What happened when estrogen receptors were introduced into ER-negative breast cancer cells?

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Growth of estrogen receptor (ER)-positive breast cancer cells is hormonally regulated, while the abstract states that most breast cancers are ER-negative and unresponsive to hormonal therapy.

To test whether hormonal control of replication could be restored, researchers transfected ER-negative MDA-MB-231 cells (clone 10A) with sense and antisense constitutive ER expression vectors. These contained either a wild-type or mutant ER gene linked to the gene for neomycin resistance aminoglycoside phosphotransferase (neo).

They used Northern blot analysis on total RNA from eight of the 10 transfectant clones produced to detect messenger RNA for ER and neo. They also used Western blot analysis on protein from one mutant and two wild-type ER sense transfectant clones to determine ER molecular weight. ER levels were measured using both enzyme immunoassay and ligand-binding methods.

To test whether the receptors were functional, the researchers examined effects of 17 beta-estradiol (E2) and/or the antiestrogen ICI 164,384 on ER-activated gene regulation, progesterone receptor induction, DNA replication and cell cycle kinetics. For the gene-regulation test, they transiently transfected the cells a second time with a reporter plasmid containing an estrogen response element linked to the chloramphenicol acetyl transferase (CAT) gene.

Messenger RNA for both ER and neo was detectable in sense and antisense clones. Both mutant and wild-type sense transfectants expressed ER protein with a molecular weight similar to that in ER-positive control cells. Ligand binding detected high ER levels in both wild-type and mutant transfectants, whereas enzyme immunoassay detected lower levels in mutant transfectants.

ER in both wild-type and mutant sense transfectants appeared functional: E2 stimulated expression of reporter-linked CAT and progesterone receptor. E2 inhibited DNA replication in wild-type sense transfectants at 10⁻¹⁰ M and in mutant sense transfectants at 10⁻⁸ M; ICI 164,384 blocked this effect.

The researchers concluded that ER-negative breast cancer cells stably transfected with either mutant or wild-type ER regained hormonal responsiveness, but E2 inhibited rather than stimulated cell growth. They suggested that reactivating quiescent ER may offer a novel approach to controlling ER-negative breast cancers.