(641) Building Towards the Future: The CFFT Lab as an expanding repository of primary cells with rare genotypes for CF research

Background: Since 2016, cell bank operations at the Cystic Fibrosis Foundation Therapeutics (CFFT) Lab have made steady progress in obtaining primary cells from cystic fibrosis (CF) patients with rare CFTR mutations. After quality control testing, basal cells isolated from lung and nasal airway epithelia, and intestinal cells from rectal biopsies are made available to the CF research community with the ultimate goal to identify agents that should be prioritized for further clinical development of potential treatments for patients with rare CFTR mutations.

Methods: All samples undergo rigorous quality control: sterility testing is performed to detect possible contamination (mycoplasma, other bacterial and fungal contamination), full CFTR gene sequencing is performed to confirm the CF genotype, and cell morphology along with growth (population doubling) is closely monitored. Standard pharmacological testing of CFTR function (electrophysiology) is performed on all fresh and out-of cryopreserved material. For several samples, we have also assessed CFTR mRNA (RT-qPCR), protein (Western blot), and CFTR function via Forskolin induced swelling (FIS) assay.

Results: To-date, our collection of cryopreserved, primary hBE cells comprises 5000 vials representing 39 genotypes from 64 donors including 7 codes with two rare CF alleles, and 29 codes with one rare CF allele and F508del. In addition, we banked 900 hNE vials comprised of 30 genotypes from 75 donors with the majority of vials containing hNE cells from people with cystic fibrosis (PwCF) with two CFTR nonsense mutations. The bank also stores over 500 vials with intestinal organoids (IOs) representing 19 genotypes from 22 donors with more than two thirds of the vials harboring IO crypts from PwCF with nonsense mutations in both CFTR alleles. Typically, primary hBE/hNE vials contain 1M cells/vial and IO vials contain on average 100-200 crypts.

Conclusions: Cell requests are initiated upon completion of the online Materials Transfer Request form at https://www.cff.org/material-transfer-request, including a brief description of the project. Requests will be granted if the internal review of the request finds that the intended research project is feasible, has scientific merit, and aligns with the CF Foundation’s mission to help people with CF to lead full and productive lives. Prior to submitting a formal cell request, we strongly recommend contacting Priyanka Bhatt (pbhatt@cff.org) or Jan Harrington (jharrington@cff.org) for cell availability and culture protocols.

Acknowledgements: We would like to thank Dr. Phil Karp (The University of Iowa) for kindly providing a substantial collection of hBE cells with rare genotypes.

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