Progestins and breast cancer hallmarks: The role of the ERK1/2 and JNK pathways in estrogen receptor positive breast cancer cells.

Progestins used in hormonal contraceptives and menopausal hormone therapy (MHT) have been linked to increased breast cancer risk. Whether the association holds for all progestins is unclear and the underlying mechanisms remain poorly understood. We directly compared the effects of four progestins (medroxyprogesterone acetate (MPA), norethisterone acetate (NET-A), levonorgestrel (LNG) and drospirenone (DRSP)) to each other and the natural progestogen progesterone (P4) on selected cancer hallmarks. To provide mechanistic insight into these effects, we assessed the role of the progesterone receptor (PR), and the extracellular signal-related kinase (ERK1/2) and c-Jun N terminal (JNK) signaling pathways. We showed that the increased proliferation of the luminal T47D breast cancer cell line by P4 and all progestins, albeit to different extents, was inhibited by PR knockdown and inhibition of both the ERK1/2 and JNK pathways. While knockdown of the PR also blocked the upregulation of MKI67 and CCND1 mRNA expression by selected progestogens, only a role for the ERK1/2 pathway could be established in these effects. Similarly, only a role for the ERK1/2 pathway could be confirmed for progestogen-induced colony formation, whereas both the ERK1/2 and JNK pathways were required for cell migration in response to the three older progestins implicated in the etiology of breast cancer, MPA, NET-A and LNG. Together our results show that all the progestins elicit their effects on cell proliferation via a mechanism requiring the PR, ERK1/2 and JNK pathways. While the ERK1/2 and JNK pathways are also required for increased cell migration by the older progestins, only a role for the ERK1/2 pathway could be established in their effects on colony formation. Notably, the cytoplasmic PR was not needed for activation of the ERK1/2 pathway by the progestogens. Given that DRSP showed significantly lower proliferation than MPA and NET-A, and that it had no effect on breast cancer cell migration and colony formation, hormonal formulations containing the newer generation progestin DRSP may provide a better benefit/risk profile towards breast cancer than those containing the older generation progestins.

Upregulation of an estrogen receptor-regulated gene by first generation progestins requires both the progesterone receptor and estrogen receptor alpha.

Progestins, synthetic compounds designed to mimic the activity of natural progesterone (P4), are used globally in menopausal hormone therapy. Although the older progestins medroxyprogesterone acetate (MPA) and norethisterone (NET) have been implicated in increased breast cancer risk, little is known regarding newer progestins, and no significant risk has been associated with P4. Considering that breast cancer is the leading cause of mortality in women, establishing which progestins increase breast cancer incidence and elucidating the underlying mechanisms is a global priority. We showed for the first time that the newer-generation progestin drospirenone (DRSP) is the least potent progestin in terms of proliferation of the estrogen-responsive MCF-7 BUS breast cancer cell line, while NET and P4 have similar potencies to estradiol (E2), the known driver of breast cancer cell proliferation. Notably, MPA, the progestin most frequently associated with increased breast cancer risk, was significantly more potent than E2. While all the progestogens enhanced the anchorage-independent growth of the MCF-7 BUS cell line, MPA promoted a greater number of colonies than P4, NET or DRSP. None of the progestogens inhibited E2-induced proliferation and anchorage-independent growth. We also showed that under non-estrogenic conditions, MPA and NET, unlike P4 and DRSP, increased the expression of the estrogen receptor (ER) target gene, cathepsin D, via a mechanism requiring the co-recruitment of ERα and the progesterone receptor (PR) to the promoter region. In contrast, all progestogens promoted the association of the PR and ERα on the promoter of the PR target gene, MYC, thereby increasing its expression under non-estrogenic and estrogenic conditions. These results suggest that progestins differentially regulate the way the PR and ER converge to modulate the expression of PR and ER-regulated genes. Our novel findings indicating similarities and differences between P4 and the progestins, emphasize the importance of comparatively investigating effects of individual progestins rather than grouping them as a class. Further studies are required to underpin the clinical relevance of PR/ERα crosstalk in response to different progestins in both normal and malignant breast tissue, to either confirm or refute their suitability in combination therapy for ER-positive breast cancer.