Breast cancer is one of the leading reasons of mortality due to cancer globally. Estrogen receptor-positive (ER +) breast cancer being a significant subtype. The therapeutic potential of Vitamin D₃ in cancer treatment has gained attention due to its ability to modulate key molecular targets and signaling pathways. This study investigates the anticancer mechanisms of Vitamin D₃ in MCF-7 breast cancer cells using network pharmacology and omics technology approach. Utilizing protein–protein interaction (PPI) networks, we identified several critical protein targets involved in breast cancer progression, including ESR1, ESR2, PGR, IGF1R, and KDR. Pathway enrichment analyses highlighted Vitamin D₃’s impact on pivotal signaling pathways such as the PI3K/Akt pathway, estrogen receptor signaling, and apoptosis regulation. In vitro studies showed that Vitamin D₃ significantly inhibited cell proliferation in MCF-7 cells. It also induced apoptosis and disrupted mitochondrial function. Flow cytometry analysis demonstrated a dose-dependent increase in apoptotic cell death and S-phase cell cycle arrest. Confocal imaging and mitochondrial membrane potential assays further supported the findings, indicating mitochondrial dysfunction and chromatin condensation. Additionally, gene expression analysis in breast invasive carcinoma tissues confirmed the relevance of ESR1 and PGR in hormone receptor-positive breast cancer. Histopathological studies on DMBA-induced mammary carcinoma revealed Vitamin D₃’s protective effects, reducing tumor malignancy severity through anti-proliferative and pro-apoptotic actions. These findings provide strong evidence for Vitamin D₃’s potential as a multi-targeted therapeutic agent in breast cancer, suggesting further investigation into its clinical applications and combination strategies with existing therapies as an adjunct or alternative in the treatment.

乳腺癌是全球癌症导致死亡的主要原因之一。雌激素受体阳性 （ER +） 乳腺癌是一种重要的亚型。维生素 D₃ 在癌症治疗中的治疗潜力因其调节关键分子靶标和信号通路的能力而受到关注。本研究采用网络药理学和组学技术方法探讨维生素 D₃ 在 MCF-7 乳腺癌细胞中的抗癌机制。利用蛋白质-蛋白质相互作用 （PPI） 网络，我们确定了参与乳腺癌进展的几个关键蛋白质靶点，包括 ESR1、ESR2、PGR、IGF1R 和 KDR。通路富集分析强调了维生素 D₃ 对关键信号通路的影响，例如 PI3K/Akt 通路、雌激素受体信号传导和细胞凋亡调节。体外研究表明，维生素 D₃ 显着抑制 MCF-7 细胞中的细胞增殖。它还诱导细胞凋亡并破坏线粒体功能。流式细胞术分析显示凋亡细胞死亡和 S 期细胞周期停滞呈剂量依赖性增加。共聚焦成像和线粒体膜电位测定进一步支持了这些发现，表明线粒体功能障碍和染色质浓缩。此外，乳腺浸润癌组织中的基因表达分析证实了 ESR1 和 PGR 在激素受体阳性乳腺癌中的相关性。对 DMBA 诱导的乳腺癌的组织病理学研究揭示了维生素 D₃ 的保护作用，通过抗增殖和促凋亡作用降低肿瘤恶性肿瘤的严重程度。 这些发现为维生素 D₃ 作为乳腺癌多靶点治疗剂的潜力提供了强有力的证据，表明进一步研究其临床应用以及与现有疗法相结合的策略作为治疗的辅助或替代。