Therapeutic approaches for people with Cystic Fibrosis carrying rare variants

The Cystic Fibrosis (CF, OMIM# 219700) is one of the most frequent and serious genetic diseases in the Caucasian population, due to variants in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR, OMIM# 602421) gene (Riordan JR et al., 1989) that cause loss of function of the CFTR protein, a chloride channel localized in the apical membrane of the epithelial cells (Lukacs GL and Verkman AS., 2012). The causative CF variants were grouped into seven classes according to the functional defect they cause (Bareil Cand Bergougnoux A, 2020). Small molecules called correctors and potentiators (CFTR modulators) have been developed to rescue the basic functional defect(s) of different variants (Hudock KM and Clancy JP,2017). Nevertheless, there are limitations that cannot be underestimated: individuals with CF carrying different variants belonging to the same class may not respond in the same way to a single pharmacological agent, many individuals with CF have rare or poorly characterized variants whose response to pharmacological treatments remains unknown (in Italy about 30% of CF people) and finally, CF individuals with the same genotype may respond differently to treatments due to their genetic background. In recent years, precision medicine has been fundamental in overcoming these problems using different primary cellular models as human bronchial epithelial cells (HBEC) or intestinal organoids each with specific limitations and strengths. HBEC are obtained by invasive procedures as bronchoscopy or lung transplantation and cannot be obtained from every desired CF individual. In contrast, intestinal organoids can be easily obtained from patients and expanded to very large numbers. Still, there is the possibility that some features of CFTR variants expressed in the context of the airway epithelium are not well represented. These observations demonstrate the need to find a personalized approach to treat CF basic defect relying on robust and highly predictive assays for the testing of therapies in CF. Human nasal epithelial cells (HNEC) may be considered an excellent predictive model given that they reproduce the functional properties of the respiratory epithelium when properly expanded and differentiated in vitro. In addition, they can be obtained by a minimally invasive procedure (nasal brushing), meaning that can be virtually applied to all individuals with CF, with a good representation of all genotypes, comprising those with rare alleles (Sondo E et al., 2022; Tomati V et al., 2022). The work of my PhD has stemmed from a wide “theratyping” project which has seen the enrollment of several Italian CF individuals aimed at evaluating the effect of available and novel CFTR modulators on orphan variants and at characterizing the mechanism of action of such variants. In this context, the work of my PhD led to the identification of individuals with CF that are likely to have a benefit from the treatment with available modulators. Moreover, we focused on the study of a selection of individuals with CF for which an unexpected response or insensitivity to CFTR modulators were reported, prompting us to investigate in detail the underlying mechanisms. To do this, we collected HNECs from all recruited CF individuals as well developed a series of robust functional, molecular and biochemical assays for define the patient's therapy and obtain a deep characterization of the specific CFTR variants. Our results demonstrate the high translational potential of the presented strategy based on the use of a reliable ex vivo model, and of a robust translational workflow integrating diagnostics and functional assays with high predictive value of the clinical efficacy of drugs and pharmacological agents investigated by these means.