Chromosomal aberrations in iris melanomas.

BACKGROUND: Uveal melanomas can develop in the choroid, ciliary body and iris. In choroidal and ciliary body melanomas, specific chromosomal changes correlate with metastatic disease. Iris melanomas have a better prognosis than choroidal melanomas, and it would be interesting to know if they share chromosomal changes. In addition, iris melanomas might harbour UV-induced mutations of tumour suppressor genes, such as PTEN and CDKN2A. METHODS: Twenty iris melanomas were analysed for chromosome 1p, 3, 6, 8, 9p and 10q abnormalities using fluorescence in situ hybridisation. These results were correlated to clinical follow-up data using statistical analyses. RESULTS: (Partial) loss of chromosome 3 was observed in nine iris melanomas, and gain of 8q was present in seven tumours. Loss of chromosome 9p was demonstrated in seven tumours, but no deletions of the PTEN region on chromosome 10 were found. Three patients died of metastatic disease, and one patient developed liver metastases, but is still alive. Univariate analysis indicated a lower disease-free survival for patients with diffuse growing melanomas (p=0.01), melanomas that lost a copy of chromosome 3 (p=0.03), or invading the ciliary body (p=0.01). In a multivariate analysis, none of the correlations were significant. CONCLUSION: Loss of chromosome 3 as well as loss of chromosomal region 9p21 (that entails tumour suppressor gene CDKN2A) plays a role in iris melanoma. A firm correlation with disease-free survival could not be established, possibly due to the small sample size.

Epistaxis or epiphora as a sign for extension of a conjunctival melanoma. A series of six patients with nasolacrimal recurrence.

PURPOSE: To characterise malignant conjunctival melanomas with extension and recurrence in the nasolacrimal system. METHODS: Localisation of the primary tumour and recurrences of 210 conjunctival melanomas treated in The Netherlands were reviewed for orbital and nasal tumours (1978-2008). Based of these cases and literature data, characteristics for nasolacrimal system extension and metastasis were reviewed. RESULTS: Six patients (3%) showed a recurrence of the primary conjunctival melanoma in the nasolacrimal system. Two of the six primary tumours were limbal tumours; the other four were diffuse tumours involving the fornix. In all six patients, the primary conjunctival melanomas were associated with primary acquired melanosis. During the follow-up period (11.6±3 years, range 3.4-28.5 years, median 8.7 years) two patients developed metastases and died. CONCLUSIONS: Patients should be advised to contact their treating ophthalmologist in the case of symptoms of epiphora, nose obstructions and epistaxis, especially non-bulbar and diffuse cases associated with primary acquired melanosis.

Angiosarcoma of the eyelid and periorbital region. Experience in Leiden with iridium192 brachytherapy and low-dose doxorubicin chemotherapy.

AIM: To report on the use of iridium(192) brachytherapy and doxorubicin chemotherapy as adjuvant therapy in 6 patients with angiosarcoma of the eyelid and periorbital region. MATERIAL AND METHODS: Tumor localization and diameter, signs of inflammation, histology and treatment are discussed in this retrospective study of 6 patients (age 46-87 yrs.) presenting with primary angiosarcoma in the eyelid. RESULTS: Six patients (4 elderly) with angiosarcoma localized in one or more eyelids, the face or multilocular were seen between 1987 and 2000. In one patient, a small nodular tumor did not recur within 4 years after radical excision. In another patient, the tumor was treated with surgery and iridium(192) wire brachytherapy and did not recur in 17 years of follow-up. In four patients with large diffuse tumors that were treated with doxorubicin, partial regression was achieved. The follow-up was >3 years (median 5 years). CONCLUSION: If radical surgery for angiosarcoma of the eyelid and periorbital region is not possible, adjuvant iridium(192) wire brachytherapy may prove beneficial. For widespread, diffusely growing tumors, and in elderly patients, low-dose (slowly, 20 mg i.v.) doxorubicin can be used weekly as adjuvant therapy, resulting in partial regression and longer survival rates than previously published by other authors.

