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Lack of estrogen receptor (ER) and presence of vimentin (VIM) associate with poor prognosis in human breast cancer. We have explored the relationships between ER, VIM, and invasiveness in human breast cancer cell lines. In the matrigel outgrowth assay, ER+/VIM‐ (MCF‐7, T47D, ZR‐75‐1), and ER‐/VIM‐ (MDA‐MB‐468, SK‐Br‐3) cell lines were uninvasive, while ER‐/VIM+ (BT549, MDA‐MB‐231, MDA‐MB‐435, MDA‐MB‐436, Hs578T) lines formed invasive, penetrating colonies. Similarly, ER‐/VIM+ cell lines were significantly more invasive than either the ER+/VIM‐ or ER‐/VIM‐ cell lines in the Boyden chamber chemoinvasion assay. Invasive activity in nude mice was only seen with ER‐/VIM+ cell lines MDA‐MB‐231, MDA‐MB‐435 and MDA‐MB‐436. Hs578T cells (ER‐/VIM +) showed hematogenous dissemination to the lungs in one of five mice, but lacked local invasion. The ER‐/VIM+ MCF‐7_ADR …