HER2 Overexpression in Retinoblastoma: A Potential Therapeutic Target?

BACKGROUND: Retinoblastoma (RB) is the most common primary intraocular malignancy. Current therapies are associated with high morbidity in the short- and long-term. Human epidermal growth factor receptor 2 (HER2) is a transmembrane protein detected in 15-30% of breast cancers, but it has also been described in other malignancies. Recently, it has been claimed that a truncated version of this protein is expressed in RB, responsive to directed therapies in vitro. We scored HER2 overexpression in RB tissue samples and discussed its potential clinical utility. METHODS: HER2 overexpression was investigated using immunohistochemistry; the overexpression was evaluated with a score ranging from 0 to 3+ according to the membranous staining pattern in archival formalin-fixed, paraffin-embedded RBs. RESULTS: A total of 60 RB cases and a RB cell line (Y79) were considered. The mean age at enucleation was 31.6 ± 31.5 months. The mean time from diagnosis to enucleation was 11.8 ± 11.2 months (range 1-44). Five (8%) cases were multifocal. HER2 overexpression was negative in all RB cases (49 cases scored 0 and 11 scored 1+) and in the Y79 cell line. CONCLUSIONS: Overall, we were not able to demonstrate the overexpression of HER2. Further studies should clarify and better elucidate the potential role of HER2-targeted therapies in RB.

Immunohistochemical analysis of retinoblastoma cell phenotype using neuronal and glial cell markers / Análise imuno-histoquímica do fenótipo de células de retinoblastoma utilizando marcadores de células neuronais e gliais

ABSTRACT Purpose: The cellular origin of retinoblastoma is uncertain as constituent tumor cells heterogeneously express markers of both immature and mature retinal cells. An immunohistochemical analysis of cellular origin may yield valuable insights into disease progression and treatment options. This study aimed to determine the cellular origin of retinoblastoma in a large case series and correlate these findings with histopathological prognostic factors. Methods: Thirty-nine retinoblastoma cases were histopathologically diagnosed and analyzed by immunohistochemistry using monoclonal antibodies against the immature neural cell marker SRY-box containing gene 2 (SOX-2), the mature neuronal cell marker microtubule-associated protein 2 (MAP2), and the mature glial cell marker glial fibrillary acidic protein (GFAP). Histopathological features were also evaluated, including patterns of growth, differentiation, vitreous seeding, and choroidal/scleral, optic nerve, and anterior chamber invasion. Two retinoblastoma cell lines, WERI-1 and Y79, were studied by immunocytochemistry using the same antibodies. Results: Expression of SOX-2 was strong in 97.4% of retinoblastoma cases, while MAP-2 was expressed in 59% of cases. Immunostaining for GFAP was positive only in reactive stromal astrocytes interspersed amongst tumor cells and in peritumoral tissue. There was no correlation between histopathological prognostic factors and immunohistochemical markers. Retinoblastoma cell lines showed strong positivity for SOX2 (90% of WERI-1 cells and 70% of Y79 cells) and MAP2 (90% of cells in both lines). GFAP was completely negative in both cell lines. Conclusion: The majority of retinoblastomas and both RB cell lines expressed an immature neural and/or a mature neuronal cell marker, but not a glial marker. These results indicate a typical neuroblast or neuronal origin and eliminate astrocyte differentiation from neural stem cells as the source of retinoblastoma.

