(623) Preliminary assessment of nasal and colon-derived primary cells theratyping for a personalized medicine approach in CF patients carrying ultra-rare genotypes.

Background: about 15-20% of CF patients are not eligible for current CFTR modulators treatment. Most of these patients bear rare or ultra-rare mutations, making it difficult for them to be enrolled in clinical trials as well. Thus, new personalized approach to evaluate their responsiveness to CFTR modulators are needed. We used both rectal organoids (colonoids) and primary nasal brushed cells (hNEC) derived from CF patients with rare mutations to evaluate their responsiveness to CFTR modulators.

Methods: Colonoids were developed from rectal biopsies and CFTR function was measured by forskolin-induced swelling (FIS) assay (3D). Moreover, the CFTR function was also evaluated by short-circuit current (Isc) in 2 dimensional (2D) rectal organoids and nasal epithelial cells based on the availability. CFTR expression was analyzed by Western blotting. The effectiveness of CFTR modulator therapy in some patients was also reported. CFTR dependent sweat was tested by optical beta adrenergic test and chloride sweat concentration was determined following the Gibson and Cooke method.

Results: FIS and short-circuit current (Isc) measurements indicated the presence of a residual CFTR function. FIS detected a strong increase by treatment with the CFTR potentiator ivacaftor (VX-770, Vertex Pharmaceuticals) and the corrector tezacaftor (VX-661,

Vertex Pharmaceuticals). In the case carrying W57G/A234D variants the combination with elexacaftor(VX-445,Vertex Pharmaceuticals) increased the swelling, also in the absence of forskolin treatment. CFTR-mediated chloride secretion assay of colonoid monolayers indicated a residual function of 7% of wild-type (WT) CFTR, a relativeenhancement of the forskolin-induced, CFTR-specific current response to VX-661 of 20% of WT CFTR reaching 28% with VX-661/VX-445 and 32% with VX-661/VX-445/VX770 combination. Increased CFTR protein levels and function were detected in both intestinal organoids and nasal brushing cultures. Patienttreatment with CFTR modulators showed a considerable decrease of sweat chloride (i.e., meanly about 60 mmol/L as compared to baseline), relevant improvement of ppFEV1 (i.e., >8) and six-minute walk test, and an increase in body mass index after the first 8 weeks of treatment (W57G/A234D, 4382delA/R1066C, F508del/G970D, F508del/Unknown).

Conclusions: Clinical parameters we recoded confirms that the response predicted on both colon and nasal epithelial cells correlates well with in vivo therapeutic endpoints, confirming the efficacy of such models as predictors of clinical effectiveness of novel therapies in patients bearing rare ultra-rare mutations for a personalized medicine approach.

Acknowledgements: Italian Cystic Fibrosis Research Foundation (grants# 7/2016,13/2018 and 9/2020; LegaItaliana Fibrosi Cistica, Associazione Veneta-Onlus; Cystic Fibrosis Foundation grant# ASSAEL08A0

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