Small-molecule eRF3a degraders rescue CFTR nonsense mutations by promoting premature termination codon readthrough.
Small-molecule eRF3a degraders rescue CFTR nonsense mutations by promoting premature termination codon readthrough.
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DOI：
10.1172/jci154571
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发表时间：
2022-09-15
影响因子：
15.9
通讯作者：
Randell, Scott H.
中科院分区：
文献类型：
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作者：
Lee, Rhianna E.;Lewis, Catherine A.;He, Lihua;Bulik-Sullivan, Emily C.;Gallant, Samuel C.;Mascenik, Teresa M.;Dang, Hong;Cholon, Deborah M.;Gentzsch, Martina;Morton, Lisa C.;Minges, John T.;Theile, Jonathan W.;Castle, Neil A.;Knowles, Michael R.;Kimple, Adam J.;Randell, Scott H.
绝大多数囊性纤维化（CF）患者现在有资格接受CF跨膜调节因子（CFTR）调节剂治疗。剩余的CF个体携带过早终止密码子（PTC）或罕见的CFTR变体，治疗选择有限。虽然使用原代气道上皮细胞可以可靠地预测临床调节剂反应，但携带罕见CFTR变体的原代细胞很少。为了克服这一障碍，可以通过小鼠Bmi-1和人TERT（hTERT）的过表达来产生细胞系。使用这种方法，我们开发了2个非CF和6个CF气道上皮细胞系，其中3个是纯合的W1282 X PTC变体。Bmi-1/hTERT细胞系重现了原代细胞形态和离子转运功能。2个F508 del-CFTR细胞系对CFTR调节剂有强烈的反应，这反映在亲本原代细胞和细胞供体的临床反应中。靶向真核释放因子3a（eRF 3a）的Cereblon E3连接酶调节剂将W1282 X-CFTR功能恢复至WT水平的约20%，当与G418配对时，将G542 X-CFTR功能恢复至WT水平的约50%。有趣的是，eRF 3a降解剂也降低了上皮钠通道（ENaC）的功能。这些研究表明，Bmi-1/hTERT细胞系忠实地反映了对CFTR调节剂的原代细胞应答，并说明了拯救CFTR无义突变的治疗方法。
The vast majority of people with cystic fibrosis (CF) are now eligible for CF transmembrane regulator (CFTR) modulator therapy. The remaining individuals with CF harbor premature termination codons (PTCs) or rare CFTR variants with limited treatment options. Although the clinical modulator response can be reliably predicted using primary airway epithelial cells, primary cells carrying rare CFTR variants are scarce. To overcome this obstacle, cell lines can be created by overexpression of mouse Bmi-1 and human TERT (hTERT). Using this approach, we developed 2 non-CF and 6 CF airway epithelial cell lines, 3 of which were homozygous for the W1282X PTC variant. The Bmi-1/hTERT cell lines recapitulated primary cell morphology and ion transport function. The 2 F508del-CFTR cell lines responded robustly to CFTR modulators, which was mirrored in the parent primary cells and in the cell donors’ clinical response. Cereblon E3 ligase modulators targeting eukaryotic release factor 3a (eRF3a) rescued W1282X-CFTR function to approximately 20% of WT levels and, when paired with G418, rescued G542X-CFTR function to approximately 50% of WT levels. Intriguingly, eRF3a degraders also diminished epithelial sodium channel (ENaC) function. These studies demonstrate that Bmi-1/hTERT cell lines faithfully mirrored primary cell responses to CFTR modulators and illustrate a therapeutic approach to rescue CFTR nonsense mutations.