Overexpression of CLDN16 in ovarian cancer is modulated by PI3K and PKC pathways.

Epithelial ovarian cancer (EOC) is the gynecological malignant tumor of poorest prognosis and higher mortality rate. Chemotherapy is the base of high-grade serous ovarian cancer (HGSOC) treatment; however, it favors the emergence of chemoresistance and metastasis. Thus, there is an urge to search for new therapeutic targets, such as proteins related to cellular proliferation and invasion. Herein, we investigated the expression profile of claudin-16 (CLDN16 protein and CLDN16 transcript) and its possible functions in EOC. In silico analysis of CLDN16 expression profile was performed using data extracted from GENT2 and GEPIA2 platforms. A retrospective study was carried out with 55 patients to evaluate the expression of CLDN16. The samples were evaluated by immunohistochemistry, immunofluorescence, qRT-PCR, molecular docking, sequencing, and immunoblotting assays. Statistical analyzes were performed using Kaplan-Meier curves, one-way ANOVA, Turkey posttest. Data were analyzed using GraphPad Prism 8.0. In silico experiments showed that CLDN16 is overexpressed in EOC. 80.0% of all EOC types overexpressed CLDN16, of which in 87% of the cases the protein is restricted to cellular cytoplasm. CLDN16 expression was not related to tumor stage, tumor cells differentiation status, tumor responsiveness to cisplatin, or patients' survival rate. When compared to data obtained from in silico analysis regarding EOC stage and degree of differentiation, differences were found in the former but not in the later, neither in survival curves. CLDN16 expression in HGSOC OVCAR-3 cells increased by 1.95-fold (p < 0.001), 2.32-fold (p < 0.001), and 6.57-fold (p < 0.001) via PKC, PI3K, and estrogen pathways, respectively. Altogether, our results suggest that despite the low number of samples included in our in vitro studies, adding to the expression profile findings, we provided a comprehensive study of CLDN16 expression in EOC. Therefore, we hypothesize that CLDN16 is a potential target in the diagnosis and treatment of the disease.

Estrogen receptor alpha (ERS1) SNPs c454-397T>C (PvuII) and c454-351A>G (XbaI) are risk biomarkers for breast cancer development.

There are several risk factors related to Breast Cancer (BC) risks and response to chemotherapy with SERMs. Recently some single nucleotide polymorphisms (SNPs) on ESR1 gene have been associated to this disease. However, data are still inconclusive. The present study aimed to investigate the association of SNPs c454-397T>C (also called PvuII) and c454-351A>G (so called XbaI) to incidence of sporadic BC; ERα expression in BC; tamoxifen hormonetherapy (HT-TMX) responsiveness. To do so, a cohort of BC patients was analyzed through retrospective data collection, immunohistochemistry to ERα protein, and genotyping for PvuII and XbaI SNPs by PCR-RFLP, confirmed by sequencing. Significant difference in PvuII alleles frequencies were found BC patients when compared to control samples. Patients with P allele have a 5.14-fold increased BC risk. We found higher P and X alleles frequencies in ERα positive BC and the pp and xx genotypes were observed exclusively in patients with HT-TMX-responsive BC. Taken together, data indicates that P allele as a novel sporadic BC biomarker whereas p and x alleles enhanced chemotherapy responsiveness.

Evaluation of the progesterone receptor status in breast cancer using three different antibodies: a comparison by Allred score system.

