Investigation of CYP17A1 and CYP19A1 gene expression levels and aromatase activity in invasive ductal breast cancer tissues

Breast cancer is the most common cancer in women worldwide, with nearly 1.7 million new cases diagnosed in 2012. It is the second most common cancer overall. This represents about 12% of all new cancer cases and 25% of all cancers in women. The numbers of incidence and mortality is increasing especially in developing countries as well as in Turkey. Breast cancer is more commonly hormone driven and the factors that modify the risk of this cancer when diagnosed premenopausally and when diagnosed postmenopausally are not the same. Extensive research and clinical observations in the past 20 years confirmed that the cessation of ovarian function at menopause does not stop the process of sex steroid hormone synthesis in females. Currently we acknowledge that multiple extra-ovarian tissues contain the same enzymatic machinery the ovary uses which can maintain a significant rate of local hormonal synthesis sufficient to cause pathological outcomes. This is commonly termed "intracrine". The term intracrinology was first coined over 2 decades ago but there are still questions to be answered, which could help us to understand the intracrine mechanisms in the breast cancer microenvironment. CYP17A1 (P450c17) and CYP19A1 (aromatase) are two of the key enzymes in the central pathways of sex steroid metabolism. In current study, local expressions of CYP17A1, CYP19A1 genes and specific activity levels of aromatase in invasive ductal breast carcinoma tissues were investigated by means of revealing their effect over peripheral and/or intratumoral estrogen production in invasive ductal breast carcinoma tissues. The relationship between these expressions and specific activity status along with the patients' known breast health risk factors and clinicopathological parameters were also reported in order to investigate the effect of tumor progression. One tumor and one peripheral mammary adipose tissue sample (P) adjacent to the tumor was obtained from each patient (n= 20) and snap frozen in liquid nitrogen and kept at -80°C until use for the extraction of total RNA and microsome isolation. Real-time polymerase chain reaction was employed for the detection of CYP17A1 and CYP19A1 gene expression. All patients were postmenopausal, diagnosed for invasive ductal breast carcinoma and classified as luminal A. Patients were divided into groups according to cilinicopathologic features and breast cancer risk factors. In addition, 12 tumor-free breast tissue samples (N) were obtained from premenopausal women with no history of breast cancer who underwent reduction mammoplasty surgery as the control group. The conversion of testosterone to 17ß-estradiol was determined via radioimmunoassay and liquid chromatography-tandem mass spectrometry, and the specific activity of aromatase in microsomal fractions were calculated. Microsomes were isolated from each specimen by employing differential centrifugation method for the activity assays. Bicinchoninic acid protein assay was used for detection and quantitation of the total protein in the samples. The acquired data pointed out the estradiol at the breast tumor microenvironment, which plays a major role in proliferation of malignant epithelial breast cancer cells, was depending on the power of local aromatization activity and the basis of this estrogen drive in the postmenopausal period is the adipose tissue adjacent to the tumor itself. The local aromatase overexpression and high aromatase activity are important factors for the survival of estrogen dependent breast carcinoma cells. Furthermore, there was a pattern consisting the combination upregulated and unaltered gene expressions of CYP17A1 and CYP19A1 which was observed to be coralated with higher aromatase activity levels in peripheral tissues compared to tumor tissues. To summarize, present study suggesting a complex breast tumor progression mechanism altered by CYP17A1 and CYP19A1 at the breast tumor microenvironment. The evaluation of various clinicopathological and disease risk factors along with the expression levels of CYP17A1 and CYP19A1 and the aromatase activity levels at breast tumor microenvironment might help clinicians to decide on treatment startegies and diagnosis for individual cases, particularly with postmenapausal status. The in-house liquid chromatography-tandem mass spectrometry method has the potential to be further developed to a commonly applied high-throughput technique for aromatase activity measurement which might be an invaluable asset for rapid and specific analysis. However, future studies must be conducted using greater sample size and addition of other key enzyme activities evaluations such as 3β-hydroxysteroid dehydrogenase and 17β-hydroxysteroid dehydrogenase in streidogenesis pathway which effect local estrogen levels for confirmation and gettting more strong and reliable results.