Differential Effects of Tamoxifen and Analogs with Nonbasic Side Chains on Cell Proliferationin Vitro

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Abstract
Structural analogs of the nonsteroidal antiestrogen tamoxifen, in which the basic dimethylaminoethoxy side chain was either absent or replaced with a variety of nonbasic side chains, were examined for their ability to inhibit the proliferation of a hormonally responsive cell line, MCF 7 human breast cancer. The degree of inhibition was compared with relative binding affinities for the estrogen receptor (RE) and a microsomal antiestrogen binding site (AEBS). All modifications resulted in loss of detectable affinity for AEBS. Replacement of the basic side chain of tamoxifen with a series of nonbasic side chains reduced affinity for RE by 78–93% except in the case of 1-(4-(1,2-diphenylbut-1-enyl)phenyl)-2,3-butanediol (ICI 145-680) where affinity was unchanged. When the basic side chain of tamoxifen was replaced by a hydroxyl group, to form the estrogenic analog ICI 141389 (Metabolite E), affinity for RE was reduced by 39%. ICI 141389 was a very weak inhibitor of MCF 7 cell growth, showing no significant growth inhibition at concentrations less than 10 μM. Despite the fact that ICI 145680 and tamoxifen had identical affinities for RE, ICI 145680 was a significantly weaker growth inhibitor than tamoxifen over the concentration range studied, i.e. 0.1–20 μm. Differences in potency were greatest at concentrations greater than 7.5 μm where the effects were not reversed by estradiol and where cytotoxicity played a major role in the decrease in cell numbers induced by tamoxifen. Like tamoxifen, ICI 145680 demonstrated both estrogen-reversible (at concentrations between 0.5–7.5 μm) and estrogen-irreversible (10–20 μm) inhibition of MCF 7 cell proliferation which was associated with a concentration-dependent accumulation of cells in the G0/G1 phase of the cell cycle. In contrast to tamoxifen, however, ICI 145680 appeared not to possess cytotoxic activity. Whereas ICI 145680 was without effect on proliferation of the RE negative human breast cancer cell line, MDA-MB-330, at doses less than 20 μm, tamoxifen inhibited growth at concentrations greater than 5 μm, but with changes in cell cycle kinetic parameters that were markedly different from those seen in RE positive cells. These data indicate that among tamoxifen analogs, a basic ether side chain is essential for binding to AEBS, side-chain structure is an important determinant of affinity for RE, potency of antagonists as growth inhibitory agents is correlated with affinity for RE but other factors including affinity for AEBS also determine potency especially in the estrogen-irreversible concentration range and a basic alkylether side chain appears essential for expression of cytotoxic activity in vitro. (Endocrinology116: 1071–1078,1985)