Human Nasal Epithelial Organoids for Therapeutic Development in Cystic Fibrosis

Abstract

We describe a human nasal epithelial (HNE) organoid model derived directly from patient samples that is well-differentiated and recapitulates the airway epithelium, including the expression of cilia, mucins, tight junctions, the cystic fibrosis transmembrane conductance regulator (CFTR), and ionocytes. This model requires few cells compared to airway epithelial monolayer cultures, with multiple outcome measurements depending on the application. A novel feature of the model is the predictive capacity of lumen formation, a marker of baseline CFTR function that correlates with short-circuit current activation of CFTR in monolayers and discriminates the cystic fibrosis (CF) phenotype from non-CF. Our HNE organoid model is amenable to automated measurements of forskolin-induced swelling (FIS), which distinguishes levels of CFTR activity. While the apical side is not easily accessible, RNA- and DNA-based therapies intended for systemic administration could be evaluated in vitro, or it could be used as an ex vivo biomarker of successful repair of a mutant gene. In conclusion, this highly differentiated airway epithelial model could serve as a surrogate biomarker to assess correction of the mutant gene in CF or other diseases, recapitulating the phenotypic and genotypic diversity of the population.

Modifying and Integrating in vitro and ex vivo Respiratory Models for Inhalation Drug Screening

“…Respiratory organoids are 3D tissue-engineered culture systems capable of mimicking essential structural aspects of airways to screen drug pharmaceutical safety and efficacy (Jung et al, 2019;Liu et al, 2020). Unlike ALI monoculture models, respiratory organoids physiologically represent the 3D respiratory airway lumen microenvironment by promoting the growth and differentiation of multiple cell types to mimic the diverse structural branching present in airways (Barkauskas et al, 2017;van der Vaart and Clevers, 2020).…”

“…Therapeutic compounds are microinjected directly into the lumen of the organoid (Hill et al, 2017), organoids are integrated within microfluidic devices (via drug diffusion in solution) (Jung et al, 2019) or implanted within in vivo models (patient-derived xenografts) ( Figure 1B; Tan et al, 2017;Berkers et al, 2019;Kim et al, 2019;Takahashi et al, 2019) to provide a physiologically relevant in vitro drug screening platform. Respiratory organoids have been used to model the specific characteristics and physiological properties numerous respiratory diseases including to cystic fibrosis (Berkers et al, 2019;Dekkers et al, 2013;Liu et al, 2020), fibrosis (Strikoudis et al, 2019), viral and bacterial infections (Chen et al, 2017;Hill et al, 2017;Paolicelli et al, 2019), and lung cancer (Jung et al, 2019;Kim et al, 2019;Takahashi et al, 2019). The delivery of novel preclinical drug compounds to organoid models of respiratory diseases to determine therapeutic efficacy and cytotoxicity has been achieved in a variety of experimentally diverse setups.…”

Current strategies and opportunities to manufacture cells for modeling human lungs

“…CF is an autosomal recessive genetic disorder caused by mutations in the epithelial chloride channel gene, CFTR, which consequently leads to accumulation of excess mucous and compromised mucociliary clearance [ 145 ], affecting multiple organs. CF phenotypes have been studied widely in primary or PSC-derived intestinal [ [146] , [147] , [148] ], rectal [ [149] , [150] , [151] ], pancreatic [ 152 ], and airway models [ 11 , 27 , 33 , 153 , 154 ].…”

Nasal Epithelial Cell-Based Models for Individualized Study in Cystic Fibrosis

“…More recently, investigators have adapted methodology utilized for intestinal organoid studies in CF to nasal spheroids. In this approach, expanded HNEs are embedded in a three-dimensional matrix (typically Matrigel, Corning, NY, USA), which supports the spherical organization of the cells without the need for mechanical agitation [ 30 , 57 , 78 , 84 ]. Much like previous spheroid models, these cultures recapitulate the mature respiratory epithelium, including ciliation, mucus production, and formation of cell-to-cell junctions [ 57 , 78 ].…”

“…By altering the matrix density, cultures can be generated with the epithelial cell apex to the inside/lumen of the spheroid or to the outside [ 57 , 78 ]. To quantify CFTR function, ion transport is stimulated through cAMP-dependent pathways and spheroid size is monitored over time [ 30 , 57 , 78 , 84 ]. Using this approach, several groups have reported the capacity of these spheroids to discriminate CF cultures from non-CF, and to demonstrate pharmacologic rescue of CFTR using modulator compounds [ 57 , 78 , 84 ].…”