RESUMO Objetivos: Este estudo visa determinar a origem do retinoblastoma em um número de casos e correlacionar essos achados com fatores prognósticos e histopatológicos conhecidos. Métodos: Trinta e nove casos de retinoblastoma foram diagnosticados e analisados com imuno-histoquímica usando marcadores de anticorpos monoclonais contra as células de retina imaturas (SOX-2: SRY-box containing gene 2), contra as células da retina maturas (MAP2: microtubule -associated protein 2) e contra as células gliais maturas (GFAP: glial fibrillar acidic protein). Foram avaliadas características microscópicas dos casos (grau de diferenciação, presença de semeadura vítrea, invasão de coroide/esclera, nervo óptico e câmara anterior). Duas linhas celulares de retinoblastoma (WERI-1 e Y79) também foram testadas, utilizando os três marcadores. Resultados: A expressão de SOX-2 foi positiva em 97,4% dos casos de retinoblastoma, enquanto MAP2 foi positivo em 59% dos casos. GFAP foi apenas positivo no estroma (astrócitos reativos). Não houve correlação entre preditores histopatológicos e marcadores imunohistoquímicos avaliados. As linhagens celulares mostraram positividade para SOX-2 (90% em WERI-1 e 70% das células Y79). Ambas as linhagens celulares se mostraram fortemente positivas con MAP2 (90%), enquanto não houve expressão de GFAP em nenhuma das linhas celulares estudadas. Conclusões: A maioria das células de retinoblastoma desta série de casos expressa marcadores de células retinianas imaturas, além de marcadores de células maduras. As linhas celulares Y79 e WERI-1 apresentaram imunomarcação para ambos os marcadores neurais em percentagens semelhantes a dos casos avaliados. Portanto, estes resultados confirmam a origem neural do tumor em particular. Alem disso, a ausência de células positivas para GFAP no tumor descarta diferenciação de astrócitos em retinoblastoma.

Immunohistochemical analysis of retinoblastoma cell phenotype using neuronal and glial cell markers.

PURPOSE:: The cellular origin of retinoblastoma is uncertain as constituent tumor cells heterogeneously express markers of both immature and mature retinal cells. An immunohistochemical analysis of cellular origin may yield valuable insights into disease progression and treatment options. This study aimed to determine the cellular origin of retinoblastoma in a large case series and correlate these findings with histopathological prognostic factors. METHODS:: Thirty-nine retinoblastoma cases were histopathologically diagnosed and analyzed by immunohistochemistry using monoclonal antibodies against the immature neural cell marker SRY-box containing gene 2 (SOX-2), the mature neuronal cell marker microtubule-associated protein 2 (MAP2), and the mature glial cell marker glial fibrillary acidic protein (GFAP). Histopathological features were also evaluated, including patterns of growth, differentiation, vitreous seeding, and choroidal/scleral, optic nerve, and anterior chamber invasion. Two retinoblastoma cell lines, WERI-1 and Y79, were studied by immunocytochemistry using the same antibodies. RESULTS:: Expression of SOX-2 was strong in 97.4% of retinoblastoma cases, while MAP-2 was expressed in 59% of cases. Immunostaining for GFAP was positive only in reactive stromal astrocytes interspersed amongst tumor cells and in peritumoral tissue. There was no correlation between histopathological prognostic factors and immunohistochemical markers. Retinoblastoma cell lines showed strong positivity for SOX2 (90% of WERI-1 cells and 70% of Y79 cells) and MAP2 (90% of cells in both lines). GFAP was completely negative in both cell lines. CONCLUSION:: The majority of retinoblastomas and both RB cell lines expressed an immature neural and/or a mature neuronal cell marker, but not a glial marker. These results indicate a typical neuroblast or neuronal origin and eliminate astrocyte differentiation from neural stem cells as the source of retinoblastoma.

Expression of SIRT2 and SIRT6 in retinoblastoma.

PURPOSE: SIRT2 and SIRT6 are members of the sirtuin family and are associated with cancer development and progression in certain tumours, but their expression in retinoblastoma has not been studied. The primary objective of our study was to determine the expression of SIRT2 and SIRT6 in human retinoblastoma cases. METHODS: Eighteen formalin-fixed paraffin-embedded blocks of retinoblastoma cases from the Ocular Pathology Registry at the Henry C. Witelson Ocular Pathology Laboratory were obtained, classified and immunostained for SIRT2 and SIRT6 using mouse monoclonal antibodies. RESULTS: Sixteen cases were poorly differentiated retinoblastoma cases. SIRT2 and SIRT6 were expressed in all cases of retinoblastoma although differences in the staining intensity were found between cases. SIRT2 and SIRT6 expression was also observed in various normal structures of the remaining ocular tissue. CONCLUSIONS: SIRT2 and SIRT6 are expressed in retinoblastoma, as well as in some normal ocular structures. While precise roles of these proteins must still be determined in retinoblastoma, their expression profiles suggest that further functional studies of both SIRT2 and SIRT6 should be pursued in this cancer.

Diagnosing Pathological Prognostic Factors in Retinoblastoma: Correlation between Traditional Microscopy and Digital Slides.

