In vivo isolation of a highly-aggressive variant of triple-negative human breast cancer MDA-MB-231 using serial orthotopic transplantation

Aim: We describe the development of a highlyinvasive, triple-negative breast cancer (TNBC) variant using serial orthotopic implantation of MDA-MB-231 human breast cancer in nude mice. Materials and Methods: MDA-MB-231 cells expressing red fluorescent protein (RFP) (1×107 cells/site) were initially injected subcutaneously in the flank of nude mice. After the subcutaneous tumors grew, they were harvested and cut into small pieces for orthotopic implantation in the right lower mammary gland. After the orthotopic tumors grew, they were resected and cut into small pieces and orthotopically reimplanted into the mammary gland of nude mice. The tumors grew and metastasized to lymph nodes. The lymph node metastases were harvested and cut into small pieces and orthotopically re-implanted into the mammary gland of nude mice. After the orthotopic tumors grew, the tumor was removed leaving residual cancer cells, which grew and metastasized to lymph nodes. The lymph node metastases were harvested, cut into pieces and orthotopically re-implanted into the mammary gland of nude mice for two cycles and then isolated. Results: The isolated variant is highly invasive in the mammary gland and metastasized to lymph nodes in 10 of 12 mice compared to 2 of 12 of the parental cell line. Conclusion: The availability of a highly invasive variant of TNBC targeting lymph nodes will be very useful for drug discovery of TNBC, a recalcitrant cancer and for mechanistic studies of its aggressiveness.