Evaluation of BubR1 expression in oral squamous cell carcinomas and benign oral lesions associated with human Papilomavirus (HPV) infection

O carcinoma oral de células escamosas (OSCC Oral Squamous Cell Carcinoma) é o câncer de cabeça e pescoço mais comum. Somente no Brasil, foram estimados 14.160 novos diagnósticos para o ano de 2009. O HPV está associado com o aumento no risco do câncer oral, mas seu papel na carcinogênese ainda é controverso. A BubR1, uma proteína importante para o checkpoint de fuso mitótico (SAC Spindle Assembly Checkpoint), tem sido associada com algumas proteínas codificadas por espécies virais e com o câncer. O objetivo do presente estudo foi avaliar a expressão de BubR1 em lesões orais benignas e amostras de OSCC com e sem metástase associadas com infecção pelo HPV. Nós realizamos imunoistoquímica para BubR1 em 16 biópsias de lesão oral benigna e em 70 biópsias de OSCC divididas em três grupos (tumores in situ, tumores invasivos sem metástase e tumores invasivos com metástase), com os respectivos linfonodos das amostras com metástase. A técnica de Nested PCR foi realizada com finalidade de detectar DNA do HPV. Nas lesões malignas, foi observada uma significante superexpressão de BubR1 associada com menor sobrevida (p = 0.0479). Houve também correlação significante (r = 1.000) de BubR1 entre as lesões com metástase e seus respectivos linfonodos. Noventa por cento dos OSCC e 100% das lesões benignas foram HPV positivos. HPV 16 e HPV 18 foram detectados em, respectivamente, 13% e 24% das amostras com OSCC HPV-positivas. O HPV teve maior prevalência (76%) nas amostras com alta expressão de BubR1 e a ausência de DNA viral não influenciou no padrão de expressão de BubR1. Esses resultados sugerem uma provável associação do HPV com a superexpressão de BubR1 em OSCC, o que não se aplica para lesões orais benignas.Oral squamous cell carcinoma (OSCC) is the most common head and neck cancer. Only in Brazil, the estimate is that 14,160 new diagnoses will be made in 2009. HPV is associated with increasing risk of oral cancer, but its role in carcinogenesis is still controversial. BubR1, an important protein in the mitotic Spindle Assembly Checkpoint (SAC), has been associated with some virus-encoded proteins and cancer. The aim of the present study was to evaluate the expression of BubR1 in non-malignant oral lesions and OSCC with and without metastasis associated with HPV infection. We performed immunohistochemistry for BubR1 in 16 non-malignant oral lesion biopsies and in 70 OSCC biopsies divided into three groups (in situ tumors, invasive tumors without metastasis and invasive tumors with metastasis) with their respective lymph nodes from samples with metastasis. Nested PCR was performed in order to detect HPV DNA. Significantly higher BubR1 expression associated with shorter survival (p = 0.0479) was observed in malignant lesions. There was also a significant correlation (r = 1.000) with BubR1 expression in lesions with metastasis and their lymph nodes. Ninety percent of OSCC and 100% of benign lesions were HPV positive. HPV 16 and HPV 18 were present in 13% and 24% of HPV-positive OSCC samples, respectively. HPV was more prevalent (76%) in samples with high BubR1 expression and the absence of viral DNA had no influence on BubR1 expression. These findings suggest that HPV could be associated with overexpression of BubR1 in OSCC, but not in benign oral lesions

In situ assessment of Mindin as a biomarker of podocyte lesions in diabetic nephropathy.

