Polyether ionophores (LAS, MON, SAL) in their sodium salts form, BET, all newly synthesized dimers 14–24, and two commonly used anticancer drugs, cisplatin and doxorubicin, were evaluated for in vitro antiproliferative activity against a panel of six cancer cell lines: human acute myeloid leukemia cell line (MV-4-11), three human breast cancer cell lines with different characteristics (JIMT-1, MCF-7, SKBR-3), and human colon adenocarcinoma cell lines including the drug-sensitive variant (LoVo), but also its drug-resistant sub-line with the ABCB1 overexpression (LoVo/DX). The gene discussed is ABCB1; the disease is breast cancer.