Background Chemotherapy is an important component in the treatment paradigm for breast cancers. However, the resistance of cancer cells to chemotherapeutic agents frequently results in the subsequent recurrence and metastasis. Identification of molecular markers to predict treatment outcome is therefore warranted. The aim of the present study was to evaluate whether expression of circulating microRNAs (miRNAs) can predict clinical outcome in breast cancer patients treated with adjuvant chemotherapy. Methodology/Principal Findings Circulating miRNAs in blood serum prior to treatment were determined by quantitative Real-Time PCR in 56 breast cancer patients with invasive ductal carcinoma and pre-operative neoadjuvant chemotherapy. Proliferating cell nuclear antigen (PCNA) immunostaining and TUNEL were performed in surgical samples to determine the effects of chemotherapy on cancer cell proliferation and apoptosis, respectively. Among the miRNAs tested, only miR-125b was significantly associated with therapeutic response, exhibiting higher expression level in non-responsive patients (n = 26, 46%; p = 0.008). In addition, breast cancers with high miR-125b expression had higher percentage of proliferating cells and lower percentage of apoptotic cells in the corresponding surgical specimens obtained after neoadjuvant chemotherapy. Increased resistance to anticancer drug was observed in vitro in breast cancer cells with ectopic miR-125b expression; conversely, reducing miR-125b level sensitized breast cancer cells to chemotherapy. Moreover, we demonstrated that the E2F3 was a direct target of miR-125b in breast cancer cells. Conclusions/Significance These data suggest that circulating miR-125b expression is associated with chemotherapeutic resistance of breast cancer. This finding has important implications in the development of targeted therapeutics for overcoming chemotherapeutic resistance in novel anti-cancer strategies.