OBJECTIVE: It was the aim of this study to determine the diagnostic accuracy of high-risk prognostic factors and morphological characteristics of retinoblastomas using digital whole slide images (WSI) generated by a scanner. METHODS: Forty-seven H&E sections of glass slides with high-risk morphological features of retinoblastoma were analyzed. Slides were scanned as WSI and reviewed. The results were compared with those obtained after reviewing the slides using a regular microscope as the gold standard. McNemar's test (MT), the percentage of agreement (POA), and sensitivity (S) and specificity (Sp) were evaluated between WSI and conventional microscopy. RESULTS: There were no differences with respect to multicentricity, growth type, rosette formation, choroidal invasion, anterior chamber invasion, extraocular extension, scleral extension, optic nerve invasion, necrosis, or Azzopardi effect between WSI analysis and light microscopy (MT, p = 1.0; POA = 100%; S = 100%, and Sp = 100%). Discordance was found in 1 case where calcification could not be found using WSI (MT, p = 1.00; POA = 97.9%; S = 100%, and Sp = 97.8%). CONCLUSION: To the best of our knowledge, this is the first report using digital pathology (WSI) to evaluate prognostic factors in eyes containing retinoblastomas. Using WSI, the pathologist was able to detect high-risk morphological features in retinoblastoma. To date, WSI is an important tool, in particular for ophthalmic pathologists examining enucleation and exenteration specimens.

Expression of focal adhesion kinase in uveal melanoma and the effects of Hsp90 inhibition by 17-AAG.

INTRODUCTION: Focal adhesion kinase (FAK) is implicated in tumor progression and metastatic cascade, and has been shown to be overexpressed in a variety of human cancers. However, the role of FAK in human uveal melanoma (UM) is not well defined. The purpose of this study was to evaluate the expression of FAK in UM tumors and normal eyes, and to determine the effect of Hsp90 inhibition on FAK expression in UM cells. METHODS: FAK expression was assessed in 39 UM specimens, FAK[pY397] expression was assessed in 51 UM specimens, and both FAK and FAK[pY397] expression were assessed in 20 normal eyes. The expression of FAK and FAK[pY397] was detected by Western blot in five UM cell lines after treatment with 10 µmol/L of 17-AAG. RESULTS: FAK was positive in 87.2% and FAK[pY397] in 90% of UM specimens. Low FAK expression was detected in non-tumor structures and in normal eyes. The cell lines with the most proliferative, invasive phenotype (92.1, SP6.5 and MKT-BR) displayed high expression of FAK[pY397], and the levels of FAK and FAK[pY397] were decreased in the presence of 17-AAG starting with 24 h of exposure. CONCLUSION: FAK and FAK[pY397] were overexpressed in human UM tumors compared to normal ocular tissue and high levels of FAK[pY397] were seen in the most aggressive UM cell lines. Hsp90 inhibition led to downregulation of FAK expression. We propose a role for FAK in the pathogenesis of UM. Future studies are needed to explore the use of Hsp90 inhibitors as a feasible approach for modulating FAK in UM.

Immunohistochemical detection of Hsp90 and Ki-67 in pterygium.

BACKGROUND: To examine the immunohistochemical expression of heat shock protein 90 (Hsp90) and Ki-67 protein in human pterygium. MATERIALS AND METHODS: Tissues obtained during pterygium surgery of 15 patients who underwent the bare-sclera procedure and 10 normal conjunctivae were studied. All of these pterygia were primary ones. Recurrent pterygia were excluded. Normal bulbar conjunctivas (2 x 2 mm) were obtained from the nasal region close to the limbus from patients during their cataract and retina surgeries. Immunohistochemical detection of Hsp90 and Ki67 was done using the streptavidin-biotin method in paraffin embedded tissue sections. RESULTS: The percentage of cells stained for Hsp90 was greater for pterygium epithelium (76 ± 10.8) than for normal conjunctiva (1.4 ± 0.8). In each pterygium sample more than 60% of cells were positive. The differences in positive cells between normal and pterygium epithelium were highly significant for Hsp90 (P < 0,001).Pterygium epithelium also showed a higher percentage of cells that stained for Ki67 (10.1 ± 9.5) than for normal conjunctiva (2.1 ± 1.9). The differences in positive cells were also statistically significant for Ki67 (P < 0.01). Although there were significant differences in the majority of samples observed. It was noted that in some samples there was no difference between normal and pterygium epithelium for Ki67. CONCLUSION: Our results indicate an abnormal expression of Hsp90 and ki-67 in pterygium samples when compared to normal conjunctiva.The finding of abnormal expression of levels of Hsp90 in pterygium samples can stimulate new research into pterygium and its recurrence. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1128478792898812.

