. Metastasis of uveal melanoma cells was inhibited by NK cell activity because disruption of in vivo NK function produced a sharp increase in the spontaneous metastasis of intraocular melanomas in nude mice. CONCLUSIONS: There is considerable variation in the susceptibility of human uveal melanomas to NK cell-mediated cytolysis. Susceptibility is closely correlated with reduced expression of MHC class I antigen expression. Disruption of NK cell function significantly increases the development of hepatic metastases from human uveal melanoma cells.

Human leukocyte antigen class I expression. Marker of poor prognosis in uveal melanoma.

PURPOSE: Because the expression of human leukocyte antigen (HLA) antigens is important for immunologic recognition of tumor cells, we determined expression of locus-specific HLA class I antigens in uveal melanoma and tested whether the level of HLA expression was related to prognosis or associated with known prognostic parameters. METHODS: Expression of HLA-A and -B antigens was determined on 30 formalin-fixed and paraffin-embedded sections of uveal melanoma by immunohistochemistry with locus-specific monoclonal antibodies and scored semiquantitatively. RESULTS: The level of expression of HLA-A and -B varied between uveal melanomas. Expression levels of HLA-A and -B were significantly correlated (P = 0.02). High HLA-B expression was significantly correlated with the presence of epithelioid cells (P = 0.04) in the tumor. Expression levels of HLA-A as well as of HLA-B, cell type, mitotic rate, Mib-1 score, and largest tumor diameter were significant predictive factors for survival. High expression of HLA-A and -B was associated with a decreased survival. Multiple Cox regression analysis with stepwise selection of covariates showed that the contribution of HLA-A expression to survival (P = 0.0003) exceeded that of tumor diameter (P = 0.02) and Mib-1 score (P = 0.04). CONCLUSIONS: Lack of expression of HLA-A as well as of HLA-B antigens on uveal melanoma is correlated with a better patient survival. Our data suggest that shedding of uveal melanoma micrometastases with a low expression of HLA class I into the systemic circulation may facilitate their removal and prevent the development of metastases. These findings support a protective role for natural killer cells in the development of metastatic disease in uveal melanoma.

Association between NM23-H1 gene expression and metastasis of human uveal melanoma in an animal model.

PURPOSE: To evaluate the role of nm23 gene expression in the development of metastases of human uveal melanomas in an animal model. METHODS: Seven human uveal melanoma cell lines and two murine skin melanoma cell lines were subjected to Northern blot analysis for the detection of nm23-H1 mRNA and to immuno-histochemistry to detect nm23 antigen. Each tumor cell line was transplanted intracamerally into nude mice, and the metastatic behavior was evaluated by histopathologic analysis of the livers and by determining host survival times. RESULTS: There was a strong inverse correlation between the levels of nm23 mRNA expression and nm23 antigen expression and the development of metastases of all seven human uveal melanomas and both murine skin melanomas transplanted intracamerally. Host survival time also was correlated with the degree of nm23 gene expression. CONCLUSIONS: The expression of nm23 mRNA and nm23 antigen in human uveal melanomas is correlated closely with reduced metastatic behavior in experimental animals and may serve as a sensitive prognostic indicator of malignancy and survival in patients with uveal melanomas.

Multidrug resistance-related proteins in primary choroidal melanomas and in vitro cell lines.

PURPOSE: Metastatic uveal melanoma is strongly resistant to chemotherapy, and multidrug resistance (MDR) may be involved. To investigate the role of MDR, the presence of the MDR-associated proteins P-glycoprotein (Pgp), MRP, and lung resistance protein (LRP) was determined on primary choroidal melanomas and cell lines. METHODS: A panel of primary choroidal melanomas was examined for the presence of MDR-associated proteins by immunohistochemical analysis. In cell lines established from four primary choroidal melanomas and one metastatic choroidal melanoma, the expression of MDR-associated proteins was determined with monoclonal antibodies in cytospin preparations and flow cytometry. In addition, the functional capacities of transporter proteins Pgp and MRP as adenosine triphosphate-driven efflux pumps were determined by measuring the cellular accumulation and efflux of the fluorescent dyes rhodamine 123 and calcein-AM, with and without the presence of specific pump inhibitors PSC833 and probenecid. RESULTS: Low levels of Pgp and MRP were detected in most primary tumors and in some cell lines. Measurable transporter function of Pgp could be determined in cell line OCM-1. Lung-resistance protein was present in all primary tumors and cell lines and showed high expression levels. CONCLUSIONS: This study revealed the involvement of LRP and at least a minor role of Pgp and MRP in chemoresistance of choroidal melanoma. Compared with cutaneous melanomas, uveal melanomas appear to express slightly higher levels of Pgp. These findings provide insights into the drug-resistant phenotype of this disease and can aid in the design of therapeutic protocols.

Neural cell adhesion molecule distribution in primary and metastatic uveal melanoma.

