Branched rolling circle amplification method for measuring serum circulating microRNA levels for early breast cancer detection

Abstract Serum circulating micro RNA s (c‐mi RNA s) are serving as useful biomarkers for cancer diagnosis. Here, we describe the development of a one‐step branched rolling circle amplification ( BRCA ) method to measure serum c‐mi RNA s levels for early diagnosis of breast cancer. Four c‐mi RNA s, c‐mi RNA 16 (c‐miR‐16), c‐mi RNA 21 (c‐miR‐21), c‐mi RNA 155 (c‐miR‐155), and c‐mi RNA 195 (c‐miR‐195) were isolated from the serum of 49 breast cancer patients (stages I‐ IV ) and 19 healthy controls, and analyzed using one‐step BRCA . The serum levels of c‐miR16, c‐miR21, c‐miR155, and c‐miR195 were higher ( P < 0.0001) in stage I breast cancer patients than healthy controls. These levels were also higher in several breast cancer molecular subtypes ( HER ‐2 over‐expression, Luminal A, Luminal B, and triple negative breast cancer) than in healthy control subjects. The diagnostic accuracy of c‐miR16, c‐miR21, c‐miR155, and c‐miR195 for early diagnosis of breast cancer was confirmed by receiver operating characteristic ( ROC ) curve assay. These results show that the BRCA method can be used to measure serum c‐mi RNA s levels, and that this method has high accuracy, sensitivity, and specificity. Moreover, both BRCA approach and quantitative real‐time PCR ( qRT ‐ PCR ) method show that the serum levels of c‐miR16, c‐miR21, c‐miR155, and c‐miR195 could be used as biomarkers to improve the early diagnosis of breast cancer, and distinguish different breast cancer molecular subtypes.