Histopathological study of lesions of the caruncle: a 15-year single center review.

INTRODUCTION: The caruncle is a modified cutaneous tissue located at the inner canthus that contains hair follicles, accessory lacrimal glands, sweat glands and sebaceous glands. These different types of tissues can give rise to a wide variety of lesions that make the clinical diagnosis difficult. The aim of the study was to investigate the most common types of caruncle lesions and the clinical and pathological correlation. METHODS: Retrospective, observational case series. Records of caruncle lesions examined at the Henry C. Witelson Ocular Pathology Laboratory, McGill University, Montreal, Canada, between 1993 and 2008 were analyzed, comparing the clinical and histopathological findings. RESULTS: A total of 42 lesions from 42 patients were analyzed. Twenty-six (61.90%) of the patients were women and 16 (38.10%) were men and the age range from 20 to 84. The main diagnoses were: 16 epithelial lesions (38.09%), 14 inflammatory lesions (31.70%), 10 melanocytic lesions (21,95%), 2 lymphoid lesions (4.87%). From the 28 cases that had a preoperative clinical hypothesis only 17 presented a histopathological confirmation of the diagnosis (60.71%). CONCLUSION: The most common caruncle lesions were epithelial tumors followed by chronic inflammation and melanocytic lesions. Although most of the lesions were benign, there was a great number of misdiagnose based on the clinical suspicious.

SPARC is expressed in human uveal melanoma and its abrogation reduces tumor cell proliferation.

BACKGROUND: Secreted protein acidic and rich in cysteine (SPARC) has been shown to play an integral role in the progression of numerous malignancies. The aim of this study was to investigate the expression of SPARC in uveal melanoma (UM). MATERIALS AND METHODS: SPARC expression was assessed in UM cell lines using RT-PCR and immunocytochemistry. Small interfering RNA directed against SPARC was used to transfect each of the cell lines, which were subsequently run in proliferation assays. SPARC expression was further investigated in 19 cases of human UM and 11 primary and 8 metastatic tumors from a rabbit xenograft model. RESULTS: The cell lines transfected with SPARC siRNA showed a significant decrease in proliferation compared to controls. All cases of human uveal melanoma demonstrated positive staining for SPARC as did all primary and metastatic tumors from the xenograft model. CONCLUSION: SPARC may represent a novel target to inhibit growth of UM.

Nódulos calcificados subepidérmicos palpebrales / Subepidermal calcified nodules palpebral

La calcinosis cutis comprende cuatro formas de presentación: calcinosis cutis metastásica, calcinosis distrófica, calcinosis idiopática y nódulos calcificados subepidérmicos. La calcinosis cutis metastásica se desarrolla como resultado de hipercalcemia o hiperfosfatemia. En la calcinosis cutis distrófica, el calcio se deposita sobre un tejido previamente dañado. La calcinosis cutis idiopática es similar a la calcificación distrófica, pero se presenta sin evidencia de enfermedad subyacente. Los nódulos calcificados subepidérmicos, también conocidos como cálculos cutáneos se observan como lesiones pequeñas, elevadas y únicas. Se ven en niños o adultos jóvenes. El aspecto puede ser verrugoso o en casos tiene apariencia suave. La localización más común es en la cara. Se presentan cinco casos con diagnóstico de nódulos calcificados subepidérmicos, localizados en el párpado, diagnosticados en la Sección de Patología Ocular del Instituto Anatomopatológico "Dr. J. A. O’Daly"