Breast cancer (BC) hormonal receptors status is assessed by immunohistochemistry (IHC), a specific, sensitive, and accessible method that guide breast cancer treatment. In this study, we evaluated progesterone receptor (PR) expression in 53 BC cases using 3 anti-PgR antibodies (AB): monoclonal (SP42 and PgR636) and polyclonal ab62621. Primary BC cases (with signed informed consent) were used to generate tissue microarray platforms, where PR expression was accessed by IHC and evaluated by the Allred score. Categorical and quantitative data are shown in percentage and mean, respectively. Concordance (CON) and correlation among ABs were analyzed by Kappa factor (Κ), Spearman's correlation coefficient (ρ) or intraclass correlation coefficient. Staining patterns of each AB were compared by paired T-Test. We noted poor CON and Κ between ab62621 vs SP42 (CON=64.1%; Κ=0.247), and ab62621 vs PgR636 (CON=62.3%; Κ=0.204), but higher CON between SP42 vs PgR636 (CON 90.6%; Κ=0.738). Data were corroborated by Mc Nemar statistical test (p=0.019, p=0.014 and p>0.05, respectively). Regarding staining intensity (SI) among PgR+ samples, we found higher proportion of weak staining and lower SI for ab62621 (48.3%; mean IS=1.6), when compared to SP42 (20.0%, mean IS=2.1, T-test p<0.01) and PgR636 (2.3, 21.9%, T-test p<0.01). Within the entire sample, similar results were observed following ρ: SP42 vs PgR636 (ρ=0.8103); ab62621 vs SP42 (ρ=0.3524); ab62621 vs PgR636 (ρ=0.4075). As for proportion of stained cells and proportion score (PS), among PgR+ samples, the mean values for ab62621 (75.4%; 4.8) were significantly higher than those of SP42 (56.3%, 4.3; T-test p<0.01) and RPG636 (60.1%; 4.2; T-test p<0.01). Similar data were found after analyzing PS for the entire sample: SP42 vs PgR636 (ρ=0.8588); SP42 vs ab62621 (ρ=0.4832); RPG636 vs ab62621 (ρ=0.4050). Our data indicate that anti-PgR monoclonal ABs, PgR636 and SP42, are, unlike ab62621, equally suitable to test BC PgR status by IHC due to their higher accuracy. Therefore, we suggest their clinical use during BC diagnosis; thus, enabling more precise therapeutic decisions to treat BC.

Comparison of immunohistochemical analysis with estrogen receptor SP1 and 1D5 monoclonal antibodies in breast cancer.

In the present study, we aimed to evaluate estrogen receptor ER-alpha status in 61 breast cancer cases using Sp1 and 1D5 monoclonal antibodies. Tissue array platforms were generated containing samples of breast cancer and positive controls that were assayed by immunohistochemistry applying monoclonal primary antibodies anti-ER alpha, SP1 and 1D5. We noted a high concordance rate (96.7%) between the referred antibodies. Moreover, we calculated the Kappa factor (0.921), indicating that 1D5 and SP1 provided overlapping ERα expression results. Indeed, we observed controversial results only in 2 samples studied, which were ER-negative when stained with 1D5 and ER-positive when assessed with SP1. Total concordance of PS was obtained (Pearson and intraclass CF, 0.7351 and 0.6193, respectively). However, concordance between the antibodies seems to be more accurate in higher PS values. An excellent IS correlation between antibodies was observed throughout the population (Spearman's CF, ρ=0.9150). Following the Allred score, 17 out of 42 positive BC samples diverged, with 1D5 always pointing to weaker staining than SP1. When calculating Spearman's CF of Total Score (TS) within the population, an excellent correlation between both the antibodies (ρ=0.9238) was noted. Nonetheless, the results were less concordant among the BC-positive cases (ρ=0.7743). Indeed, 20 samples were differentially classified using the antibodies (only 3 had higher TS with 1D5). Considering the mean TS of all samples or of invasive ductal carcinoma, SP1 provided higher scores than 1D5 (p<0.05). We recommend the use of the anti-ER RMAb SP1 due to the high probability that the BC ERα status can be determined accurately as the reagent provides higher IS. Therefore, the A-score was higher than the MMAb 1D5. Ultimately, higher IS and A-score decrease the possibility of ERα status misinterpretation and, consequently, inappropriate BC treatment that would compromise the patient's quality of life and overall survival. We recommend the use of anti-ER RMAb SP1 due to the high probability that the BC ER status can be determined accurately as the reagent provides higher IS, therefore higher A-score, than the MMAb 1D5.

Comparison of DGGE and immunohistochemistry in the detection of TP53 variants in a Brazilian sample of sporadic breast tumors.

