Molecular genetic analysis of primary renal epithelial tumours with granular oncocytic cytoplasms / Análisis genético molecular de tumores epiteliales renales primarios con citoplasma oncocítico granular

SUMMARY: The group of primary renal tumours with granular-oncocytic cytoplasm is a very heterogeneous group, in its histological origin and biological behavior resulting in many diagnostic problems. In this study 57 renal epithelial tumours with granular oncocytic cells were analyzed using fluorescence in situ hybridisation (FISH), array comparative genomic hybridisation (aCGH) and polymerase chain reaction (PCR). The results of analysis in renal oncocytoma (RO) did not indicate the presence of the gene mutations or chromosomal abnormalities. Sporadic renal hybrid oncocytic/chromophobe tumours (HOCT) had multiple numerical aberrations of chromosomes 1, 2, 6, 9, 10, 13, 17, 20, 21 and 22. This type of tumour had no mutations in the VHL, c-kit, PDGFRA, and FLCN genes. Oncocytic papillary renal cell carcinoma (O-PRCC) had numerical abnormalities of chromosomes 7 and 17 and the loss of the Y chromosome. Cytogenetic analysis of 20 pigmented microcystic chromophobe renal cell carcinomas (PMChRCC) showed monosomy as the most frequent aberration in all analyzed chromosomes 1, 2, 5, 10, 13, 17 and 21. One case of chromophobe renal cell carcinoma (ChRCC) with hyaline globules had a mutation in the distal part of exon 3 of the VHL gene. Absence of genetic disorders in usual RO is common result, but we have established absence of genetic disorders even in rare variants. Variety of genetic alterations detected in sporadic renal HOCT proves it to be a separate entity, not a variant of ChRCC, while PMChRCC is an uncommon variant of ChRCC. O-PRCC is a subtype of papillary renal cell carcinoma.

RESUMEN: El grupo de tumores renales primarios con citoplasma granular-oncocítico es un grupo muy heterogéneo, en su origen histológico y comportamiento biológico, resultando en problemas de diagnóstico. En el estudio se analizaron 57 tumores epiteliales renales con citoplasma oncocítico granular mediante hibridación fluorescente in situ (FISH), hibridación genómica comparativa de matriz (aCGH) y reacción en cadena de la polimerasa (PCR). Los resultados del análisis en oncocitoma renal (RO) no indicaron la presencia de mutaciones genéticas ni anomalías cromosómicas. Los tumores oncocíticos / cromófobos híbridos renales esporádicos (HOCT) tenían múltiples aberraciones numéricas de los cromosomas 1, 2, 6, 9, 10, 13, 17, 20, 21 y 22. No se observaron mutaciones en este tipo de tumor en el VHL, c-kit, PDGFRA y genes FLCN. El carcinoma de células renales papilar oncocítico (O-PRCC) tenía anomalías numéricas de los cromosomas 7 y 17 y la pérdida del cromosoma Y. El análisis citogenético de 20 carcinomas de células renales cromófobos microquísticos pigmentados (PMChRCC) mostró que la monosomía era la aberración más frecuente en todos los cromosomas analizados 1, 2, 5, 10, 13, 17 y 21. Un caso de carcinoma de células renales cromófobo (CCRc) hialino tenía una mutación en la parte distal del exón 3 del gen VHL. La ausencia de trastornos genéticos en la OI habitual es un resultado común, pero hemos establecido la ausencia de trastornos genéticos incluso en variantes raras. Varias alteraciones genéticas detectadas en esporádica HOCT renal demuestran que es una entidad separada, no una variante de ChRCC, mientras que PMChRCC es una variante poco común de ChRCC. O-PRCC es un subtipo de carcinoma papilar de células renales.

The NER-related gene GTF2H5 predicts survival in high-grade serous ovarian cancer patients / 부인종양

