PT  - JOURNAL ARTICLE
AU  - John D. Lueck
AU  - Daniel T. Infield
AU  - Adam L. Mackey
AU  - R. Marshall Pope
AU  - Paul B. McCray
AU  - Christopher A. Ahern
TI  - Engineered tRNA suppression of a CFTR nonsense mutation
AID  - 10.1101/088690
DP  - 2016 Jan 01
TA  - bioRxiv
PG  - 088690
4099  - http://biorxiv.org/content/early/2016/11/20/088690.short
4100  - http://biorxiv.org/content/early/2016/11/20/088690.full
AB  - Ten percent of human diseases are caused by ‘nonsense’ mutations that lead to premature truncation of the protein reading frame. Small molecules that promote read-through of such PTC have significant clinical promise but current iterations suffer from low in vivo efficacy and the nonselective amino acid incorporation. Alternatively, while gene-modifying approaches, such as CRISPR/Cas9, represent a long-term solution, such treatments may be far from reaching the clinical setting. Building on previous work by our group and others, we describe a tRNA engineering approach that enables the conversion of an in frame nonsense stop mutation to the naturally occurring amino acid, thus rescuing the full-length wild type protein. Data is presented demonstrating the functionality of the approach with the rescue of CFTR W1282X, a human mutation that causes cystic fibrosis (CF). The stringency of the approach is confirmed by mass spectrometry in a model protein indicating the encoding of only tryptophan at the TGA suppression site. The data describe the first use of an edited tRNA to repair a CF causative mutation and serve a proof of principle for the eventual use of codon-edited tRNA for the therapeutic rescue of PTC disease codons.