Merkel cell carcinoma of the eyelid review of the literature and report of patients with Merkel cell carcinoma showing spontaneous regression.

PURPOSE: To evaluate the clinical and histopathologic characteristics of Merkel cell carcinoma (MCC) of the eyelid. DESIGN: Retrospective case series and literature review. PARTICIPANTS: Three consecutive patients with MCC of the eyelid who were referred to the Ocular Oncology Unit of Leiden University Medical Center, Netherlands. METHODS: Clinical records and histopathologic material of patients with eyelid MCC were reviewed. The clinical presentation and treatment were evaluated. MAIN OUTCOME MEASURES: Clinical and histopathologic description of eyelid MCC, with histologic proof of spontaneous regression of the tumor. RESULTS: Three patients with MCC of the eyelid were included. Diagnosis was made by pathologic investigation and immunohistochemistry (S100, cytokeratin 20, epithelial membrane antigen, chromogranin). Two of the patients showed histologically proven complete spontaneous regression after nonradical excision of the tumor. After local excision, none of the MCCs demonstrated local recurrence, without regional or distant metastases. Mean clinical follow-up was 50 months. CONCLUSIONS: Nonocular MCC is known to recur in 66% of patients and to be lethal in almost 33%. Merkel cell carcinoma of the eyelid is a rare malignancy that can not be recognized clinically. Clinical differential diagnosis must be made with a chalazion, and histopathologic differential diagnosis must be made with small cell carcinomas. Close follow-up of these patients is advised because of the potential high recurrence rate and lymphatic spread. The immunologic phenomenon of spontaneous regression points out the importance of the immune system in this disease.

Impression cytology of melanocytic conjunctival tumours using the Biopore membrane.

PURPOSE: To compare a new Biopore membrane impression cytology method with the routinely used exfoliative cytology in patients with a melanocytic lesion of the conjunctiva. METHODS: Sixty-eight consecutive patients with a conjunctival melanocytic lesion underwent Biopore membrane impression cytology as well as exfoliative cytology. A histologic sample was also available in 26 cases. All Biopore samples were stained immediately with RAL 555. Both Biopore and exfoliative cytology samples were assessed by two cytopathologists and graded into four different categories of atypia. RESULTS: Twenty-three out of 26 Biopores and 20 out of 24 for the exfoliative smears correlated with the corresponding histologic sample. Biopore cytology resulted in higher numbers of cells with a greater density compared to exfoliative cytology. CONCLUSIONS: Biopore cytology can be used for cytologic sampling of conjunctival melanocytic lesions. Because of the larger amount and higher density of cells obtained with the Biopore membrane, interpretation by a pathologist is easier and faster. Sampling of the fornix, caruncula, and ocular material in children is difficult with the Biopore method, and exfoliative cytology seems to be the favorable test in those situations.

Screening for conjunctival melanoma metastasis: literature review.

Local tumour control in conjunctival melanoma has improved in recent years. However the tumour-related death rate of these patients is still 14% at 5 years up to 33% at 15 years. With the introduction of sentinel node biopsies for conjunctival melanomas with a poor prognosis and screening for locoregional and distant metasases prognosis might be improved.

Immunophenotypic markers to differentiate between benign and malignant melanocytic lesions.