Diabetic nephropathy (DN) is the leading cause of chronic kidney disease and end-stage renal failure worldwide. Several mechanisms are involved in the pathogenesis of this disease, which culminate in morphological changes such as podocyte injury. Despite the complex diagnosis and pathogenesis, limited attempts have been made to establish new biomarkers for DN. The higher concentration of Mindin protein in the urine of patients with type 2 diabetes mellitus suggests that it plays a role in DN. Therefore, this study investigated whether in situ protein expression of Mindin can be considered a potential DN biomarker. Fifty renal biopsies from patients diagnosed with DN, 57 with nondiabetic glomerular diseases, including 17 with focal segmental glomerulosclerosis (FSGS), 14 with minimal lesion disease (MLD) and 27 with immunoglobulin A nephropathy (IgAN), and 23 adult kidney samples from autopsies (control group) were evaluated for Mindin expression by immunohistochemistry. Podocyte density was inferred by Wilms' tumor 1 (WT1) immunostaining, while foot process effacement was assessed by transmission electron microscopy. Receiver operative characteristic (ROC) analysis was performed to determine the biomarker sensitivity/specificity. Low podocyte density and increased Mindin expression were observed in all cases of DN, regardless of their class. In the DN group, Mindin expression was significantly higher than that in the FSGS, MCD, IgAN and control groups. Higher Mindin expression was significantly positively correlated with foot process effacement only in class III DN cases. Furthermore, Mindin protein presented high specificity in the biopsies of patients with DN (p < 0.0001). Our data suggest that Mindin may play a role in DN pathogenesis and is a promising biomarker of podocyte lesions

IGF2 and IGF1R in pediatric adrenocortical tumors: roles in metastasis and steroidogenesis

Deregulation of the IGF system observed in human tumors indicates a role in malignant cell transformation and in tumor cell proliferation. Although overexpression of the IGF2 and IGF1R genes was described in adrenocortical tumors (ACTs), few studies reported their profiles in pediatric ACTs. In this study, the IGF2 and IGF1R expression was evaluated by RT-qPCR according to the patient’s clinical/pathological features in 60 pediatric ACT samples, and IGF1R protein was investigated in 45 samples by immunohistochemistry (IHC). Whole transcriptome and functional assays were conducted after IGF1R inhibition with OSI-906 in NCI-H295A cell line. Significant IGF2 overexpression was found in tumor samples when compared with non-neoplastic samples (P1μM) showed limited reduction of cell viability, suggesting that OSI-906 alone could not be a suitable therapy to abolish carcinoma cell growth.236481493CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESPSem informação2010/07020-

A simplified approach using Taqman low-density array for medulloblastoma subgrouping

Abstract Next-generation sequencing platforms are routinely used for molecular assignment due to their high impact for risk stratification and prognosis in medulloblastomas. Yet, low and middle-income countries still lack an accurate cost-effective platform to perform this allocation. TaqMan Low Density array (TLDA) assay was performed using a set of 20 genes in 92 medulloblastoma samples. The same methodology was assessed in silico using microarray data for 763 medulloblastoma samples from the GSE85217 study, which performed MB classification by a robust integrative method (Transcriptional, Methylation and cytogenetic profile). Furthermore, we validated in 11 MBs samples our proposed method by Methylation Array 450 K to assess methylation profile along with 390 MB samples (GSE109381) and copy number variations. TLDA with only 20 genes accurately assigned MB samples into WNT, SHH, Group 3 and Group 4 using Pearson distance with the average-linkage algorithm and showed concordance with molecular assignment provided by Methylation Array 450 k. Similarly, we tested this simplified set of gene signatures in 763 MB samples and we were able to recapitulate molecular assignment with an accuracy of 99.1% (SHH), 94.29% (WNT), 92.36% (Group 3) and 95.40% (Group 4), against 97.31, 97.14, 88.89 and 97.24% (respectively) with the Ward.D2 algorithm. t-SNE analysis revealed a high level of concordance (k = 4) with minor overlapping features between Group 3 and Group 4. Finally, we condensed the number of genes to 6 without significantly losing accuracy in classifying samples into SHH, WNT and non-SHH/non-WNT subgroups. Additionally, we found a relatively high frequency of WNT subgroup in our cohort, which requires further epidemiological studies. TLDA is a rapid, simple and cost-effective assay for classifying MB in low/middle income countries. A simplified method using six genes and restricting the final stratification into SHH, WNT and non-SHH/non-WNT appears to be a very interesting approach for rapid clinical decision-making