Tumor cell adhesion, detachment, and aggregation play an important part in tumor invasion and metastasis, and a variety of cell adhesion molecules have been found on tumor cells. Cell adhesion molecules, including those of the immunoglobulin superfamily, are associated with the development of metastatic behavior in cutaneous melanomas. The neural cell adhesion molecule (NCAM) belongs to this family. To investigate its possible role in the development metastatic behavior of uveal melanomas, the authors studied immunohistochemically the expression of NCAM by using an antibody that recognizes all three major isoforms of NCAM and an antibody that recognizes the HNK-1 epitope present on some isoforms of NCAM. The authors studied 32 primary uveal melanomas from 32 patients (among these, 12 were rapidly metastasizing and 16 slowly metastasizing) and 29 metastases from 19 patients. From 13 patients the primary, as well as the metastatic, tumors were available. With one exception, all HNK-1 positive primary and metastatic tumors were also positive for NCAM. NCAM was significantly more expressed in aggressive, rapidly metastasizing primary tumors (P = .02 and .04, respectively) and in metastases. HNK-1 was significantly (P = .04) more expressed in larger tumors. In liver metastases HNK-1 immunoreactivity was significantly (P = .005) less frequently expressed than NCAM. Therefore, NCAM isoforms that lack the HNK-1 epitope might play a role in the organ specific metastatic behavior of uveal melanomas.

No demonstrated effect of pre-enucleation irradiation on survival of patients with uveal melanoma.

PURPOSE: To evaluate the hypothetical effect of pre-enucleation irradiation on survival of patients with uveal melanoma. METHODS: In a prospective study between 1978 and 1990, 145 patients with uveal melanoma were treated by irradiation in two fractions of 4 Gy before enucleation. A historical control group of 89 patients with uveal melanoma treated by enucleation alone was operated on between 1971 and 1990. Patients were followed up until December 1992 or until death. The mean follow-up period was 65 months in the irradiated group and 88 months in the control group. RESULTS: The preoperatively irradiated group of patients showed no significant improvement of the survival rate after 7 1/2 years (75.9%) compared with the control group (72.1%). Preoperative irradiation was not associated with survival (P = .93), as assessed by Cox proportional hazard analysis, adjusted for age, gender, tumor location, tumor size, cell type, and year of enucleation. Women in both the irradiated and control groups had a better prognosis than men (P = .002). CONCLUSION: Preoperative irradiation in this nonrandomized study had no effect on survival of patients with uveal melanoma.

Iris-claw phakic intraocular lens for high myopia.

PURPOSE: To evaluate the efficacy, safety, predictability, and stability of implanting a polymethylmethacrylate (PMMA) phakic intraocular lens (PIOL) (the Artisan myopia lens) to correct high myopia. METHODS: An Artisan myopia lens was implanted in 78 consecutive eyes of 49 patients with preoperative myopia that ranged from -6.25 to -28.00 D. Mean patient age was 42.4 years. Mean follow-up was 10.7 months and all patients were followed for at least 6 months; 45 eyes had follow-up of 12 months, and 10 eyes had 24 months. The desired outcome was emmetropia in all eyes except for those eyes with preoperative myopia greater then -23.00 D. RESULTS: Fifty-three eyes (67.9%) had a postoperative refraction at the last follow-up examination within +/-1.00 D of emmetropia, and 39 eyes (50.0%) had a postoperative refraction +/- within 0.50 D of emmetropia. The postoperative refraction remained stable during the entire follow-up period. Mean spectacle-corrected visual acuity improved from 20/32 preoperatively to 20/25 postoperatively. Mean postoperative uncorrected visual acuity was 20/32. There was no significant change in endothelial cell density from baseline. We did not encounter major complications. CONCLUSION: Implantation of the Artisan myopia lens to correct high myopia resulted in a stable and fairly predictable refractive outcome. A significant endothelial cell change was not detected.

DNA flow cytometry in uveal melanoma: the effect of pre-enucleation irradiation.

BACKGROUND: For uveal melanoma it has been demonstrated that aneuploidy correlates with worse clinical outcome. However, a striking variation in incidence of aneuploidy is reported for uveal melanomas. METHODS: Flow cytometry was used to study retrospectively DNA-ploidy of 132 uveal melanomas on paraffin embedded material. Thirty five patients received 2 x 4 Gy doses of irradiation 24 and 48 hours before enucleation. Correlation between DNA-ploidy and histopathological grading, largest tumour diameter, tumour height, tumour location, scleral invasion, and TNM classification was assessed. Survival analysis methods were used to investigate the predictive value of these variables on clinical outcome. RESULTS: Of the tumours 37% were aneuploid and 63% were diploid. Intratumour ploidy heterogeneity was minimal (92% concordance). A strong correlation (p = 0.009) was found between DNA-ploidy and cell type. No correlation was found between DNA-ploidy and other conventional prognostic variables. Irradiated melanomas were significantly more aneuploid than non-irradiated tumours (p < or = 0.01). CONCLUSION: In survival analysis DNA-ploidy and the largest tumour diameter were significant in predicting metastatic outcome (p < or = 0.03 and 0.01 respectively); histological cell type and tumour location were of borderline significance.