Bt549

The TNBC cell lines MDA-MB-231 (PerkinElmer, Inc. CT), Hs578T (American Type Culture Collection, ATCC), BT549, and 4T1-Luc (Japanese Collection of Research Bioresources Cell Bank, JCRB), the normal murine mammary gland epithelial cell line NMuMG (ATCC) and the normal human embryonic kidney cell line HEK293 (JCRB) were cultured in MEM or RPMI 1640 (Gibco, MD) containing 10% fetal bovine serum (FBS), and streptomycin-penicillin (100 U/ml). Normal human mammary epithelial MCF10A (ATCC) cells were cultured in Mammary Epithelial Cell Growth Medium (MEGM), including hEGF, insulin, hydrocortisone, and bovine pituitary extract (SingleQuotsTM Kit, Lonza, CA) containing streptomycin-penicillin (100 U/ml). Cells were incubated at 37 °C in an atmosphere of 5% CO₂. All cell lines were authenticated by short tandem repeat (STR) profiling by Macrogen Inc (Seoul, South Korea).

Among thirteen BC and two non-cancerous breast epithelial cell lines, BT-549, HCC1419, HCC1954, and Hs578T cell lines were purchased from the Japanese Collection of Research Bioresources Cell Bank (Osaka, Japan), and BT-474, MCF-7, and MCF-12A were obtained from the laboratory of Professor David Sidransky at Johns Hopkins University (Baltimore, MD, USA), following the material transfer agreement. Other cell lines were obtained from the American Type Culture Collection (Manassas, VA, USA). These cell lines were cultured in a medium consisting of RPMI 1640 (Sigma-Aldrich, St. Louis, MO, USA) and 10% fetal bovine serum in an atmosphere of 5% CO₂ at 37 °C [12 (link)].
For clinical specimens, BC patients who were operated on at Nagoya University Hospital from March 2002 to November 2009 and had available postoperative surveillance data spanning more than five years were evaluated in this study. The cancerous specimens were resected at approximately 1.5 mm in diameter and frozen immediately at −80 °C. The resected BC specimens were histologically diagnosed and categorized according to the Union for International Cancer Control (UICC) staging system for BC (8th edition). The perioperative pharmacological treatment of each patient was determined by physician discretion based on the general condition and pathological features.

The TNBC cell lines MDA-MB-231 (PerkinElmer Inc), BT549, 4T1-Luc (Japanese Collection of Research Bioresources Cell Bank, JCRB) and normal mouse fibroblast line NIH/3T3 (ATCC) were cultured in MEM, DMEM or RPMI 1640 (Gibco, MD) containing 10% fetal bovine serum (FBS), and streptomycin-penicillin (100 U/ml). Cells were incubated at 37 °C in an atmosphere of 5% CO₂. All cell lines were authenticated by short tandem repeat (STR) profiling by Macrogen Inc (Seoul, South Korea).