In silico analysis and theratyping of an ultra-rare CFTR genotype (W57G/A234D) in primary human rectal and nasal epithelial cells

  • iScience. 2023 Oct 12;26(11):108180. doi: 10.1016/j.isci.2023.108180.
Karina Kleinfelder  1 ,  Virginia Lotti  1 ,  Adriana Eramo  2 ,  Felice Amato  3  4 ,  Stefania Lo Cicero  2 ,  Germana Castelli  2 ,  Francesca Spadaro  5 ,  Alessia Farinazzo  1 ,  Daniele Dell'Orco  6 ,  Sara Preato  1 ,  Jessica Conti  1 ,  Luca Rodella  7 ,  Francesco Tomba  7 ,  Angelo Cerofolini  7 ,  Elena Baldisseri  8 ,  Marina Bertini  8 ,  Sonia Volpi  8 ,  Valeria Rachela Villella  3  4 ,  Speranza Esposito  3  4 ,  Immacolata Zollo  3  4 ,  Giuseppe Castaldo  3  4 ,  Carlo Laudanna  1 ,  Eric J Sorsher  9 ,  Jeong Hong  9 ,  Disha Joshi  9 ,  Garry Cutting  10 ,  Marco Lucarelli  11  12 ,  Paola Melotti  8 ,  Claudio Sorio  1
Affiliations
  • 1. Department of Medicine, University of Verona, Division of General Pathology, 37134 Verona, Italy.
  • 2. Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, 00161 Rome, Italy.
  • 3. Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, 80131 Naples, Italy.
  • 4. CEINGE-Biotecnologie Avanzate Franco Salvatore S.c.a.r.l., 80145 Naples, Italy.
  • 5. Confocal Microscopy Unit, Core Facilities, Istituto Superiore di Sanità, 00161 Rome, Italy.
  • 6. Department of Neurosciences, Biomedicine and Movement Sciences, Section of Biological Chemistry, University of Verona, 37134 Verona, Italy.
  • 7. Endoscopic Surgery Unit, Azienda Ospedaliera Universitaria Integrata Verona, 37126 Verona, Italy.
  • 8. Cystic Fibrosis Centre, Azienda Ospedaliera Universitaria Integrata Verona, 37126 Verona, Italy.
  • 9. Department of Pediatrics, Division of Pulmonary, Allergy/Immunology, Cystic Fibrosis & Sleep, Emory University, Atlanta, GA 30322, USA.
  • 10. McKusick-Nathans Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA.
  • 11. Department of Experimental Medicine, Sapienza University of Rome, 00185 Rome, Italy.
  • 12. Pasteur Institute, Cenci Bolognetti Foundation, Sapienza University of Rome, 00161 Rome, Italy.
PMID: 38026150 DOI: 10.1016/j.isci.2023.108180
Abstract

Mutation targeted therapy in cystic fibrosis (CF) is still not eligible for all CF subjects, especially for cases carrying rare variants such as the CFTR genotype W57G/A234D (c.169T>G/c.701C>A). We performed in silico analysis of the effects of these variants on protein stability, which we functionally characterized using colonoids and reprogrammed nasal epithelial cells. The effect of mutations on cystic fibrosis transmembrane conductance regulator (CFTR) protein was analyzed by western blotting, forskolin-induced swelling (FIS), and Ussing chamber analysis. We detected a residual CFTR function that increases following treatment with the CFTR modulators VX661±VX445±VX770, correlates among models, and is associated with increased CFTR protein levels following treatment with CFTR correctors. In vivo treatment with VX770 reduced sweat chloride concentration to non-CF levels, increased the number of CFTR-dependent sweat droplets, and induced a 6% absolute increase in predicted FEV1% after 27 weeks of treatment indicating the relevance of theratyping with patient-derived cells in CF.

Keywords
Human specimen; In silico biology; Integrative aspects of cell biology; Molecular medicine; Pharmacoinformatics; Therapy.
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