Triple-negative breast cancer (TNBC), a subset of breast cancers, have poorer survival than other breast cancer types. Recent studies have demonstrated that the abnormal Hedgehog (Hh) pathway is activated in TNBC and that these treatment-resistant cancers are sensitive to inhibition of the Hh pathway. Smoothened (Smo) protein is a vital constituent in Hh signaling and an attractive drug target. Vismodegib (VIS) is one of the most widely studied Smo inhibitors. But the clinical application of Smo inhibitors is limited to adult patients with BCC and AML, with many side effects. Therefore, it’s necessary to develop novel Smo inhibitor with better profiles. Twenty [1,2,4]triazolo[4,3-a]pyridines were designed, synthesized and screened as Smo inhibitors. Four of these novel compounds showed directly bound to Smo protein with stronger binding affinity than VIS. The new compounds showed broad anti-proliferative activity against cancer cell lines in vitro, especially triple-negative breast cancer cells. Mechanistic studies demonstrated that TPB15 markedly induced cell cycle arrest and apoptosis in MDA-MB-468 cells. TPB15 blocked Smo translocation into the cilia and reduced Smo protein and mRNA expression. Furthermore, the expression of the downstream regulatory factor glioma-associated oncogene 1 (Gli1) was significantly inhibited. Finally, TPB15 demonstrated greater anti-tumor activity in our animal models than VIS with lower toxicity. Hence, these results support further optimization of this novel scaffold to develop improved Smo antagonists.

三阴性乳腺癌（TNBC）是乳腺癌的一个子集，其生存率较其他乳腺癌类型差。最近的研究表明，TNBC中激活了异常的Hedgehog（Hh）途径，这些对治疗有抵抗力的癌症对Hh途径的抑制敏感。平滑蛋白（Smo）是Hh信号中的重要组成部分，并且是有吸引力的药物靶标。Vismodegib（VIS）是研究最广泛的Smo抑制剂之一。但是，Smo抑制剂的临床应用仅限于成年BCC和AML患者，具有许多副作用。因此，有必要开发出性能更好的新型Smo抑制剂。二十[1,2,4]三唑[4,3- a对吡啶进行设计，合成和筛选，作为Smo抑制剂。这些新化合物中的四个显示与Smo蛋白直接结合，结合亲和力比VIS强。新化合物在体外对癌细胞系，尤其是三阴性乳腺癌细胞显示出广泛的抗增殖活性。机理研究表明，TPB15明显诱导MDA-MB-468细胞的细胞周期停滞和凋亡。TPB15阻止Smo易位到纤毛中，并降低Smo蛋白和mRNA表达。此外，下游调节因子神经胶质瘤相关癌基因1（Gli1）的表达受到明显抑制。最后，与我们的动物模型相比，TPB15具有更强的抗肿瘤活性VIS具有较低的毒性。因此，这些结果支持该新型支架的进一步优化以开发改良的Smo拮抗剂。