Cancer and Metabolism Laboratory, Faculty of Medicine, Saint-Joseph University, Beirut, Lebanon.
BMC Cancer. 2024 May 21;24(1):615. doi: 10.1186/s12885-024-12379-3.
Breast cancer (BC) is the most commonly diagnosed cancer in women. Treatment approaches that differ between estrogen-positive (ER+) and triple-negative BC cells (TNBCs) and may subsequently affect cancer biomarkers, such as H19 and telomerase, are an emanating delight in BC research. For instance, all-trans-Retinoic acid (ATRA) could represent a potent regulator of these oncogenes, regulating microRNAs, mostly let-7a microRNA (miR-let-7a), which targets the glycolysis pathway, mainly pyruvate kinase M2 (PKM2) and lactate dehydrogenase A (LDHA) enzymes. Here, we investigated the potential role of ATRA in H19, telomerase, miR-let-7a, and glycolytic enzymes modulation in ER + and TNBC cells.
MCF-7 and MDA-MB-231 cells were treated with 5 µM ATRA and/or 100 nM fulvestrant. Then, ATRA-treated or control MCF-7 cells were transfected with either H19 or hTERT siRNA. Afterward, ATRA-treated or untreated MDA-MB-231 cells were transfected with estrogen receptor alpha ER(α) or beta ER(β) expression plasmids. RNA expression was evaluated by RT‒qPCR, and proteins were assessed by Western blot. PKM2 activity was measured using an NADH/LDH coupled enzymatic assay, and telomerase activity was evaluated with a quantitative telomeric repeat amplification protocol assay. Student's t-test or one-way ANOVA was used to analyze data from replicates.
Our results showed that MCF-7 cells were more responsive to ATRA than MDA-MB-231 cells. In MCF-7 cells, ATRA and/or fulvestrant decreased ER(α), H19, telomerase, PKM2, and LDHA, whereas ER(β) and miR-let-7a increased. H19 or hTERT knockdown with or without ATRA treatment showed similar results to those obtained after ATRA treatment, and a potential interconnection between H19 and hTERT was found. However, in MDA-MB-231 cells, RNA expression of the aforementioned genes was modulated after ATRA and/or fulvestrant, with no significant effect on protein and activity levels. Overexpression of ER(α) or ER(β) in MDA-MB-231 cells induced telomerase activity, PKM2 and LDHA expression, in which ATRA treatment combined with plasmid transfection decreased glycolytic enzyme expression.
To the best of our knowledge, our study is the first to elucidate a new potential interaction between the estrogen receptor and glycolytic enzymes in ER + BC cells through miR-let-7a.
乳腺癌（BC）是女性最常见的癌症。雌激素阳性（ER+）和三阴性乳腺癌细胞（TNBCs）之间的治疗方法不同，可能随后影响癌症生物标志物，如 H19 和端粒酶，这是 BC 研究中的一个令人欣喜的发现。例如，全反式视黄酸（ATRA）可能是这些癌基因的有效调节剂，调节 microRNAs，主要是 let-7a microRNA（miR-let-7a），其靶向糖酵解途径，主要是丙酮酸激酶 M2（PKM2）和乳酸脱氢酶 A（LDHA）酶。在这里，我们研究了 ATRA 在 ER+和 TNBC 细胞中调节 H19、端粒酶、miR-let-7a 和糖酵解酶的潜在作用。
用 5 µM ATRA 和/或 100 nM 氟维司群处理 MCF-7 和 MDA-MB-231 细胞。然后，用 ATRA 处理或对照 MCF-7 细胞转染 H19 或 hTERT siRNA。此后，用雌激素受体 alpha ER(α)或 beta ER(β)表达质粒转染 ATRA 处理或未处理的 MDA-MB-231 细胞。通过 RT‒qPCR 评估 RNA 表达，通过 Western blot 评估蛋白质。使用 NADH/LDH 偶联酶测定法测量 PKM2 活性，通过定量端粒重复扩增协议测定法评估端粒酶活性。使用 Student's t-test 或单因素方差分析对重复数据进行分析。
我们的结果表明，MCF-7 细胞对 ATRA 的反应比 MDA-MB-231 细胞更敏感。在 MCF-7 细胞中，ATRA 和/或氟维司群降低了 ER(α)、H19、端粒酶、PKM2 和 LDHA，而 ER(β)和 miR-let-7a 增加。用 ATRA 处理或不处理 H19 或 hTERT 敲低的结果与 ATRA 处理后的结果相似，并且发现了 H19 和 hTERT 之间的潜在相互联系。然而，在 MDA-MB-231 细胞中，上述基因的 RNA 表达在 ATRA 和/或氟维司群处理后发生了变化，但对蛋白质和活性水平没有显著影响。在 MDA-MB-231 细胞中转染 ER(α)或 ER(β)可诱导端粒酶活性、PKM2 和 LDHA 的表达，其中 ATRA 联合质粒转染可降低糖酵解酶的表达。
据我们所知，我们的研究首次通过 miR-let-7a 阐明了 ER+BC 细胞中雌激素受体和糖酵解酶之间的新的潜在相互作用。
