The Hedgehog/Glioma-associated oncogene (Hh/Gli) signaling pathway plays an essential role in embryonic development and tissue homeostasis. Aberrant regulation of this pathway has been linked to various human malignancies. Gli1, the downstream transcription factor of the Hh pathway, is the ultimate effector of the canonical Hh pathway and has been identified as a common regulator of several tumorigenic pathways prevalent in Hh-independent cancers. Thus Gli1 represents a unique and promising drug target for a wide range of cancers. However, the identification and development of small molecules that directly target Gli1 protein have progressed slowly, due to an insufficient efficacy and selectivity. Herein, we developed novel small-molecule Gli1 degraders based on the hydrophobic tagging (HyT) strategy. The Gli1 HyT degrader 8e potently inhibited the proliferation of Gli1-overexpressed HT29 colorectal cancer cells, induced Gli1 degradation with a DC₅₀ value of 5.4 μM in HT29 and achieved 70% degradation at 7.5 μM in MEF^PTCH1−/− and MEF^SUFU−/−cell lines, via proteasome pathway. Compared to the canonical Hh antagonist Vismodegib, 8e exhibited much stronger potency in suppressing the mRNA expression of Hh target genes in Hh-overactivated MEF^PTCH1−/− and Vismodegib resistant MEF^SUFU−/− cells. Our study provides small molecule Gli1 degraders effectively interfering with both canonical and noncanonical Hh signaling and overcoming current Smoothened (SMO) antagonists resistance, which might pave a new avenue for developing therapeutic modalities targeting Hh/Gli1 signaling pathway.

Hedgehog/胶质瘤相关癌基因 (Hh/Gli) 信号通路在胚胎发育和组织稳态中发挥重要作用。该通路的异常调节与多种人类恶性肿瘤有关。 Gli1 是 Hh 通路的下游转录因子，是经典 Hh 通路的最终效应子，并已被确定为 Hh 独立癌症中常见的几种致瘤通路的共同调节因子。因此，Gli1 代表了针对多种癌症的独特且有前途的药物靶点。然而，由于功效和选择性不足，直接靶向Gli1蛋白的小分子的鉴定和开发进展缓慢。在此，我们开发了基于疏水标记（HyT）策略的新型小分子 Gli1 降解剂。 Gli1 HyT 降解剂8e有效抑制 Gli1 过表达的 HT29 结直肠癌细胞的增殖，在 HT29 中以 5.4 μM 的 DC ₅₀值诱导 Gli1 降解，并在 MEF ^PTCH1−/−和 MEF ^SUFU−/中以 7.5 μM 的浓度实现 70% 的降解。 ⁻细胞系，通过蛋白酶体途径。与经典的 Hh 拮抗剂 Vismodegib 相比， 8e在抑制 Hh 过度激活的 MEF ^PTCH1−/−和 Vismodegib 耐药性 MEF ^SUFU−/−细胞中 Hh 靶基因 mRNA 表达方面表现出更强的效力。 我们的研究提供了小分子 Gli1 降解剂，可有效干扰经典和非经典 Hh 信号传导，并克服当前的 Smoothened (SMO) 拮抗剂耐药性，这可能为开发针对 Hh/Gli1 信号通路的治疗方式开辟新途径。