Brane Andrew, Sutko Madeline, Tollefsbol Trygve O
Department of Biology, University of Alabama at Birmingham, 3100 Science & Engineering Complex-East Science Hall, 902 14th Street South, Birmingham, AL 35205, USA.
Comprehensive Cancer Center, University of Alabama at Birmingham, 1802 6th Avenue South, Birmingham, AL 35294, USA.
Int J Mol Sci. 2025 Jan 30;26(3):1210. doi: 10.3390/ijms26031210.
Breast cancer (BC) is a widespread malignancy that affects the lives of millions of women each year, and its resulting financial and healthcare hardships cannot be overstated. These issues, in combination with side effects and obstacles associated with the current standard of care, generate considerable interest in new potential targets for treatment as well as means for BC prevention. One potential preventive compound is Withaferin A (WFA), a traditional medicinal compound found in winter cherries. WFA has shown promise as an anticancer agent and is thought to act primarily through its effects on the epigenome, including, in particular, the methylome. However, the relative importance of specific genes' methylation states to WFA function remains unclear. To address this, we utilized human BC cell lines in combination with CRISPR-dCas9 fused to DNA methylation modifiers (i.e., epigenetic editors) to elucidate the importance of specific genes' promoter methylation states to WFA function and cancer cell viability. We found that targeted demethylation of promoters of the tumor suppressors and within MDA-MB-231/MCF7 cells resulted in around 1.7×/1.5× and 1.2×/1.3× increases in expression, respectively. Targeted methylation of the promoter of the oncogene within MDA-MB-231/MCF7 cells resulted in 0.5×/0.8× decreases in gene expression. These changes to , , and were also associated with decreases in cell viability of around 25%/50%, 5%/35%, and 12%/16%, respectively, for MDA-MB-231/MCF7 cells. When given in combination with WFA in both mutant and wild type cells, we discovered that targeted methylation of the promoter was able to modulate the anticancer effects of WFA, while targeted methylation or demethylation of the promoters of and had no significant effect on viability decreases from WFA treatment. Taken together, these results indicate that , , and may be important targets for future in vivo studies that may lead to epigenetic editing therapies and that WFA may have utility in the prevention of BC through its effect on promoter methylation independent of function.
乳腺癌（BC）是一种广泛存在的恶性肿瘤，每年影响着数百万女性的生活，其造成的经济和医疗负担不容小觑。这些问题，再加上当前护理标准所带来的副作用和障碍，引发了人们对新的潜在治疗靶点以及乳腺癌预防方法的浓厚兴趣。一种潜在的预防化合物是睡茄内酯A（WFA），它是在酸浆中发现的一种传统药用化合物。WFA已显示出作为抗癌剂的潜力，并且被认为主要通过其对表观基因组的作用来发挥作用，特别是对甲基化组的作用。然而，特定基因的甲基化状态对WFA功能的相对重要性仍不清楚。为了解决这个问题，我们将人类乳腺癌细胞系与融合了DNA甲基化修饰剂（即表观遗传编辑器）的CRISPR-dCas9结合使用，以阐明特定基因的启动子甲基化状态对WFA功能和癌细胞活力的重要性。我们发现，在MDA-MB-231/MCF7细胞中，肿瘤抑制基因 和 的启动子靶向去甲基化分别导致表达增加约1.7倍/1.5倍和1.2倍/1.3倍。在MDA-MB-231/MCF7细胞中，癌基因 的启动子靶向甲基化导致基因表达降低0.5倍/0.8倍。对于MDA-MB-231/MCF7细胞，这些对 、 和 的变化还分别与细胞活力降低约25%/50%、5%/35%和12%/16%相关。当在 突变型和野生型细胞中与WFA联合使用时，我们发现 的启动子靶向甲基化能够调节WFA的抗癌作用，而 和 的启动子靶向甲基化或去甲基化对WFA治疗导致的活力降低没有显著影响。综上所述，这些结果表明， 、 和 可能是未来体内研究的重要靶点，这些研究可能会带来表观遗传编辑疗法，并且WFA可能通过其对 启动子甲基化的作用独立于 功能而在预防乳腺癌方面具有效用。
