Nanobody-Driven Stabilization Synergistically Rescues F508del-CFTR and Reveals an Alternative Active State of the Channel

Defects in protein trafficking underlie many genetic diseases, including cystic fibrosis (CF), where the common F508del mutation destabilizes the cystic fibrosis transmembrane conductance regulator (CFTR) channel, leading to its degradation. To enhance current CFTR modulator therapies, we used lipid nanoparticles to deliver mRNA encoding T2a, a nanobody that thermally stabilizes CFTR by binding nucleotide-binding domain 1 (NBD1). When combined with clinically-approved correctors, T2a significantly improved F508del-CFTR maturation, plasma membrane expression, and channel activity. Single-channel recording revealed that nanobody binding sustained channel activity by promoting both full and sub-conductance gating states and protecting F508del-CFTR against thermal deactivation. Cryo-EM analysis identified a novel conformation of CFTR where NBD1 adopts an alternative geometry enabling pore formation in the absence of NBD dimerization. Our findings establish a new paradigm to correct protein trafficking by stabilizing misfolded domains with targeted nanobodies and demonstrate a broadly applicable framework to treat CF and related protein misfolding diseases.