Pharmacological Modulation of BMAL1 Alters Circadian Dynamics and the Macrophage Immune Response

The master transcription factor BMAL1-CLOCK is crucial for orchestrating rhythmic gene expression and governing circadian rhythms. Pharmacological manipulation of this central circadian regulator requires accessible protein cavities and selective compounds. By targeting BMAL1 through a resident cavity in its PAS-B domain, we demonstrate that the small-molecule Core Circadian Modulator (CCM) penetrates and substantially expands this pocket. Biochemical and cellular analyses validate CCM's target engagement selectivity, enabling direct access to BMAL1's transcriptional activities. CCM induces dose-dependent changes in PER2-Luc macrophage oscillations and selectively modifies expression patterns of circadian controlled genes. In activated macrophages, CCM modulates BMAL1-CLOCK controlled inflammatory and phagocytic pathways to promote their downregulation. Our findings demonstrate the feasibility of directly targeting BMAL1-CLOCK as a strategy for circadian regulation and targeting circadian-regulated processes To investigate the specificity and effects of pharmacological manipulation of Bmal1, bone-marrow derived macrophages from both myeloid cell specific Bmal1-KO mice and littermate controls were treated with CCM. The effect of inflammation was monitored: this was done under both basal and stimulated conditions.

主转录因子BMAL1-CLOCK在协调节律性基因表达、调控昼夜节律过程中发挥关键作用。对这一核心昼夜节律调控因子实施药理学操控，需依赖具备可及性的蛋白质空腔与高选择性小分子化合物。本研究通过靶向BMAL1的PAS-B结构域内的固有空腔，证实小分子核心昼夜节律调控剂（Core Circadian Modulator，CCM）可结合该空腔并使其显著扩张。生化与细胞实验验证了CCM的靶点结合选择性，使其能够直接调控BMAL1的转录活性。CCM可诱导携带PER2-Luc荧光素酶报告基因的巨噬细胞振荡呈现剂量依赖性变化，并选择性改变昼夜节律调控基因的表达模式。在活化巨噬细胞中，CCM可调控BMAL1-CLOCK介导的炎症与吞噬通路，促进其表达下调。本研究结果证实，直接靶向BMAL1-CLOCK可作为昼夜节律调控以及靶向昼夜节律调控过程的可行策略。为探究BMAL1药理学操控的特异性与效应，研究人员对髓系细胞特异性BMAL1基因敲除（Bmal1-KO）小鼠及其同窝对照小鼠的骨髓来源巨噬细胞施以CCM处理，并在基础状态与炎症刺激条件下监测炎症相关效应。