OBJECTIVE: We aimed to evaluate the prognostic and predictive value of the nucleotide excision repair-related gene GTF2H5, which is localized at the 6q24.2-26 deletion previously reported by our group to predict longer survival of high-grade serous ovarian cancer patients. METHODS: In order to test if protein levels of GTF2H5 are associated with patients' outcome, we performed GTF2H5 immunohistochemical staining in 139 high-grade serous ovarian carcinomas included in tissue microarrays. Upon stratification of cases into high- and low-GTF2H5 staining categories (> and < or = median staining, respectively) Kaplan-Meier and log-rank test were used to estimate patients' survival and assess statistical differences. We also evaluated the association of GTF2H5 with survival at the transcriptional level by using the on-line Kaplan-Meier plotter tool, which includes gene expression and survival data of 855 high-grade serous ovarian cancer patients from 13 different datasets. Finally, we determined whether stable short hairpin RNA-mediated GTF2H5 downregulation modulates cisplatin sensitivity in the SKOV3 and COV504 cell lines by using cytotoxicity assays. RESULTS: Low expression of GTF2H5 was associated with longer 5-year survival of patients at the protein (hazard ratio [HR], 0.52; 95% CI, 0.29 to 0.93; p=0.024) and transcriptional level (HR, 0.80; 95% CI, 0.65 to 0.97; p=0.023) in high-grade serous ovarian cancer patients. We confirmed the association with 5-year overall survival (HR, 0.55; 95% CI, 0.38 to 0.78; p=0.0007) and also found an association with progression-free survival (HR, 0.72; 95% CI, 0.54 to 0.96; p=0.026) in a homogenous group of 388 high-stage (stages III-IV using the International Federation of Gynecology and Obstetrics staging system), optimally debulked high-grade serous ovarian cancer patients. GTF2H5-silencing induced a decrease of the half maximal inhibitory concentration upon cisplatin treatment in GTF2H5-silenced ovarian cancer cells. CONCLUSION: Low levels of GTF2H5 are associated with enhanced prognosis in high-grade serous ovarian cancer patients and may contribute to cisplatin sensitization.

Somatic mutations in breast and serous ovarian cancer young patients: a systematic review and meta-analysis / Mutações somáticas em pacientes jovens com câncer de mama e epitelial de ovário: revisão e metanálise

Summary Objective: our aim was to evaluate whether somatic mutations in five genes were associated with an early age at presentation of breast cancer (BC) or serous ovarian cancer (SOC). Methods: COSMIC database was searched for the five most frequent somatic mutations in BC and SOC. A systematic review of PubMed was performed. Young age for BC and SOC patients was set at ≤35 and ≤40 years, respectively. Age groups were also classified in <30years and every 10 years thereafter. Results: twenty six (1,980 patients, 111 younger) and 16 studies (598, 41 younger), were analyzed for BC and SOC, respectively. In BC, PIK3CA wild type tumor was associated with early onset, not confirmed in binary regression with estrogen receptor (ER) status. In HER2-negative tumors, there was increased frequency of PIK3CA somatic mutation in older age groups; in ER-positive tumors, there was a trend towards an increased frequency of PIK3CA somatic mutation in older age groups. TP53 somatic mutation was described in 20% of tumors from both younger and older patients; PTEN, CDH1 and GATA3 somatic mutation was investigated only in 16 patients and PTEN mutation was detected in one of them. In SOC, TP53 somatic mutation was rather common, detected in more than 50% of tumors, however, more frequently in older patients. Conclusion: frequency of somatic mutations in specific genes was not associated with early-onset breast cancer. Although very common in patients with serous ovarian cancer diagnosed at all ages, TP53 mutation was more frequently detected in older women.

Resumo Objetivo: avaliar se mutações somáticas em câncer de mama e seroso de ovário estão associados com pacientes jovens. Métodos: com base no COSMIC, foram selecionados os cinco genes mais frequentes mutados em câncer de mama e seroso de ovário. Em seguida, realizou-se uma revisão sistemática no PubMed. Pacientes jovens foram classificadas com ≤35 anos e ≤40 anos para câncer de mama e seroso de ovário, respectivamente. Classificaram-se também as pacientes em grupos etários de ≤30 anos, separados a cada 10 anos. Resultados: vinte e seis (1.980 pacientes, 111 jovens) e 16 estudos (598, 41 jovens) foram selecionados para câncer de mama e seroso de ovário, respectivamente. Em câncer de mama, pacientes jovens apresentaram baixa frequência de mutações somáticas em PIK3CA. Tumor HER2 negativo foi associado a mutações somáticas em PIK3CA no grupo etário mais avançado, e em tumores ER positivos foi observada uma tendência a essa associação. Mutações somáticas em TP53 foram observadas em 20% dos tumores, em ambos os grupos (≤35 anos vs. ≥35 anos). Mutações somáticas em PTEN, CDH1 e GATA3 foram analisadas em 16 pacientes e apenas uma apresentou mutação em PTEN. Em câncer seroso de ovário, mutações somáticas em TP53 foram detectadas em mais que 50% dos tumores; entretanto, foram mais frequentes em pacientes idosas. Conclusão: a frequência de mutações somáticas nos genes selecionados não foi associada com pacientes jovens. Embora muito comum em pacientes com câncer seroso de ovário, mutações somáticas em TP53 foram mais frequentes em pacientes mais velhas.