BACKGROUND/AIMS: The authors investigated the expression of S100A1, S100A6, S100B, MelanA, and CEA in conjunctival naevi, primary acquired melanosis (PAM), conjunctival melanoma, and uveal melanoma in order to assess their potential usefulness in the pathological differential diagnosis of these entities. METHODS: Paraffin embedded sections of 18 conjunctival naevi, 14 PAM, 16 conjunctival melanomas, and 20 uveal melanomas were immunostained for S100A1, S100A6, S100B, MelanA, and CEA, and expression was scored semiquantitatively. RESULTS: Expression of S100A1 differed significantly between conjunctival naevi and conjunctival melanoma, with percentages of positive cells of 30.6% and 71.4%, respectively. Conjunctival melanomas had high average scores for S100A1 and S100B (71.4%, 62.9%, respectively), while uveal melanomas also had high S100A1 but low S100B scores (88.5%, 18.5%, respectively). MelanA was highly variable; naevi and uveal melanoma had higher average scores than conjunctival melanoma. CEA was hardly detectable in all four groups. CONCLUSION: S100A1 seems to be a possible candidate to differentiate conjunctival naevi from conjunctival melanoma. S100B seems to differentiate between uveal melanoma and conjunctival melanoma. However, the study size was small and therefore the data have to be confirmed by others.

Ciliary tissue transplantation in the rabbit.

Irreversible damage of the ciliary body can be responsible for prolonged ocular hypotony and phthisis bulbi, which, currently, cannot be treated. The aim of this study was to achieve survival of morphologically normal ciliary tissue (CT) transplants in the anterior chamber of a rabbit's eye. Outbred female New Zealand albino rabbits received CT allografts, which were placed on to the surface of the host iris. We evaluated the influence of ciclosporin (CsA), VEGF and donor perfusion on graft survival. Operated eyes were assessed clinically and histologically, and revascularization of the grafts was determined by fluorescein angiography. All grafts became dark and ischemic during the first five to seven days after transplantation. Reperfusion of the grafted tissue was complete at approximately ten days after transplantation. In untreated animals, transplants became infiltrated by inflammatory cells, which led to destruction of the tissue. This was prevented by systemic use of CsA. Transplants treated with VEGF prior to transplantation had fewer ischemic areas but epithelial cell survival was not improved. Whole body donor perfusion prior to preparation of the grafts resulted in less inflammation and, histologically, in a better quantity and quality of the epithelial cells in the CT transplants. Ciliary tissue can be successfully transplanted but the ciliary epithelium suffers from ischemia and in untreated animals the whole transplant is rejected in the classical fashion. If the donor is perfused and the host immunosuppressed, histologically normal ciliary epithelium can be preserved together with rapid revascularization, minimal inflammation and good survival of the transplant, although fibrosis continued to occur during the two months after transplantation.

Topical alpha-interferon in recurrent conjunctival papilloma.

PURPOSE: To report the effect of topical alpha-interferon (alpha-IF) in two cases of recurrent conjunctival papillomas. METHODS: One patient with a conjunctival papilloma and one patient with a limbus-based papilloma were treated with excision and cryotherapy. Recurrences included one in situ carcinoma. Retreatment included systemic and topical alpha-IF. RESULTS: In Case 1, the papillomas regressed with systemic alpha-IF followed by topical application of alpha-IF (2.8 x 106 U) for 1 year against recurrence. In Case 2, the papillomas disappeared with topical alpha-IF over 1 year. No recurrences were seen during follow-up (84 and 91 months, respectively). The possible side effect of topical alpha-IF was superficial keratitis. Retrospective polymerase chain reaction tests for HPV were positive and showed type 6 in Case 1 and type 33 in Case 2. CONCLUSION: Topical alpha-IF can be used as adjuvant therapy in recurrent conjunctival papillomas.

Expression of epidermal growth factor receptor: risk factor in uveal melanoma.

PURPOSE: To investigate the prognostic significance of the expression of epidermal growth factor receptor (EGFR) in uveal melanoma. EGFR is a transmembrane glycoprotein, and its expression has been correlated with the development of metastases in various malignancies. METHODS: Frozen sections from 22 primary uveal melanomas were examined for EGFR expression by a three-step immunoperoxidase staining, using a mouse anti-human EGFR IgG2b monoclonal antibody. The results were compared with patient survival and clinical and histopathologic parameters. RESULTS: EGFR expression could not be determined on one tumor due to excessive pigmentation. Two patients died of causes unrelated to melanoma, and two patients were lost to follow-up. Out of 21 tumors, six tumors showed immunoreactivity for EGFR. Five of these six patients (83%) died due to metastases, compared with 2 (17%) of 12 patients with no EGFR expression (Kaplan-Meier analysis P = 0.0004). EGFR-positive tumors tended to have a greater tumor prominence and a higher mitotic rate. CONCLUSIONS: The expression of EGFR was significantly correlated with death due to metastatic disease and therefore can be regarded as an important prognostic factor in human uveal melanoma.

