DataSheet1_A small molecule inhibitor of Notch1 modulates stemness and suppresses breast cancer cell growth.pdf

Although breast cancer stem cells (BCSCs) are well characterized, molecularly targeting and eradicating this sub-population remains a challenge in the clinic. Recent studies have explored several signaling pathways that govern stem cell activation: We and others established that the Notch1 signaling plays a significant role in the proliferation, survival, and differentiation of BCSCs. Earlier, we reported that a newly developed small molecule, ASR490, binds to the negative regulatory region (NRR: The activation switch of the Notch receptor) of Notch1. In vitro results demonstrated that ASR490 significantly inhibited BCSCs (ALDH+ and CD44+/CD24–) and breast cancer (BC) growth at nM concentrations, and subsequently inhibited the colony- and mammosphere-forming abilities of BCSCs and BCs. ASR490 downregulated the expressions of Notch1 intracellular domain (NICD: The active form of Notch1) and its downstream effectors Hey1 and HES1. Inhibition of Notch1-NICD facilitated autophagy-mediated growth inhibition by triggering the fusion of autophagosome and autolysosome in BCSCs. ASR490 was found to be non-toxic to healthy cells as compared to existing Notch1 inhibitors. Moreover, oral administration of ASR490 abrogated BCSC and BC tumor growth in the in vivo xenograft models. Together our results indicate that ASR490 is a potential therapeutic agent that inhibits BC tumor growth by targeting and abolishing Notch1 signaling in BCSCs and BC cells.

尽管乳腺癌干细胞（breast cancer stem cells, BCSCs）已得到充分表征，但在临床中通过分子靶向手段清除这一亚群仍是一项挑战。近期研究已探索了多种调控干细胞激活的信号通路：本团队及其他研究团队均证实，Notch1信号通路在乳腺癌干细胞的增殖、存活与分化过程中发挥重要作用。此前，本团队曾报道一种新型小分子化合物ASR490可结合至Notch1的负调控区（negative regulatory region, NRR：Notch受体的激活开关）。体外实验结果显示，在纳摩尔浓度下，ASR490可显著抑制乳腺癌干细胞（ALDH+及CD44+/CD24–表型）与乳腺癌（breast cancer, BC）细胞的增殖，并可进一步抑制乳腺癌干细胞及乳腺癌细胞的集落形成与乳腺球形成能力。ASR490可下调Notch1胞内结构域（Notch1 intracellular domain, NICD：Notch1的活性形式）及其下游效应分子Hey1与HES1的表达水平。抑制Notch1-NICD可通过触发乳腺癌干细胞中自噬体与自溶酶体的融合，增强自噬介导的生长抑制效应。与现有Notch1抑制剂相比，ASR490对正常细胞无毒性。此外，在体内异种移植模型中，口服给予ASR490可完全阻断乳腺癌干细胞及乳腺癌肿瘤的生长。综上，本研究结果表明，ASR490是一种潜在的治疗性药物，可通过靶向并阻断乳腺癌干细胞及乳腺癌细胞中的Notch1信号通路，从而抑制乳腺癌肿瘤的生长。