Inflammatory Activity of Epithelial Stem Cell Variants from Cystic Fibrosis Lungs Is Not Resolved by CFTR Modulators

Authors

Shan Wang
Suchan Niroula
Ashley Hoffman
Melika Khorrami
Melina Khorrami
Feng Yuan
Grace N Gasser
Soon Choi
Bovey Liu
Justin Li
Mark L Metersky
Matthew Vincent
Christopher P Crum
Richard C Boucher
Harry Karmouty-Quintana
Howard J Huang
Ajay Sheshadri
Burton F Dickey
Kalpaj R Parekh
John F Engelhardt
Frank D McKeon
Wa Xian

Publication Date

11-1-2023

Journal

American Journal of Respiratory and Critical Care Medicine

Abstract

Rationale

CFTR (cystic fibrosis transmembrane conductance regulator) modulator drugs restore function to mutant channels in patients with cystic fibrosis (CF) and lead to improvements in body mass index and lung function. Although it is anticipated that early childhood treatment with CFTR modulators will significantly delay or even prevent the onset of advanced lung disease, lung neutrophils and inflammatory cytokines remain high in patients with CF with established lung disease despite modulator therapy, underscoring the need to identify and ultimately target the sources of this inflammation in CF lungs.

Objectives

To determine whether CF lungs, like chronic obstructive pulmonary disease (COPD) lungs, harbor potentially pathogenic stem cell “variants” distinct from the normal p63/Krt5 lung stem cells devoted to alveolar fates, to identify specific variants that might contribute to the inflammatory state of CF lungs, and to assess the impact of CFTR genetic complementation or CFTR modulators on the inflammatory variants identified herein.

Methods

Stem cell cloning technology developed to resolve pathogenic stem cell heterogeneity in COPD and idiopathic pulmonary fibrosis lungs was applied to end-stage lungs of patients with CF (three homozygous CFTR:F508D, one CFTR F508D/L1254X; FEV1, 14–30%) undergoing therapeutic lung transplantation. Single-cell–derived clones corresponding to the six stem cell clusters resolved by single-cell RNA sequencing of these libraries were assessed by RNA sequencing and xenografting to monitor inflammation, fibrosis, and mucin secretion. The impact of CFTR activity on these variants after CFTR gene complementation or exposure to CFTR modulators was assessed by molecular and functional studies.

Measurements and Main Results

End-stage CF lungs display a stem cell heterogeneity marked by five predominant variants in addition to the normal lung stem cell, of which three are proinflammatory both at the level of gene expression and their ability to drive neutrophilic inflammation in xenografts in immunodeficient mice. The proinflammatory functions of these three variants were unallayed by genetic or pharmacological restoration of CFTR activity.

Conclusions

The emergence of three proinflammatory stem cell variants in CF lungs may contribute to the persistence of lung inflammation in patients with CF with advanced disease undergoing CFTR modulator therapy.

Keywords

cystic fibrosis, CFTR modulators, lung inflammation, lung stem cells, neutrophils.

Comments

This article has been corrected. See Am J Respir Crit Care Med. 2024 July 15; 210(2): 249.

PMID: 37695863