Late ophthalmological complications after total body irradiation in non-human primates.

PURPOSE: To investigate the long-term effects of total body irradiation (TBI) on the incidence and time course of ocular complications. MATERIALS AND METHODS: Rhesus monkeys treated with TBI photon doses up to 8.5 Gy and proton doses up to 7.5 Gy were studied at intervals up to 25 years post-irradiation. They were compared with control groups with a similar age distribution. Cataract formation and ocular fundus lesions were scored according to a standardized protocol. Fluorescein angiography and histopathology was performed in selected animals. RESULTS: Cataract formation occurred after a latent period of 3-5 years. Significant cataract induction was observed for photon-doses of 8 and 8.5 Gy and beyond 20 years after proton irradiation. The severity of the lesions represents significant impairment of vision and would require cataract surgery if similar results occurred in human bone marrow transplant patients. Fluorescein angiography demonstrated a normal pattern of retinal vessels in 13 out of 14 animals (93%) from the irradiated group and in eight out of nine animals (89%) from the control group. No additional lesions apart from age-related degenerative changes could be demonstrated. Histological evaluation revealed no radiation-associated vasculopathy. CONCLUSIONS: Radiation alone for doses up to 8.5 Gy of photons does not carry a potential risk for fundus pathology, whereas clinically important cataract induction should be anticipated within 5 years after photon doses of 8.0 and 8.5 Gy and proton doses in excess of 2.5 Gy.

Histopathological findings in human choroidal melanomas after transpupillary thermotherapy.

AIMS: The effect of transpupillary thermotherapy (TTT) on human choroidal melanomas was investigated by means of histopathology. METHODS: Before enucleation TTT was performed in 11 eyes with a xenon are photocoagulator with a red filter or a diode laser at 810 nm. The exposure time was 1 minute; the estimated temperature at the top of the tumour was about 65 degrees C. RESULTS: Seven of 11 tumours developed necrosis to a maximum depth of 3.9 mm with a sharp demarcation between the necrotic and the viable part of the tumour. The depth correlated with penetration of heat into the tumour. Scattered small haemorrhages in the transitional zone between the necrotic and the viable part of the tumour were observed in three eyes but large haemorrhages were absent. Ocular media were not affected owing to the low rate of absorption of radiation at 810 nm. TTT did not cause significant scleral damage. Intrascleral tumour cells with a viable appearance were observed in one eye, where the tumour was almost totally necrotic. CONCLUSION: Results show that TTT has potential as a conservative therapeutic treatment for choroidal melanomas.

Superior orbital fissure syndrome caused by intraorbital spread of a cutaneous squamous cell carcinoma and not detected on computed tomography and magnetic resonance imaging.

Intraneural and perineural spread of a squamous cell carcinoma of the frontal region via the orbit to the cranial cavity is a rare cause of a superior orbital fissure syndrome. This mode of tumor spread, for which a previously excised invasive malignant tumor is responsible, is rarely reported. The absence of an intraorbital mass and the fact that computed tomography (CT) and magnetic resonance imaging (MRI) are normal make the diagnosis extremely difficult. We describe a 76-year-old patient in whom both intraneural and perineural tumor growth deep into the orbit caused acute oculer motility disturbances, visual impairment, and ocular dysfunction with subsequent cerebral and vascular dysfunction. Surgery, radiotherapy, and orbital exenteration did not prevent spread of the tumor toward vital intracranial and intracerebral structures. Even the advanced diagnostic modalities of CT and MRI failed to reveal the cause of the pathologic process in our patient. In such a case, one should look carefully for both intraneural and perineural tumor invasion on previous histologic material and, if positive, treat the patient with radical surgery on purely clinical grounds as early as possible.

