Melatonin and estradiol action on let-7 family members in triple-negative breast cancer cells

The widely studied tumor suppressor miRNAs of the let-7 family are involved in many cellular processes through the regulation of several signaling pathways. They shared identical seed sequences and have variable stem-loop regions. The let-7 family regulates crucial physiological functions in cells. However, low expression is frequently found, and it was associated with cell proliferation, migration, and metastasis in triple-negative breast cancer (BC). Although the MDA-MB231 BC cell line is triple-negative, it has been shown to express estrogen receptor beta. Let-7 has a regulatory relationship with estrogen, acting as a negative regulator of estrogen receptors. The recovery or restoration of let-7 expression seems to be a key point to prevent breast cancer growth and development. In this context, melatonin (Mel) has gained notoriety due to its versatile and essential mechanisms for antagonizing tumor features, which involve modulation of differential miRNA expression. Studies have demonstrated that Mel can alter the expression of some members of the let-7 family in some contexts. Objective: We aimed to demonstrate the effects of Mel and 17beta-estradiol (E2) treatments on the proliferation of triple-negative BC and assess the expression profile of six let-7 family members (let-7a/b/c/g/i/f). Methods: MDA-MB231 cells were treated with 3 mM Mel and 100 nM E2, and the two drugs were combined (ME2) and were replaced daily for up to 96h. Cell proliferation was evaluated in treated cells. RNAs were extracted, and reverse transcription was performed. RT-PCR was carried out for the detection of selected let-7 family members. Results: Our preliminary results showed a significant decrease in the proliferation assay in the cell line after Mel and ME2 treatments. Cells presented a lower rate of proliferation at 72h of Mel and ME2 treatments. Mel treatment leads to slightly and significantly increased expression of let-7c/g/f (Fold:1.939; 1.132; 1.624, respectively), and the let-7c/g also showed higher expression related to ME2. Cells treated with ME2 showed a significant increase of let-7b/i/f (Fold:1.112; 1.123; 1.506, respectively) compared to CC and E2. Let-7c (Fold: 0.879) presented a significant decrease compared to CC. Additionally, lower expression of let-7a/b/g/i (Fold: 0.707; 0.823; 0.662; 0.759, respectively) was observed in cells treated with E2. Conclusion: Our results indicate that Mel can minimize E2 effects and inhibit the proliferation of triple-negative BC cells through the regulation of specific miRNAs. It seems to recover the expression of tumor suppressor miRNAs of the let-7 family in triple-negative BC. Supported by: FAPESP 2025/08621-1, 2022/04174-2, and 2018/24224-9, and CNPq 444345/2024-8.