Study the Expression of Two Circular RNAs, hsa_circ_0003227 and hsa_circ_0001666, in the Primary Breast Cancer Cell Line BT-20 and the Metastatic Breast Cancer Cell Line MCF-7.

Breast cancer is one of the most prevalent cancers globally and remains a significant cause of cancer-related mortality among women despite advancements in early detection and treatment. The heterogeneity of breast cancer arises from a complex interplay of genetic and environmental factors. Early-stage breast cancer is often asymptomatic, with initial signs including subtle changes in breast morphology and localized swelling, emphasizing the need for reliable diagnostic tools for early detection. Recent research has highlighted the potential of molecular biomarkers, particularly non-coding RNAs such as circular RNAs (circRNAs), in cancer diagnosis. CircRNAs, a unique subset of non-coding RNAs, are characterized by their covalently closed-loop structure, which confers exceptional stability and resistance to exonuclease degradation. They are present in various body fluids and have demonstrated regulatory roles in transcription, translation, and as microRNA sponges, making them promising candidates for cancer diagnostics and prognostics. This study focuses on evaluating the diagnostic potential of two circRNAs, hsa_circ_0001666 and hsa_circ_0003227, by examining their expression in normal, tumor, and metastatic breast cancer cell lines. Breast cancer cell lines representing normal (MCF-10A), tumor (MCF-7), and metastatic (BT-20) stages were cultured for analysis. Total RNA was extracted using a column-based RNA extraction kit, and RNA quality was assessed through NanoDrop spectrophotometry and agarose gel electrophoresis. Complementary DNA (cDNA) synthesis was performed using random hexamers, and the expression levels of hsa_circ_0001666 and hsa_circ_0003227 were quantified using Real-Time Quantitative Reverse Transcription PCR (RT-qPCR), with beta-actin serving as the internal control. Statistical analyses were conducted using SPSS software to evaluate differences in expression levels across cell lines. A significant downregulation of hsa_circ_0001666 and hsa_circ_0003227 was observed in tumor and metastatic cell lines compared to normal breast cell lines (P < 0.05). These results suggest that the expression of these circRNAs correlates with the progression of breast cancer, with decreased levels observed as cells transition from normal to tumorigenic and metastatic stages. The findings of this study indicate that hsa_circ_0001666 and hsa_circ_0003227 have potential utility as diagnostic biomarkers for breast cancer. Their significant expression changes across different stages of breast cancer highlight their relevance in early detection and disease monitoring. This study reinforces the potential of RNA-based biomarkers, particularly circRNAs, in cancer diagnosis and treatment. However, in vitro findings require validation in clinical samples and larger cohorts. Future research should explore their roles in breast cancer progression and integration into non-invasive diagnostics.