HLA expression in a primary uveal melanoma, its cell line, and four of its metastases.

BACKGROUND: The level of HLA expression on a tumour may influence the immunological response against this tumour, and vice versa. HLA expression was determined in a primary uveal melanoma, its metastases, and on a cell line derived from this melanoma, and the presence and type of infiltrate in tissue sections were also studied. METHODS: Immunohistochemistry with monoclonal antibodies (MAbs) against HLA class I and II, T cells, NK cells, and macrophages. RESULTS: Primary and metastatic lesions, as well as the cell line showed high levels of expression of the monomorphic determinants of HLA class I. Expression of the polymorphic HLA-A2 and HLA-A3 antigens was decreased on metastases to the skin and liver. HLA-Bw4 expression was low on all lesions, as well as expression of HLA class II. Tumour infiltrating cells consisted mainly of CD3, CD4, and CD8 positive cells. Expression on the cell line corresponded to expression on the primary tumour. CONCLUSION: The primary uveal melanoma as well as the cell line showed a high expression of monomorphic and polymorphic HLA-A antigens, while metastases showed a high expression of monomorphic and a lower expression of polymorphic antigens. This variation in expression may support tumour cell escape from NK cells as well as CTL mediated lysis.

A histopathological, ultrastructural and immunohistochemical study of congenital hereditary retinoschisis.

OBJECTIVE: To confirm our earlier histopathological and electron microscopic findings in congenital hereditary retinoschisis (CHRS) in two additional globes and to further evaluate the nature and origin of the intraretinal filaments by means of immunohistochemical analysis. PATIENTS: Three white men with CHRS, aged 83 years (patient I) (two globes), 55 years (patient 2) (two globes) and 33 years (patient 3, nephew of patient 2) (one globe). OUTCOME MEASURES: Findings on histopathological study and electron microscopy (patient I) and immunohistochemical analysis (all five globes). RESULTS: Histopathological examination showed extensive extracellular deposition of amorphous material positive for periodic acid-Schiff reagent in the outer schisis layer and focally in the macula. Ultrastructurally, the amorphous material represented filaments measuring 8 to 12 nm in diameter within degenerated Müller cells, with accumulation of these filaments in adjacent extracellular spaces. Similar, less severe changes were seen in the superonasal retina. Immunohistochemical studies showed focal reactivity for glial fibrillary acidic protein (GFAP) in the retina adjacent to the schisis cavity in all five globes, focal reactivity for S-100 protein in four retinas, rare focal staining for vimentin and neurofilaments in two retinas each and no reactivity for type I keratin or actin. CONCLUSIONS: The present study corroborates our previous work and provides pathological evidence that the retinal disorder extends beyond the limits of the schisis. The results of the immunohistochemical analysis are consistent with a glial cell origin of the filaments. We postulate that defective Müller cells produce GFAP and possibly S-100 protein, which accumulate within the retina and secondarily result in degeneration of these cells and schisis formation.

Differential expression of HLA-A and B-alleles on uveal melanoma as determined by immuno-histology.

HLA molecules play an important role in the presentation of antigens to the immune system, including tumor-specific antigens. Uveal melanomas vary in the level of expression of monomorphic HLA molecules. However, since the HLA system is polymorphic and since antigen-presentation may be linked to the expression of specific HLA alleles, the authors wondered whether allelic differences in expression existed on uveal melanomas. In order to test this, tissue sections from 23 uveal melanomas were stained in an indirect immunoperoxidase technique with monoclonal antibodies against monomorphic and polymorphic determinants of HLA molecules. All uveal melanomas showed a high level of expression of the monomorphic determinants of HLA-Class I. The polymorphic HLA-Class I molecules A2, A3, Bw4 and Bw6 varied in expression, with a higher expression of HLA-A than of HLA-B. A low level of expression of both ß2-microglobulin and HLA-B locus products was associated with a large tumor diameter. Expression of HLA-Class II molecules was low (0 to 35%). The observation that expression of the HLA-A allelic products was higher than of the HLA-B subtypes may have implications for the search of tumorspecific peptides for immunotherapeutic use: it may be worthwile to select peptides that specifically bind to HLA-A and not to HLA-B.