The presence of TP53 gene mutations in breast cancer has been associated with worse prognosis. These mutations interfere with the ability of the p53 protein, a transcription factor, to regulate the expression of target genes. Unlike the wild-type protein, which is rapidly degraded in cells, mutated forms have increased half-life and accumulate in tumor cells. Immunohistochemistry (IHC) is widely used in Brazil in the determination of breast cancer patients' prognosis. However, this technique is not able to detect many altered forms of the p53 protein (false-negative results) and readily detects the accumulation of wild-type p53 (false-positive results) that is associated with non-tumoral processes. For these reasons, we have set out to compare the efficiency of IHC with a molecular technique that detects gene variations at the DNA level in the evaluation of Brazilian patients with sporadic breast cancer. We have used denaturing gradient gel electrophoresis (DGGE) to study the TP53 status in 45 tumors, finding 26 allelic variants, most of them located in exon 4. Comparing the two techniques, IHC showed a false-negative rate of 64% and a false-positive rate of 50%. These results confirm the inability of IHC to correctly detect TP53 status, reason because it should not be routinely used to establish prognosis of breast cancer patients in Brazilian Pathology Laboratories. We recommend the utilization of a screening method, such as DGGE, followed by sequencing of altered exonic fragments to correctly detect TP53 gene variants and establish the prognosis of breast cancer patients.

Lesão pigmentada do mamilo e aréola - doença de Paget pigmentada ou melanoma? / Pigmented lesion of the nipple and areola - pigmented Paget's disease or melanoma?

O câncer de mama é a neoplasia mais frenquente na mulher. A pele da mama também pode ser acometida por câncer, como carcinoma mamário em estádios avançados, doença de Paget e raramente melanoma. Relata-se o caso de paciente de 43 anos que procurou o serviço com história de mancha cutânea na mama direita há mais de um ano. Havia tratado com antimicóticos e corticóides tópicos sem melhora. Ao exame clínico apresentava mácula na região central da mama direita, irregular, de coloração avermelhada e amarronzada, associada à área palpável mal delimitada na mesma mama. As principais suspeitas diagnósticas eram melanoma cutâneo e doença de Paget atípica. Tinha mamografia de um ano antes sem achados suspeitos e biópsia da pele sugerindo Paget ou melanoma e indicando estudo imunohistoquímico para definir diagnóstico. Foi solicitada revisão anatomopatológica e nova mamografia. À mamografia e á ultra-sonografia complementar foi visto nódulo espiculado correspondente à área palpável. Foi feita biópsia que diagnosticou carcinoma ductal infiltrante. A revisão da biópsia da pele, associada ao estudo imunohistoquímico (IMH), mostrou tratar-se de doença de Paget pigmentada da mama. Conforme a literatura, a doença de Paget pigmentada mamária é rara e tem como principal diagnóstico diferencial o melanoma in situ. Isso alerta para avaliação minuciosa clínico-patológica diante de lesões pigmentadas da pele mamária, sendo imprescindível o estudo IMH.

Breast cancer is the most common cancer in women. The skin of the breast can also be affected by cancer, as for breast cancer in advanced stages, Paget’s disease and rarely melanoma. We report the case of 43 years old patient who looked for this service with a history of skin spot in the right breast for over one year. It had been treated with antifungal and topical steroids without any successful improvement. In a clinical examination, an irregular red and brown macula was noticed in the central region of right breast, coupled with badly palpable area in the same breast. The main suspected diagnosis were skin melanoma and Paget’s atypical disease. There was a mammography from one year earlier without any suspicious findings and a biopsy of the skin suggesting Paget or melanoma and also indicating immunohistochemical studies to define diagnosis. An anatomopathological review and a new mammography were requested. An angled nodule corresponding to palpable was noticed in the additional mammography and ultrasound. An infiltrating ductal carcinoma was diagnosed through a biopsy. The review of the skin biopsy, together with the immunohistochemical study, confirmed the existence of pigmented Paget’s disease in the breast. According to the bibliography, the pigmented mammary Paget’s disease is rate and it has as its main diagnosis differential the melanoma in situ. It warns for a thorough clinical and pathological evaluation whenever pigmented lesions are perceived in the breast skin, prompting to an indispensable immunohistochemical study.

