Evaluation of miRNA-9 and miRNA-34a as potential biomarkers for diagnosis of breast cancer in Iranian women.

PURPOSE: MicroRNAs are involved in diverse biological processes and their dysregulation is a common event in various diseases including breast cancer. Breast cancer is a major threat to women's health. This study was designed to examine the expression levels of miR-9 and miR-34a in breast tumor tissue samples and plasma of breast cancer patients, compare their expression pattern between tissue samples and plasma samples of patients and analyze their relationship with tumor clinical features. Also, the potential of these miRNAs as diagnostic biomarkers for breast cancer was investigated. MATERIALS AND METHODS: The expression levels of miR-9, miR-34a and CDH1 were measured by real-time reverse transcription polymerase chain reaction and ΔΔct method. Data were analyzed using t-test and one-way ANOVA. The sensitivity and specificity of miRNAs were determined by receiver operating characteristic (ROC) curve. RESULTS AND DISCUSSION: The expression levels of miR-9 and miR-34a were significantly down-regulated in tumor tissues compared to healthy tissues (fold change = 0.26, p = 0.0051 for miR-9 and fold change = 0.55, p = 0.021 for miR-34a). While no significant difference was observed in the expression levels of miR-9 (p = 0.205) and miR-34a (p = 0.132) in plasma samples of patients compared to normal plasma. CDH1 expression in tumor tissue was not significantly different from normal tissue (p = 0.33). We found that expression level of miR-9 in patients with tumor size larger than 5 cm (p = 0.026) and expression level of miR-34a in patients with higher stage (lll & lV, p = 0.03) were significantly down-regulated. Also miR-34a expression level was positively correlated with patient's age (p = 0.03). CONCLUSION: According to the ROC curves, the area under the curve (AUC) of miR-9 in tissue was 0.71 (p = 0.009) with sensitivity 83.33% and specificity 70.37%. The AUC for miR-34a in tissue was 0.72 (p = 0.007) with sensitivity 72% and specificity 76%. Thus miR-9 and miR-34a have the capability for distinguishing tumor tissues from healthy tissues and the study of their expression levels in tissue may be used as a biomarker for the diagnosis of breast cancer patients from healthy women.

HER-2/neu gene amplification assessment in breast cancer patients in Isfahan province by real time PCR, differential PCR and immunohistochemistry.

INTRODUCTION: The amplification status of proto oncogene HER-2/neu is one of the major molecular prognosis markers in breast cancer and recent adjuvant treatment with Trastuzumab has increased a request for the evaluation of HER-2/neu status in breast cancer. The aim of our study was the evaluation of HER-2/neu amplification status in malignant breast cancer by PCR techniques such as differential PCR and real time PCR and comparison of results of two methods together and with IHC results in some specimens. METHODS: 86 malignant breast cancer tissue specimens were analysed initially by differential PCR and then by real time PCR. Sections from paraffin-embedded or fresh tissue samples were homogenized by squash and then DNA extraction was performed on cell suspension. A standard curve was initially plotted using BioEasy SYBR Green I for using 2(-ddct) method. A 98bp fragment of HER-2/neu gene was co-amplified in the same reaction tube with a 150bp fragment of INFγ gene for differential PCR. RESULTS: The IHC results existed only for 27 of 83 assessed samples by dPCR and for 30 of 86 assessed samples by real time PCR. 29 out of 83 (35%) samples tested by dPCR and 28 out of 86 (32.5%) samples tested by real time PCR have HER-2/neu gene amplification. CONCLUSION: There was concordance between the results of real time PCR and differential PCR in 61 of 83 specimens (73.5%) tested by both method. Furthermore, in comparison of IHC results with these two methods, 70% concordance between IHC and differential PCR, 63% concordance between IHC and real time PCR and 55.5% concordance between three methods were observed.