Chemically modified hCFTR mRNAs recuperate lung function in a mouse model of cystic fibrosis

Abstract

Gene therapy has always been a promising therapeutic approach for Cystic Fibrosis (CF). However, numerous trials using DNA or viral vectors encoding the correct protein resulted in a general low efficacy. In the last years, chemically modified messenger RNA (cmRNA) has been proven to be a highly potent, pulmonary drug. Consequently, we first explored the expression, function and immunogenicity of human (h)CFTR encoded by cmRNAhCFTR in vitro and ex vivo, quantified the expression by flow cytometry, determined its function using a YFP based assay and checked the immune response in human whole blood. Similarly, we examined the function of cmRNAhCFTR in vivo after intratracheal (i.t.) or intravenous (i.v.) injection of the assembled cmRNAhCFTR together with Chitosan-coated PLGA (poly-D, L-lactide-co-glycolide 75:25 (Resomer RG 752 H)) nanoparticles (NPs) by FlexiVent. The amount of expression of human hCFTR encoded by cmRNAhCFTR was quantified by hCFTR ELISA, and cmRNAhCFTR values were assessed by RT-qPCR. Thereby, we observed a significant improvement of lung function, especially in regards to FEV0.1, suggesting NP-cmRNAhCFTR as promising therapeutic option for CF patients independent of their CFTR genotype.

Recent Developments in mRNA-Based Protein Supplementation Therapy to Target Lung Diseases

“…A significant decrease in chloride concentration (around 50%) was also observed, indicating a restoration of CFTR in the duct compartment of submucosal glands and thus leading to improved chloride absorption. 20 A separate study from Robinson et al 181 confirmed nasal application of chemically modified CFTR mRNA can recover up to 55% of the net chloride efflux characteristic of healthy mice. Bangel-Ruland et al 182 demonstrate restoration of cAMP-induced CFTR current following transfection of CFBE41ocells with wild-type CFTR-mRNA similar to the values seen in 16HBE14o-control cells.…”

“…178 Similarly, DNA-based vectors (viral and plasmid) were tested by Alton's group (pGM169/GL67A), reaching phase II clinical trials with modest improvement in FEV 1 after repeated administration but no improvement in patient's quality of life. 179,180 Haque et al 20 have observed a significant improvement in CFTR protein translation, expression, and function in vitro (CFBE41o-and 16HBE14o-) and in vivo (CFTR-knockout mice) by administering chemically modified human CFTR (hCFTR) mRNAs complexed with chitosan-coated poly(lactic-co-glycolic acid) (PLGA) nanoparticles. The study also showed a substantial improvement in FEV 0.1 up to 89% of the level of a healthy control group.…”

“…TLR3, TLR7, TLR8, and RIG-I activations were significantly reduced when mRNA contained modified nucleosides such as m5C, m6A, 5-methyluridine (m5U), J-UTP, and 2-thiouridine (s2U). 20,150,230,234 RNAdependent protein kinase (PKR) arbitrated the immune response, and translation inhibition (by phosphorylating the alpha subunit of translation initiation factor 2 [eIF-2a]) can be escaped using J-UTPor m5C-modified nucleosides. 235 m5C is explicitly recognized by the mRNA export adaptor ALYREF, and it increases mRNA-binding affinity and associated mRNA export.…”

“…248 This increases the amount of mRNA needed for reaching effective dose levels in the lung. 20 Another hurdle emerges in the lung itself. The lung has a capillary system that consists of mostly small and non-fenestrated capillaries.…”

“…The recent advancements in the field of nanocarriers suggest the possibility to customize particles for target organs. 20,21 Based on our research interest in lungs, this review focuses on protein replacement therapy of lung disease, especially monogenetic diseases, such as cystic fibrosis (CF) and surfactant protein B (SP-B) deficiency, as well as multifactorial diseases, such as chronic obstructive pulmonary disease (COPD) and asthma. To achieve targeted therapy options, lung structure and methods to reach specific lung cell populations are critical.…”