Cutis calcinosis has four clinical presentations: metastatic calcinosis cutis, dystrophic calcinosis, idiopathic calcinosis and sub epidermal calcified nodules. Metastatic cutis calcinosis occurs as a result of hypercalcaemia or hyperphosphatemia. In dystrophic calcinosis cutis, calcium is deposited over a previously damaged tissue. Similar to dystrophic calcinosis, idiopathic calcinosis cutis is presented without any subjacent illness. Sub epidermal calcified nodules, also known as a cutaneous calculus are tiny, unique and elevated lesions. It is appear in children or young adults. The clinical aspect may be since soft lesions to verrugous appearance. The most common site to be seen is on the face. We present five eyelid sub epidermal calcified nodules diagnosed in Ocular Pathology Section at The Instituto Anatomopatologico "Dr. Jose A. O’Daly"

Expression of C-kit in retinoblastoma: a potential therapeutic target.

BACKGROUND: C-kit is a transmembrane tyrosine kinase protein thought to play an important role in tumourigenesis. With the development of the compound imatinib mesylate, which specifically inhibits tyrosine kinase receptors, C-kit has emerged as a potential therapeutic target. This study aims to determine the immunoexpression of C-kit in retinoblastoma and correlate this expression with histopathological prognostic features. METHODS: Eighty-four paraffin-embedded retinoblastomas were collected from the Henry C Witelson Ocular Pathology Registry. C-kit immunostaining was used according to the protocol provided by Ventana Medical System Inc., Arizona. Immunoreactivity was correlated with the presence or absence of invasion into the choroid and optic nerve and the degree of tumour differentiation. Odds ratios were calculated to quantify differences in C-kit expression between tumours with different patterns of invasion and differentiation. RESULTS: Twenty-one slides (25%) were excluded from analysis because of the presence of extensive tissue necrosis or the absence of sufficient optic nerve tissue for analysis. Overall, C-kit expression was identified in 33/63 specimens analysed (52.38%). Two of the 13 tumours without choroidal or optic nerve invasion (15.4%) were positive for C-kit. C-kit expression was seen in 31 of the 50 tumours with extraretinal invasion (62%, p<0.01), 26 of 44 specimens with choroidal involvement (59.9%, p<0.2), and 20 of the 29 with optic nerve involvement (68.96%, p<0.02). Fourteen of 25 moderate or well-differentiated specimens (56%) and 19 of 38 undifferentiated specimens (50%) displayed positivity for C-kit (p>0.5). CONCLUSIONS: More than half the retinoblastomas in this study expressed C-kit. The expression of C-kit strongly correlated with histopathological features of a worse prognosis including optic nerve and choroidal invasion.

Tumores óseos orbitarios / Orbital bone tumors

Reportar la frecuencia de los tumores óseos orbitarios. En un período de ocho años (1998-2005) se realizaron 117 899 biopsias en el Instituto Anatomopatológico "Dr. J. A. O’Daly" de la Universidad Central de Venezuela, de las cuales se identificaron 1 826 biopsias óseas, de las cuales 17 (0,93 por ciento) correspondieron a tumores óseos orbitarios. Se investigaron las variables edad, sexo, localización y tipo de neoplasia. En la revisión se identificaron 10 tumores óseos primarios (8 benignos y 2 malignos), 6 casos de pseudotumores (35 por ciento) y un tumor metastásico (6 por ciento). El osteoma y la displasia fibrosa fueron las condiciones más frecuentes. Los tumores óseos primarios de la órbita son un grupo heterogéneo de condiciones que constituyen el 0,6 por ciento - 2 por ciento de todos los tumores orbitarios.

To evaluate the rate of orbital bone tumours. Our experience over a 8-year period in Instituto Anatomopatologico "Dr. J. A. O’Daly" at the Universidad Central de Venezuela, yielded 17 (0.93 percent) cases of orbital bone tumours from a total of 117 899 biopsies. Data were collected on patient age, sex, tumour localization and type of neoplastic disorders. Results: Ten cases (59 percent) were primary tumours of orbital bone (8 benign and 2 malignant), six patients with tumour-like lesions (35 percent) and one bone metastases. The two most commonly encountered entities were osteoma and fibrous dysplasia. Primary tumours of orbital bone constitute 0,6 percent - 2 percent of all orbital tumours.