Cicatriz radial/lesão esclerosante complexa: aspectos radiológicos com correlação clínica, ultra-sonográfica e anatomopatológica / Radial scar/complex sclerosing lesion: radiologic features with clinical, ultrasonographic and pathologic correlation

OBJETIVO: Estudar, retrospectivamente, as várias formas de apresentação da cicatriz radial/lesão esclerosante complexa (CR/LEC) na mamografia, correlacionando-as com o exame clínico e os achados ultra-sonográficos. Os achados histopatológicos e a associação da CR/LEC com hiperplasia atípica e carcinoma são discutidos. MATERIAIS E MÉTODOS: Foi realizado estudo retrospectivo de 926 lesões impalpáveis em 901 pacientes submetidas a biópsia excisional após localização pré-cirúrgica, do arquivo do Centro de Diagnóstico por Imagem e do Hospital Santa Rita, Vitória, ES, no período de outubro de 1993 a dezembro de 2001, nas quais 57 pacientes tiveram diagnóstico histopatológico de CR/LEC. RESULTADOS: A idade variou de 31 a 84 anos (média de 49 anos). As lesões foram palpáveis em dez casos. Na mamografia, 48 casos se apresentaram como distorção arquitetural, e com a mesma freqüência o nódulo espiculado e a densidade assimétrica, quatro casos cada. As microcalcificações foram detectadas na mamografia em 14 casos e em 20 quando o espécime cirúrgico foi avaliado. A ultra-sonografia foi realizada em 51 casos, tendo expressão em 17 como área hipoecóica irregular com atenuação acústica posterior. Houve 42 casos de CR/LEC sem ou com proliferação típica, nove casos com proliferação epitelial atípica e seis casos com carcinoma infiltrativo associado. CONCLUSÃO: Não é possível fazer diagnóstico diferencial de CR/LEC com câncer pelos métodos de imagem e a biópsia excisional deve ser realizada

OBJECTIVE: To review the different types of radial scar/complex sclerosing lesion (RS/CSL) seen on clinical, mammography, and ultrasound examinations. The histopathology findings and the association of RS/CSL with atypical hyperplasia and malignancy are discussed. MATERIALS AND METHODS: We performed a retrospective study of patients from the files of the "Centro de Diagnóstico por Imagem and Hospital Santa Rita" - Vitória, ES, Brazil, of the period between October, 1993 and December, 2001. A total of 926 non-palpable lesions of 901 patients submitted to excision biopsy after pre-surgical localization were studied. Fifty-seven patients had pathology proven RS/CSL diagnosis. RESULTS: The age of the patients ranged from 31 to 84 years (average 49 years). There were palpable lesions in 10 patients (17.9%). The mammographic signs that suggested the need of biopsy were divided into three groups: architectural distortion (48 patients; 85.7%), stellate nodules (4 patients; 7.1%), and asymmetric densities (4 patients; 7.1%). Microcalcifications were detected by mammography alone in 14 patients (25%) and in the surgical specimen in 20 (35.7%) patients. Among 51 patients submitted to ultrasound, 17 had irregular hypoechoic areas with posterior acoustic shadowing. Of the 57 patients with RS/CSL, 42 had ductal hyperplasia, 9 had atypical ductal proliferation and 6 had infiltrative carcinoma. CONCLUSION: Differential diagnosis of RS/CSL from carcinomas can not be made using imaging methods and excision biopsy is mandatory

Esquistossomose mansônica: forma tumoral cerebral: relato de caso / Schistosomiasis mansoni: cerebral tumor form: case report

Relatamos um caso de esquistossomose mansônica no Sistema Nervoso Central, apresentando-se sob a forma tumoral. O paciente, de 16 anos, apresentou história de cefaléia há 2 meses, acompanhada de vômitos, tonteira e lipotímia, 15 dias antes da internaçao. O exame físico foi normal. A tomografia axial computadorizada do crânio revelou área hipodensa córtico-subcortical na regiao parietal direita, com efeito de massa sobre ventrículo lateral homolateral, hipercaptante à injeçao de contraste e de aspecto nodular. Feita ressecçao cirúrgica, o exame histopatológico evidenciou tecido encefálico com denso infiltrado crônico, granulomatose multifocal e áreas de necrose circundando ovos de Schistossoma mansoni. A biópsia retal identificou ovos viáveis de S. mansoni. Foi feito tratamento com praziquantel 60 mg/kg/dia durante 7 dias e dexametasona 12 mg/dia. Chamamos a atençao para esta forma rara de neuroesquistossomose, com manifestaçao tumoral de efeito de massa e reaçoes sorológicas negativas, inclusive no liquor. O diagnóstico somente foi estabelecido através da intervençao cirúrgica.