Mutations in the gene encoding BMPR2 (bone morphogenetic protein type 2 receptor) are the major cause of heritable pulmonary arterial hypertension (PAH). Point mutations in the BMPR2 ligand-binding domain involving cysteine residues (such as C118W) are causative of PAH and predicted to cause protein misfolding. Using heterologous overexpression systems, we showed previously that these mutations lead to retention of BMPR2 in the endoplasmic reticulum but are partially rescued by chemical chaperones. Here, we sought to determine whether the chemical chaperone 4-phenylbutyrate (4PBA) restores BMPR2 signaling in primary cells and in a knockin mouse harboring a C118W mutation. First, we confirmed dysfunctional BMP signaling in dermal fibroblasts isolated from a family with PAH segregating the BMPR2 C118W mutation. After BMP4 treatment, the induction of downstream signaling targets (Smad1/5, ID1 [inhibitor of DNA binding 1], and ID2) was significantly reduced in C118W mutant cells. Treatment with 4PBA significantly rescued Smad1/5, ID1, and ID2 expression. Pulmonary artery smooth muscle cells isolated from the lungs of heterozygous mice harboring the Bmpr2 C118W mutation exhibited significantly increased proliferation. In the presence of 4PBA, hyperproliferation was dramatically reduced. Furthermore, in vivo, 4PBA treatment of Bmpr2 C118W mice partially rescued Bmpr2 expression, restored downstream signaling, and improved vascular remodeling. These findings demonstrate in primary cells and in a knockin mouse that the repurposed small-molecule chemical chaperone 4PBA might be a promising precision medicine approach to treat PAH in patients with specific subtypes of BMPR2 mutation involving cysteine substitutions in the ligand-binding domain.

编码 BMPR2（骨形态发生蛋白 2 型受体）的基因突变是遗传性肺动脉高压 (PAH) 的主要原因。BMPR2 配体结合域中涉及半胱氨酸残基（如 C118W）的点突变是 PAH 的致病因素，预计会导致蛋白质错误折叠。使用异源过表达系统，我们之前表明这些突变导致 BMPR2 保留在内质网中，但部分被化学伴侣拯救。在这里，我们试图确定化学伴侣 4-苯基丁酸酯 (4PBA) 是否能恢复原代细胞和携带 C118W 突变的敲入小鼠中的 BMPR2 信号。首先，我们证实了从 PAH 分离BMPR2的家族中分离出的真皮成纤维细胞中功能失调的 BMP 信号C118W突变。BMP4 处理后，下游信号转导目标（Smad1/5、ID1 [DNA 结合抑制剂 1] 和 ID2）的诱导在 C118W 突变细胞中显着减少。用 4PBA 治疗显着挽救了 Smad1/5、ID1 和 ID2 的表达。从携带Bmpr2 C118W 突变的杂合小鼠肺部分离的肺动脉平滑肌细胞表现出显着增加的增殖。在 4PBA 的存在下，过度增殖显着减少。此外，在体内，4PBA 处理Bmpr2C118W 小鼠部分挽救了 Bmpr2 的表达，恢复了下游信号，并改善了血管重塑。这些发现在原代细胞和敲入小鼠中表明，改变用途的小分子化学伴侣 4PBA 可能是一种有前途的精准医学方法，用于治疗BMPR2突变特定亚型患者的 PAH，涉及配体结合域中的半胱氨酸取代。