Malignización de retinocitoma en retinoblastoma en una preescolar con retinoblastoma bilateral: reporte de un caso / Retinocytoma maligninant in retinoblastoma in a preschool bilateral retinoblastoma: case report

El retinoblastoma es el tumor intraocular maligno primario más frecuente en la infancia. No obstante, es un tumor raro, suponiendo alrededor del 3 por ciento de todos los cánceres infantiles. El retinocitoma es considerado por muchos autores una variedad benigna del retinoblastoma. Presentamos el caso clínico de un preescolar femenino quien presentó leucocoria bilateral y estrabismo. Se planteó el diagnóstico clínico de retinoblastoma bilateral. En vista del tamaño del tumor y de haber descartado la invasión extraocular, se realizó enucleación de ambos globos oculares, aplicando 3 ciclos de quimioterapia entre ambas cirugías. El estudio histopatológico reportó, para el globo ocular derecho, retinoblastoma poco diferenciado con probable origen en un retinocitoma, según la clasificación de Howarth. La paciente falleció 4 meses después.

Patología ocular para el patólogo general / Ocular pathology for the general pathologist

La Patología Ocular incluye el estudio de los procesos o enfermedades que afectan el globo ocular (las estructuras que lo componen) y sus anexos. Esta especialidad ha avanzado en el tiempo en la misma proporción que lo han hecho la Patología General y la Oftalmología, de las cuales se nutren. El objetivo de la presente revisión es la actualización de términos y procedimientos de utilidad para los patólogos generales, que reciben muestras relacionadas con el campo de la Oftalmología. Los especialistas en Oftalmología y sus pacientes se beneficiarán de un óptimo estudio y, con el consiguiente reporte final se accederá a un tratamiento adecuado. Se revisan así mismo, los tópicos más resaltantes y comunes en la práctica diaria de la Patología Ocular.

Distrofina como herramienta diagnóstica / Dystrophin how implement diagnóstic

Las distrofias musculares constituyen un amplio grupo de enfermedades con una forma de presentación y un curso clínico variable; entre ellas, las distrofinopatías, con herencia ligada al cromosoma X como la distrofia muscular tipo Duchenne y tipo Becker son entidades poco frecuentes. Se estudiaron las biopsias de 5 pacientes pedíatricos con diagnóstico de distrofia muscular, con microscopia convencional, histoquímica enzimática e inmunorreacciones para distrofina. En tres casos se diagnosticó distrofia muscular tipo Duchenne y en dos casos, distrofia muscular tipo Becker. A través de esta presentación se pone en relieve la importancia de la realización de inmunorreacción para distrofina en el diagnóstico de estas entidades, del estudio del grupo familiar, de proporcionar consejo genético y de diferenciarlas de otras miopatías

Fusión espleno-gonadal continua: reporte de un caso / Continuous splenic-gonadal fusion: case report

La fusión espleno-gonadal es una anomalia congénita rara. Es el resultado de la fusión de los esbozos esplénicos y gonadal durante el desarrollo embriológico. Se han descrito aproximadamente 90 casos en la literatura

Patología de la ampolla de vater estudio clínico-morfológico / Pathologic study morphologic-clinic of the ampulla of vater

Se examinaron en un estudio retrospectivo (1993-1997) 29 casos de biopsias endoscópicas de la ampolla de Vater y 5 especímenes de pancreatoduodenectomía (operación de Whipple) del Instituto Anatomopatológico "Dr. J.A. O'Daly". En todos los casos se evaluó la información clínica disponible y las evaluaciones complementarias. Las biopsias se clasificaron según el diagnóstico histológico en lesiones inflamatorias, tumores benignos y malignos. Los tumores malignos fueron los más frecuentes (40 por ciento) y entre éstos los adenocarcinomas bien diferenciados de tipo intestinal. Entre los tumores benignos se identificaron pólipos tubulares, vellosos y túbulo-vellosos y las lesiones inflamatorias correspondieron a inflamación crónica con y sin regudización. Los cambios displásicos y el carcinoma in situ se vieron en dos casos. No se observó asociación del carcinoma ampular a residuos adenomatosos. Así como tampoco se evidenció relación del cáncer de la ampolla de Vater a poliposis colónica familiar. No se pudo evaluar la eficacia del tratamiento propuesto, ni determinar la sobrevida en esta población debido a la falta de seguimiento de los casos