Department of Internal Medicine, Division of Hematology/Oncology, Brody School of Medicine, East Carolina University, Greenville, NC, 27834, USA.
Department of Clinical and Diagnostic Sciences, School of Health Professions, University of Alabama at Birmingham, Birmingham, AL, 35294, USA.
Adv Biol Regul. 2023 May;88:100957. doi: 10.1016/j.jbior.2023.100957. Epub 2023 Jan 28.
Colorectal cancer (CRC) remains a challenging disease to treat due to several factors including stemness and epithelial to mesenchymal transition (EMT). Dysfunctional signaling pathways such as Notch and TGF-β contribute to these phenomena. We previously found that cells expressing constitutively active Notch1 also had increased expression of Smad3, an important member of the TGF-β signaling pathway. We hypothesized that Smad3, mediates the Notch-induced stemness and EMT observed in CRC cells. The human colorectal carcinoma cell line HCT-116, stably transduced with constitutively active Notch-1 (ICN) or a GFP-vector control was treated with different combinations of TGF-β1, DAPT (a Notch inhibitor), or SIS3 (a Smad3 inhibitor). Western blot analysis was performed to determine the effects of Smad3 stimulation and inhibition on Notch and potential downstream EMT-related targets, CD44, Slug and Snail. Smad3 inhibition induced a decrease in Notch1 and Notch3 receptor expression and effectively inhibited CD44, Slug, and Snail expression. Colosphere forming ability was also reduced in cells with inhibited Smad3. These results indicate a key role of TGF-β signaling in Notch1-induced tumorigenesis, and suggest a potential use for Smad3 inhibitors in combination with Notch1 inhibitors that are already in use for CRC treatments.
结直肠癌（CRC）仍然是一种具有挑战性的疾病，其治疗困难的原因有几个，包括干性和上皮间质转化（EMT）。功能失调的信号通路，如 Notch 和 TGF-β，促成了这些现象。我们之前发现，表达组成性激活 Notch1 的细胞也增加了 TGF-β信号通路的重要成员 Smad3 的表达。我们假设 Smad3 介导了 CRC 细胞中 Notch 诱导的干性和 EMT。我们用人结直肠癌细胞系 HCT-116，稳定转染组成性激活 Notch-1（ICN）或 GFP-载体对照，用不同组合的 TGF-β1、DAPT（Notch 抑制剂）或 SIS3（Smad3 抑制剂）进行处理。通过 Western blot 分析来确定 Smad3 刺激和抑制对 Notch 和潜在的 EMT 相关下游靶标 CD44、Slug 和 Snail 的影响。Smad3 抑制诱导 Notch1 和 Notch3 受体表达下降，并有效抑制 CD44、Slug 和 Snail 的表达。抑制 Smad3 的细胞集落形成能力也降低了。这些结果表明 TGF-β 信号在 Notch1 诱导的肿瘤发生中起着关键作用，并表明 Smad3 抑制剂与 Notch1 抑制剂联合使用可能对已经用于 CRC 治疗的 Notch1 抑制剂有一定作用。