Hamster Greene melanoma in the rabbit eye: immunosuppressive treatment to improve this tumor model.

BACKGROUND: The hamster Greene melanoma (HGM) implanted in the anterior chamber of the rabbit eye has been used to study experimental therapies for human uveal melanoma. However, the occurrence of spontaneous necrosis limits the value of this model for long-term evaluation of experimental treatments. In the present study we tested the hypothesis that an immune response is the cause of this necrosis and that prevention of the immune response can prolong the experimentation time METHODS: HGM was implanted in the anterior chamber of control, presensitized and immunosuppressed rabbits. The effects of sensitization and immunosuppression were assessed by clinical and histological observation RESULTS: Sensitization led to a significant slowing down of tumor growth, but not to a difference in necrosis. Immunosuppressive treatment with cyclosporin A improved the success rate of implantation and decreased the amount of necrosis in the tumor CONCLUSION: The immune response plays a role in the occurrence of necrosis. However, although immunosuppressive treatment with cyclosporin A decreased the amount of necrosis, significant necrosis still occurred, suggesting that other factors like angiogenesis play a part as well and still limit the usefulness of this model in the long-term evaluation of experimental therapies.

Tapioca melanoma of the iris.

Clinical identification of tapioca melanoma of the iris is important because its medical treatment may differ from that of other malignant iris melanomas. The characteristic iris nodules must be differentiated from granulomatous uveitis, metastases, and Lisch nodules (neurofibromatosis). We will discuss the anterior segment findings, secondary glaucoma, and fluorescein iris angiographic and histopathologic data from two patients, one with a single nodular type and one with a seeding type of tapioca melanoma of the iris.

Transpupillary thermotherapy (TTT) by infrared irradiation of choroidal melanoma.

We studied the destructive effect of hyperthermia at sub-photocoagulation level of 45-60 degrees C on melanomas. Optimal conditions for spreading of heat into tissue are a wavelength of 700-900 nm, a temperature of 45-60 degrees C, an exposure time of 1 minute or more, and a beam diameter of several millimeters. In hamsters with subcutaneous melanomas we obtained a tumour necrosis of 6 mm depth at 60 degrees C and one minute exposure time. We performed transpupillary thermotherapy (TTT) with a diode laser at 810 nm in patients with choroidal melanomas prior to enucleation. Treatment is based on the fortunate situation that irradiation at this wavelength combines optimal tissue penetration with a low absorption by clear ocular media of 5% or less. In 3 TTT-treated eyes histopathology showed a depth of necrosis of 0.9, 3.4, and 3.9 mm. TTT may become a new useful treatment modality for choroidal melanoma but its ultimate value has yet to be assessed.

Possible relation between HLA and ABO type and prognosis of uveal melanoma.

Not only local parameters but also genetic determinants like the HLA genes may play a role in the development and clinical behavior of malignant tumors. In skin melanoma the presence of HLA-B40 is associated with a poor prognosis. We tested the hypothesis that the clinical behavior of uveal melanoma is influenced by the HLA type of the patient. The HLA types of 44 patients with uveal melanoma had been determined before operation with a view to using the cornea of the enucleated eye for an HLA-matched corneal transplantation. We compared the ABO and HLA types of the patients with the development of metastases and with the ten-year patient survival. An association was observed between the presence of HLA-B40 and death due to metastasis of uveal melanoma. We conclude that the HLA type of the patient may influence the clinical behavior not only of skin melanoma but also of uveal melanoma.