Analysis of Ki-67 expression in women with breast cancer: Comparative evaluation of two different methodologies by immunophenotyping.

The Ki-67 antigen is a nuclear protein with proven prognostic value in different neoplasms and recognizes the predictive value in breast cancer (BC). No consensus exists on the ideal cutoff point. In this study, Ki-67 expression was evaluated in samples of BC by flow cytometry (FC) and compared with immunohistochemical (IHC) examination. For this, the BC tissue samples were sectioned, macerated, filtered, and marked with anti-Ki-67 FITC and anti-CD45 V500 antibodies. We selected the neoplastic cells according to CD45 expression and size and internal complexity (FSC × SSC) using the Infinicity 1.7 software. Lymphocytes were negative control. We compared the results with IHC analyses carried out in parallel and independently. The expression of Ki-67 was evaluated in both methodologies through Bland-Altman analysis. Among the 44 samples analyzed, only three showed bias higher than the established confidence interval (mean bias 2.1%, p = 0.62), with no significant difference for the perfect mean bias (0%). Therefore, one can state that FC provides results equivalent to IHC analysis and possibly analyzes more cells simultaneously. The results obtained in this study show the absence of observational bias through software analysis in a larger number of tumor cell populations. We can conclude that FC may be a promising alternative method for investigating Ki-67 in solid tumours.

Evaluation of ER, PR and HER2 markers by flow cytometry for breast cancer diagnosis and prognosis.

BACKGROUND AND AIMS: Laboratory diagnosis of breast cancer (BC) is done by morphological analysis and immunohistochemistry (IHC). However, this methodology still has some limitations. The aim of this study is to validate flow cytometry (FC) immunophenotyping to investigate diagnostic and prognostic markers of BC. METHODS: Tumor samples from surgical specimens of patients previously diagnosed with BC, were first sliced and then macerated together with PBS. Then, sample was filtered and the single cell suspension obtained was labeled with antibodies against estrogen (ERα), progesterone (PR) and HER2 receptors and CD45. The results were compared, in terms of sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV), with reference methods. RESULTS: Results obtained comparing FC with reference methods were: ERα detection (sensitivity: 75%; specificity: 90%; PPV: 96.7%; NPV: 47.4%); PR detection (sensitivity: 72%; specificity: 70%; PPV: 79.3%; NPV: 60.8%); HER2 detection (sensitivity: 80%; specificity: 90.2%; PPV: 66.7%; NPV: 94.9%). CONCLUSION: The results obtained show the capacity of this methodology on BC markers differentiation. FC, together with morphological analysis and IHC can overcome individual limitations of each methodology and provide reliable results on a faster and efficient manner, resulting in improvements on BC diagnosis and prognosis.

Association of TNFRSF1A and IFNLR1 Gene Polymorphisms with the Risk of Developing Breast Cancer and Clinical Pathologic Features.

Several genes have been associated with breast cancer (BC) susceptibility. The tumor necrosis factor receptor superfamily, member 1A (TNFRSF1A), and interferon lambda receptor 1 (IFNLR1) genes encode receptors that mediate the action of inflammatory cytokines. Previous studies have demonstrated the association of the variants rs1800693 (TNFRSF1A) and rs4649203 (IFNLR1) with some inflammatory diseases. The present study aimed to verify a possible association of these variants with BC, its clinical pathologic features, as well as epidemiological data in a Brazilian population. A total of 243 patients and 294 individuals without history of BC were genotyped for these polymorphisms through TaqMan® SNP genotyping assays by qPCR. For the TNFRSF1A gene, no significant results were found. For IFNLR1, the AA genotype (p = 0.008) and the A allele (p = 0.02) were significantly associated with a lower risk of developing BC. When analyzing the age, it was observed that each increase of one year contributes to the development of BC (p < 0.001). Also, the smoking habit (p < 0.001) and body mass index (p = 0.018) increase the risk of disease development. Analyzing progesterone receptor factor an association was found with the AA genotype of the IFNLR1 (p = 0.02). The findings suggest that polymorphism in the immune-related IFNLR1 gene contribute to BC susceptibility in a Brazilian population. These findings can contribute to the further understanding of the role this gene and pathways in BC development.