Ko Wooree, Porter Joseph J, Spelier Sacha, Sorensen Emily G, Bhatt Priyanka, Gabell Jeffrey T, van der Windt Isabelle, Couch Tyler, Coote Kevin, Mense Martin, Beekman Jeffrey M, Lueck John D
Department of Pharmacology and Physiology, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, United States.
Department of Pediatric Respiratory Medicine, Wilhelmina Children's Hospital, University Medical Center, Utrecht University, 3584 EA, Utrecht, The Netherlands.
Nucleic Acids Res. 2025 Jul 8;53(13). doi: 10.1093/nar/gkaf675.
Nonsense mutations arise from single nucleotide substitutions that result in premature termination codons (PTCs). PTCs result in little to no full-length protein production and decreased mRNA stability due to the nonsense-mediated mRNA decay (NMD) pathway. We provide evidence that anticodon-edited (ACE-) tRNAs efficiently suppress the most prevalent cystic fibrosis (CF)-causing PTCs, promoting significant rescue of endogenous cystic fibrosis transmembrane conductance regulator (CFTR) transcript abundance and channel function in different model systems. We show that our best-performing ACE-tRNA, which decodes all UGA PTCs to a leucine amino acid, markedly rescues CFTR function from the most prevalent CF-causing PTCs, all of which arose from nonleucine encoding codons. Using this single ACE-tRNA variant, we demonstrate significant rescue of CFTR function in an immortalized airway cell line and two different primary CF patient-derived intestinal cell models with CFTR nonsense mutations. Further, we demonstrate that leucine substitution CFTR variants are highly functional. Thus, ACE-tRNAs have promise as a platform therapeutic for CF and other nonsense-associated diseases.
无义突变源于单核苷酸替换，导致过早终止密码子（PTCs）的出现。由于无义介导的mRNA衰变（NMD）途径，PTCs导致几乎没有全长蛋白质产生，并且mRNA稳定性降低。我们提供的证据表明，反密码子编辑（ACE-）tRNA能有效抑制最常见的导致囊性纤维化（CF）的PTCs，在不同模型系统中显著促进内源性囊性纤维化跨膜传导调节因子（CFTR）转录本丰度和通道功能的恢复。我们表明，我们表现最佳的ACE-tRNA，它将所有UGA PTCs解码为亮氨酸氨基酸，能显著从最常见的导致CF的PTCs中恢复CFTR功能，所有这些PTCs均来自非亮氨酸编码密码子。使用这种单一的ACE-tRNA变体，我们在永生化气道细胞系以及两种不同的源自CF患者的原发性肠道细胞模型中证明了CFTR功能的显著恢复，这些模型均带有CFTR无义突变。此外，我们证明亮氨酸替代的CFTR变体具有高度功能性。因此，ACE-tRNAs有望成为治疗CF和其他无义相关疾病的平台疗法。