Gene expression profile of ABC transporters and cytotoxic effect of ibuprofen and acetaminophen in an epithelial ovarian cancer cell line in vitro / Perfil da expressão gênica dos transportadores ABC e efeito citotóxico do ibuprofeno e acetaminofen em uma linhagem celular de câncer epitelial de ovário in vitro

PURPOSES: To determine the basic expression of ABC transporters in an epithelial ovarian cancer cell line, and to investigate whether low concentrations of acetaminophen and ibuprofen inhibited the growth of this cell line in vitro. METHODS: TOV-21 G cells were exposed to different concentrations of acetaminophen (1.5 to 15 μg/mL) and ibuprofen (2.0 to 20 μg/mL) for 24 to 48 hours. The cellular growth was assessed using a cell viability assay. Cellular morphology was determined by fluorescence microscopy. The gene expression profile of ABC transporters was determined by assessing a panel including 42 genes of the ABC transporter superfamily. RESULTS: We observed a significant decrease in TOV-21 G cell growth after exposure to 15 μg/mL of acetaminophen for 24 (p=0.02) and 48 hours (p=0.01), or to 20 μg/mL of ibuprofen for 48 hours (p=0.04). Assessing the morphology of TOV-21 G cells did not reveal evidence of extensive apoptosis. TOV-21 G cells had a reduced expression of the genes ABCA1, ABCC3, ABCC4, ABCD3, ABCD4 and ABCE1 within the ABC transporter superfamily. CONCLUSIONS: This study provides in vitro evidence of inhibitory effects of growth in therapeutic concentrations of acetaminophen and ibuprofen on TOV-21 G cells. Additionally, TOV-21 G cells presented a reduced expression of the ABCA1, ABCC3, ABCC4, ABCD3, ABCD4 and ABCE1 transporters. .

OBJETIVOS: Determinar a expressão básica dos transportadores ABC em uma linhagem celular do câncer epitelial de ovário, e investigar se o acetaminofen e o ibuprofeno em baixas concentrações são capazes de inibir o crescimento desta linhagem celular in vitro. MÉTODOS: A linhagem celular TOV-21 G foi exposta a diferentes concentrações de acetaminofen (1,5 a 15 µg/mL) e ibuprofeno (2,0 a 20 µg/mL), de 24 a 48 horas. O crescimento celular foi avaliado utilizando-se um ensaio de viabilidade celular. A morfologia celular foi determinada por meio da microscopia de fluorescência. O perfil de expressão gênica foi estabelecido por um painel de 42 genes da superfamília de transportadores ABC. RESULTADOS: Observou-se um decréscimo significativo no crescimento das células TOV-21 G expostas a 15 µg/mL de acetaminofen durante 24 (p=0,02) e 48 horas (p=0,01), ou a 20 µg/mL de ibuprofeno por 48 horas (p=0,04). Ao avaliar a morfologia das células cultivadas, não foi observada evidência de apoptose extensiva. A linhagem de células estudada subexpressa os genes de ABCA1, ABCC3, ABCC4, ABCD3, ABCD4 e ABCE1 na superfamília de transportadores ABC. CONCLUSÕES: Este estudo fornece evidências in vitro referentes aos efeitos inibidores do crescimento de concentrações terapêuticas do acetaminofen e ibuprofeno na linhagem celular testada. Além disso, as células TOV-21 G apresentaram uma expressão reduzida de genes dos transportadores ABCA1, ABCC3, ABCC4, ABCD3, ABCD4 e ABCE1. .

DNA Hypomethylation-Mediated Overexpression of Carbonic Anhydrase 9 Induces an Aggressive Phenotype in Ovarian Cancer Cells

PURPOSE: Both genetic and epigenetic alterations can lead to abnormal expression of metastasis-regulating genes in tumor cells. Recent studies suggest that aberrant epigenetic alterations, followed by differential gene expression, leads to an aggressive cancer cell phenotype. We examined epigenetically regulated genes that are involved in ovarian cancer metastasis. MATERIALS AND METHODS: We developed SK-OV-3 human ovarian carcinoma cell xenografts in mice. We compared the mRNA expression and DNA methylation profiles of metastatic tissues to those of the original SK-OV-3 cell line. RESULTS: Metastatic implants showed increased mRNA expression of the carbonic anhydrase 9 (CA9) gene and hypomethylation at CpG sites in the CA9 promoter. Treatment of wild-type SK-OV-3 cells with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine reduced methylation of the CA9 promoter and increased CA9 mRNA expression. Eight CpGs, which were located at positions -197, -74, -19, -6, +4, +13, +40, and +86, relative to the transcription start site, were hypomethylated in metastatic tumor implants, compared to that of wild-type SK-OV-3. Overexpression of CA9 induced an aggressive phenotype, including increased invasiveness and migration, in SK-OV-3 cells. CONCLUSION: Alterations in the DNA methylation profile of the CA9 promoter were correlated with a more aggressive phenotype in ovarian cancer cells.