Biology subjects

Mense, M.

Publications and source records attributed to Mense, M..

4 recordsLinked to original sources

bioRxiv2024

Upregulation of a nonsense mediated decay (NMD) insensitive CFTR mRNA isoform has therapeutic potential for the treatment of 3 Prime CFTR PTC variants

BackgroundNonsense or Premature Termination Codon (PTC) mutations of the CFTR gene are pathogenic and found in [~]10% of North American people with cystic fibrosis. PTC mutations induce Nonsense-Mediated mRNA Decay (NMD), leading to a substantial ([~]80-90%) reduction in full-length mRNA. This reduction is a key contributor to PTC mutation-related pathology. Various approaches to evade NMD and preserve the impacted mRNA transcript have been explored but have not progressed to clinical development, leaving NMD a significant hurdle for PTC readthrough therapy. MethodsAntisense oligonucleotides (ASOs) were tiled across intron 22 splice donor (SD) and acceptor (SA) sites of the CFTR gene. Immortalized airway cells were treated with SD and SA ASOs, and those yielding the greatest increase in a CFTR NMD-insensitive mRNA isoform (e22 trunc mRNA) were tested in combinations. Top SD/SA ASO pairs were assessed for their impact on e22 trunc mRNA via ddPCR, e22 trunc protein via western blot, and CFTR-mediated chloride (Cl-) transport via transepithelial electrophysiological measurements in immortalized and primary human bronchial epithelial (hBE) cell cultures. ResultsWe demonstrate that e22 trunc mRNA generates a truncated CFTR protein whose Cl- transport function can be enhanced with elexacaftor/tezacaftor/ivacaftor (ETI) treatment. ASO and ETI treatment in combination restore [~]20% and 25% of wild-type CFTR Cl- transport function in immortalized epithelial and primary hBE cells homozygous for CFTR W1282X, respectively. ConclusionsThis study lays the groundwork for advancing ASO-mediated upregulation of e22 trunc mRNA and protein as a therapeutic approach for cystic fibrosis caused by 3-terminal CFTR PTC mutations.

genomics↗

bioRxiv2024

Anticodon Engineered Transfer RNAs (ACE-tRNAs) are a Platform Technology for Suppressing Nonsense Mutations

Nonsense mutations arise from single nucleotide substitutions that result in premature termination codons (PTCs). PTCs result in little to no full-length protein production and loss of mRNA expression through the nonsense-mediated mRNA decay (NMD) pathway. We demonstrate that anticodon engineered (ACE-) tRNAs efficiently suppress the most prevalent cystic fibrosis (CF) causing PTCs, promoting significant rescue of endogenous cystic fibrosis transmembrane conductance regulator (CFTR) transcript abundance and channel function in different model systems. We demonstrate that our best-performing ACE-tRNA, that decodes all UGA PTCs to a leucine amino acid, markedly rescues CFTR channel function from the most prevalent CF causing PTCs that arise from non-leucine encoding codons. Using this single ACE-tRNA variant, we demonstrate significant rescue of CFTR channel function in an immortalized airway cell line and two different primary CF patient-derived intestinal cell models with CFTR nonsense mutations. Thus, ACE-tRNAs have promise as a platform therapeutic for CF and other nonsense-associated diseases.

molecular biology↗

bioRxiv2023

In Vitro Modulator Responsiveness of 655 CFTR Variants Found in People With CF

BackgroundIn 2017, the US Food and Drug Administration initiated expansion of drug labels for the treatment of cystic fibrosis (CF) to include CF transmembrane conductance regulator (CFTR) gene variants based on in vitro functional studies. This study aims to identify CFTR variants that result in increased chloride (Cl-) transport function by the CFTR protein after treatment with the CFTR-modulator combination elexacaftor/tezacaftor/ivacaftor (ELX/TEZ/IVA). These data may benefit people with CF (pwCF) who are not currently eligible for modulator therapies. MethodsPlasmid DNA encoding 655 CFTR variants and wild-type (WT) CFTR were transfected into Fisher Rat Thyroid cells that do not natively express CFTR. After 24 hours of incubation with control or TEZ and ELX, and acute addition of IVA, CFTR function was assessed using the transepithelial current clamp conductance assay. Each variants baseline activity, responsiveness to IVA alone, and responsiveness to the TEZ/ELX/IVA combination were measured in three different laboratories. Western blots were conducted to evaluate CFTR protein maturation and complement the functional data. Results and Conclusions253 variants not currently approved for CFTR modulator therapy showed low baseline activity (<10% of normal CFTR Cl- transport activity). For 152 of these variants, treatment with ELX/TEZ/IVA improved the Cl- transport activity by [≥]10% of normal CFTR function, which is suggestive of clinical benefit. ELX/TEZ/IVA increased CFTR function by [≥]10 percentage points for an additional 140 unapproved variants with [≥]10% but <50% of normal CFTR function at baseline. These findings significantly expand the number of rare CFTR variants for which ELX/TEZ/IVA treatment should result in clinical benefit.

pharmacology and toxicology↗

bioRxiv2020

A molecular atlas of proximal airway identifies subsets of known airway cell types revealing details of the unique molecular pathogenesis of Cystic Fibrosis

Introduction/AbstractCystic fibrosis (CF) is a lethal autosomal recessive disorder that afflicts in excess of 70,000 people globally. People with CF experience multi-organ dysfunction resulting from aberrant electrolyte transport across polarized epithelia due to mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. CF-related lung disease is by far the most significant determinant of morbidity and mortality. In this study we report results from a multi-institute consortium in which single cell transcriptomics were applied to define disease-related changes to the proximal airway of CF donors (n=19) undergoing transplantation for end-stage lung disease compared to the proximal airway of previously healthy lung donors (n=19). We found that all major airway epithelial cell types were conserved between control and CF donors. Disease-dependent differences were observed, including an overabundance of epithelial cells transitioning to specialized ciliated and secretory cell subtypes coupled with an unexpected decrease in cycling basal cells. This study developed a molecular atlas of the proximal airway epithelium that will provide insights for the development of new targeted therapies for CF airway disease.

cell biology↗