(670) TGF-β1 diminishes efficacy of highly effective CFTR modulators, an effect reversed by losartan

Background: The introduction of highly effective modulator therapies (HEMT) has dramatically improved the prognosis for those with cystic fibrosis (CF). The triple combination of elexacaftor, tezacaftor, and ivacaftor (ETI) has benefited many, but not all, of those with F508del, the most common mutation in CFTR. Heterogeneous responses to ETI suggests additional factors influence the efficacy of even highly effective CFTR modulators. Transforming growth factor-β1 (TGF-β1) is not only linked to worse pulmonary outcomes in CF but can also diminish the efficacy of first/second generation modulators. Losartan, used clinically as an angiotensin receptor blocker (ARB), exerts anti-TGF-β effects independent of its ARB property. In this study, we sought to determine whether TGF-β1 reduces the efficacy of ETI on functional F508del-CFTR recovery in primary CF bronchial epithelial (CFBE) cells and whether losartan could reverse these effects.

Methods: Homozygous F508del CFBE cells were re-differentiated at the air-liquid interface. Cells were treated with basolateral losartan (10 µM) every 2 days for at least 3 weeks. Cells were then treated for 24h with recombinant TGF-β1 (5 ng/mL) and ETI (1/5/1 µM) or vehicle control. Cells were mounted in Ussing chambers and CFTR-dependent short-circuit currents were recorded upon addition of CFTRinh-172 (10 µM) after ivacaftor (1 µM) and forskolin (10 µM) stimulation in the presence of amiloride (10 µM). Airway surface liquid (ASL) volume was estimated by meniscus scanning. CFTR, TNF-α, and COX-2 expression was measured by qPCR. TNF-α and active/total TGF-β1 levels were measured by ELISA. Nasal fluid from CF patients was collected by Leukosorb.

Results: Sweat chloride concentrations and poor lung function response to ETI were associated with elevated levels of TGF-β1 in the upper airway of CF patients. TGF-β1 caused a significant reduction in ETI-corrected F508del-CFTR function, thereby increasing ASL absorption rates and inducing mucus hyperconcentration in primary CFBE cells in vitro. TGF-β1 decreased CFTR mRNA expression but also induced expression of TNF-α and COX-2. Losartan and EXP3179, the losartan metabolite with no ARB activity, improved ETI-corrected F508del-CFTR function in the presence of TGF-β1. Losartan further reversed TGF-β1-induced increases in TNF-α and COX-2 expression. NS-398, a specific COX-2 inhibitor, rescued TGF-β1 inhibition of ETI-corrected F508del-CFTR function.

Conclusions: These data show that elevated levels of TGF-β1 in the upper airway correlate with higher sweat chloride concentrations and less improvement in lung function after starting ETI in those with CF. Our in vitro data further show that TGF-β1 impairs the function of ETI-corrected F508del-CFTR by reducing CFTR mRNA and by increasing expression of TNF-α and COX-2. Losartan and EXP3179 partially reverse the effects of TGF-β1 in ETI-corrected F508del CFBE cells in vitro. These data set the stage for clinical studies using losartan for those with CF on ETI but with poor clinical responses.

Acknowledgements: Supported by NIH (R01 HL133240 to MS) and CFF (BENGTA19CO to CB; SALATH14G0, SALATHE16G0 to MS).

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